96 well plates  (Thermo Fisher)


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    Name:
    96-Well Plates
    Description:
    These are polystyrene 96-well round-bottom plates suitable for use with Thermo Scientific BCA-RAC (reducing agent compatible) assays. Related Products Pierce Microplate BCA Protein Assay Kit - Reducing Agent Compatible Pierce BCA Protein Assay Kit - Reducing Agent Compatible
    Catalog Number:
    15045
    Price:
    None
    Applications:
    Protein Assays and Analysis|Protein Biology|Protein Quantitation
    Size:
    5 plates
    Category:
    Kits and Assays, Protein Analysis Kits, Protein Quantitation Assay Kits, Reagents & Standards
    Score:
    85
    Buy from Supplier


    Structured Review

    Thermo Fisher 96 well plates
    CD13 chemical inhibitors or antibodies significantly slow growth and migration of RA FLS in vitro . FLS were seeded on <t>96-well</t> plates overnight. An Essen Incucyte system was used for both growth ( A ) and migration ( B ) assays. Growth was calculated as the difference in percent confluence from time 0. Migration was measured in a scratch wound assay using relative wound density. All data is expressed as a ratio to untreated FLS of the same cell line. mean±SEM n≥4 (*p≤0.05)
    These are polystyrene 96-well round-bottom plates suitable for use with Thermo Scientific BCA-RAC (reducing agent compatible) assays. Related Products Pierce Microplate BCA Protein Assay Kit - Reducing Agent Compatible Pierce BCA Protein Assay Kit - Reducing Agent Compatible
    https://www.bioz.com/result/96 well plates/product/Thermo Fisher
    Average 78 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    96 well plates - by Bioz Stars, 2020-01
    78/100 stars

    Images

    1) Product Images from "Localization, Shedding, Regulation and Function of Aminopeptidase N/CD13 on Fibroblast like Synoviocytes"

    Article Title: Localization, Shedding, Regulation and Function of Aminopeptidase N/CD13 on Fibroblast like Synoviocytes

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0162008

    CD13 chemical inhibitors or antibodies significantly slow growth and migration of RA FLS in vitro . FLS were seeded on 96-well plates overnight. An Essen Incucyte system was used for both growth ( A ) and migration ( B ) assays. Growth was calculated as the difference in percent confluence from time 0. Migration was measured in a scratch wound assay using relative wound density. All data is expressed as a ratio to untreated FLS of the same cell line. mean±SEM n≥4 (*p≤0.05)
    Figure Legend Snippet: CD13 chemical inhibitors or antibodies significantly slow growth and migration of RA FLS in vitro . FLS were seeded on 96-well plates overnight. An Essen Incucyte system was used for both growth ( A ) and migration ( B ) assays. Growth was calculated as the difference in percent confluence from time 0. Migration was measured in a scratch wound assay using relative wound density. All data is expressed as a ratio to untreated FLS of the same cell line. mean±SEM n≥4 (*p≤0.05)

    Techniques Used: Migration, In Vitro, Scratch Wound Assay Assay

    2) Product Images from "MRI-Based Assessment of Intralesional Delivery of Bone Marrow-Derived Mesenchymal Stem Cells in a Model of Equine Tendonitis"

    Article Title: MRI-Based Assessment of Intralesional Delivery of Bone Marrow-Derived Mesenchymal Stem Cells in a Model of Equine Tendonitis

    Journal: Stem Cells International

    doi: 10.1155/2016/8610964

    MRI data from cell suspensions in agar gel showing (a) PD (top row) and TRUFI (bottom row) images ranging from cell-free gel (CFG) up to 1 × 10 6 cells in a 96-well plate. (b) Signal-to-noise ratios (SNR) corresponding to the TRUFI images in (a). (c) Normal tendon following serial injection of 20, 10, 5, and 1 × 10 6 SPIO-labeled MSCs in order along the palmar aspect of the limb. SPIO-associated signal is only visible in the tissues surrounding the SDFT (arrows) ( ∗∗∗∗ p
    Figure Legend Snippet: MRI data from cell suspensions in agar gel showing (a) PD (top row) and TRUFI (bottom row) images ranging from cell-free gel (CFG) up to 1 × 10 6 cells in a 96-well plate. (b) Signal-to-noise ratios (SNR) corresponding to the TRUFI images in (a). (c) Normal tendon following serial injection of 20, 10, 5, and 1 × 10 6 SPIO-labeled MSCs in order along the palmar aspect of the limb. SPIO-associated signal is only visible in the tissues surrounding the SDFT (arrows) ( ∗∗∗∗ p

    Techniques Used: Magnetic Resonance Imaging, Injection, Labeling

    3) Product Images from "Effects of Azithromycin, Metronidazole, Amoxicillin, and Metronidazole plus Amoxicillin on an In Vitro Polymicrobial Subgingival Biofilm Model"

    Article Title: Effects of Azithromycin, Metronidazole, Amoxicillin, and Metronidazole plus Amoxicillin on an In Vitro Polymicrobial Subgingival Biofilm Model

    Journal:

    doi: 10.1128/AAC.04974-14

    Matrix of minimum similarity coefficient values for the microbial profiles for 9 in vitro biofilms from three 96-well plates (plate I, plate II, and plate III) and for these in vitro biofilms versus mean values for in vivo biofilms from 178 subjects. Cells with thicker borders include minimum similarity coefficient values for comparisons among biofilms from the same plate.
    Figure Legend Snippet: Matrix of minimum similarity coefficient values for the microbial profiles for 9 in vitro biofilms from three 96-well plates (plate I, plate II, and plate III) and for these in vitro biofilms versus mean values for in vivo biofilms from 178 subjects. Cells with thicker borders include minimum similarity coefficient values for comparisons among biofilms from the same plate.

    Techniques Used: In Vitro, In Vivo

    4) Product Images from "A human monoclonal antibody against small envelope protein of hepatitis B virus with potent neutralization effect"

    Article Title: A human monoclonal antibody against small envelope protein of hepatitis B virus with potent neutralization effect

    Journal:

    doi: 10.1080/19420862.2015.1134409

    Neutralization of HBV infectivity in HepaRG cells (A) and HepG2/NTCP cells (B) by G12 or HBIG. Cells seeded in 96-well plates were incubated overnight with cell culture derived HBV particles in the absence or presence of serial 4-fold dilutions of G12
    Figure Legend Snippet: Neutralization of HBV infectivity in HepaRG cells (A) and HepG2/NTCP cells (B) by G12 or HBIG. Cells seeded in 96-well plates were incubated overnight with cell culture derived HBV particles in the absence or presence of serial 4-fold dilutions of G12

    Techniques Used: Neutralization, Infection, Incubation, Cell Culture, Derivative Assay

    Characterization of 21 human anti-S mAbs to identify those with high affinity for the S protein. (A) ELISA method. Purified S protein (red) or anti-human Fab antibody (blue) were coated on 96-well plates, and incubated with human Fab antibodies. Bound
    Figure Legend Snippet: Characterization of 21 human anti-S mAbs to identify those with high affinity for the S protein. (A) ELISA method. Purified S protein (red) or anti-human Fab antibody (blue) were coated on 96-well plates, and incubated with human Fab antibodies. Bound

    Techniques Used: Enzyme-linked Immunosorbent Assay, Purification, Incubation

    5) Product Images from "The functionalized amino acid (S)-Lacosamide subverts CRMP2-mediated tubulin polymerization to prevent constitutive and activity-dependent increase in neurite outgrowth"

    Article Title: The functionalized amino acid (S)-Lacosamide subverts CRMP2-mediated tubulin polymerization to prevent constitutive and activity-dependent increase in neurite outgrowth

    Journal: Frontiers in Cellular Neuroscience

    doi: 10.3389/fncel.2014.00196

    ( S )-LCM does not affect tubulin-CRMP2 binding. (A) 96-well plates coated with 200 ng tubulin were incubated with increasing concentrations of recombinant CRMP2 or heat-denatured CRMP2. The Y axis displays the OD 450 absorbance of the ELISA using CRMP2-specific antibodies. CRMP2 bound to tubulin with half-saturation concentration of ~607 nM. Binding of heat-denatured CRMP2 demonstrated non-saturable background binding to tubulin. (B) Competitive binding assay revealed that ( S )-LCM does not abrogate the binding of CRMP2 to tubulin. For (A,B) , all measurements were performed in sextuplicate and error bars indicate standard error of the mean. Most of the error bars are smaller than the symbols.
    Figure Legend Snippet: ( S )-LCM does not affect tubulin-CRMP2 binding. (A) 96-well plates coated with 200 ng tubulin were incubated with increasing concentrations of recombinant CRMP2 or heat-denatured CRMP2. The Y axis displays the OD 450 absorbance of the ELISA using CRMP2-specific antibodies. CRMP2 bound to tubulin with half-saturation concentration of ~607 nM. Binding of heat-denatured CRMP2 demonstrated non-saturable background binding to tubulin. (B) Competitive binding assay revealed that ( S )-LCM does not abrogate the binding of CRMP2 to tubulin. For (A,B) , all measurements were performed in sextuplicate and error bars indicate standard error of the mean. Most of the error bars are smaller than the symbols.

    Techniques Used: Laser Capture Microdissection, Binding Assay, Incubation, Recombinant, Enzyme-linked Immunosorbent Assay, Concentration Assay, Competitive Binding Assay

    6) Product Images from "Inhibition of the AIF/CypA complex protects against intrinsic death pathways induced by oxidative stress"

    Article Title: Inhibition of the AIF/CypA complex protects against intrinsic death pathways induced by oxidative stress

    Journal: Cell Death & Disease

    doi: 10.1038/cddis.2013.518

    CypA-siRNA and AIF(370–394) peptide block the oxidative stress-induced increase of intracellular Ca2+ concentration. ( a and b ) HT-22 cells were treated with 2 mM glutamate for 12–14 h. The intracellular concentration of Ca 2+ was assessed by the fluorescent Ca 2+ indicator Fluo-4 AM in 96-well plates. ( b ) Cell viability assessment of HT-22 cells received AIF(370–394) peptide at the indicated concentrations, followed by exposure to 2 mM glutamate for 12–14 h, either in a standard culture medium (black bars) or in a medium without Ca 2+ (gray bars) ( n =4, *** P
    Figure Legend Snippet: CypA-siRNA and AIF(370–394) peptide block the oxidative stress-induced increase of intracellular Ca2+ concentration. ( a and b ) HT-22 cells were treated with 2 mM glutamate for 12–14 h. The intracellular concentration of Ca 2+ was assessed by the fluorescent Ca 2+ indicator Fluo-4 AM in 96-well plates. ( b ) Cell viability assessment of HT-22 cells received AIF(370–394) peptide at the indicated concentrations, followed by exposure to 2 mM glutamate for 12–14 h, either in a standard culture medium (black bars) or in a medium without Ca 2+ (gray bars) ( n =4, *** P

    Techniques Used: Blocking Assay, Concentration Assay

    7) Product Images from "Anti-heparan Sulfate Peptides That Block Herpes Simplex Virus Infection in Vivo"

    Article Title: Anti-heparan Sulfate Peptides That Block Herpes Simplex Virus Infection in Vivo

    Journal:

    doi: 10.1074/jbc.M110.201103

    G2 also blocks entry of representative members of β- and γ-herpesvirus subfamilies (CMV and HHV-8). A , a monolayer of cultured RPE cells grown in a 96-well plate was pretreated with G1, G2, or Cp at 0.5 m m concentration. A mock-treated
    Figure Legend Snippet: G2 also blocks entry of representative members of β- and γ-herpesvirus subfamilies (CMV and HHV-8). A , a monolayer of cultured RPE cells grown in a 96-well plate was pretreated with G1, G2, or Cp at 0.5 m m concentration. A mock-treated

    Techniques Used: Cell Culture, Concentration Assay

    G1 and G2 peptides block HSV-1 entry into human target cells. HeLa cells ( panel A ) and primary cultures of human CFs ( panel B ) were tested. Cells in 96-well plates were pretreated for 60 min with indicated m m concentrations of G1, G2, or Cp peptides.
    Figure Legend Snippet: G1 and G2 peptides block HSV-1 entry into human target cells. HeLa cells ( panel A ) and primary cultures of human CFs ( panel B ) were tested. Cells in 96-well plates were pretreated for 60 min with indicated m m concentrations of G1, G2, or Cp peptides.

    Techniques Used: Blocking Assay

    8) Product Images from "Vibrio natriegens as Host for Expression of Multisubunit Membrane Protein Complexes"

    Article Title: Vibrio natriegens as Host for Expression of Multisubunit Membrane Protein Complexes

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2018.02537

    Mrp expression confers a growth advantage to V. natriegens at high [Li + ]. Cells were transformed with either the pVc-Mrp (closed circles) or with the pBAD-TOPO plasmid (empty circles) and grown aerobically in LBS in the presence of 0.2% L -arabinose and 400 mM LiCl at 37°C in a 96-well plate. Averages and standard deviations from three biological replicates are shown.
    Figure Legend Snippet: Mrp expression confers a growth advantage to V. natriegens at high [Li + ]. Cells were transformed with either the pVc-Mrp (closed circles) or with the pBAD-TOPO plasmid (empty circles) and grown aerobically in LBS in the presence of 0.2% L -arabinose and 400 mM LiCl at 37°C in a 96-well plate. Averages and standard deviations from three biological replicates are shown.

    Techniques Used: Expressing, Transformation Assay, Plasmid Preparation

    9) Product Images from "A synthetic lethal siRNA screen identifying genes mediating sensitivity to a PARP inhibitor"

    Article Title: A synthetic lethal siRNA screen identifying genes mediating sensitivity to a PARP inhibitor

    Journal:

    doi: 10.1038/emboj.2008.61

    PARP-inhibitor synthetic lethality screen with protein kinase siRNA library. ( A ) HTS method. CAL51 cells plated in 96-well plates were transfected with siRNA. Each transfection plate contained 80 experimental siRNAs (SMARTPools of four different siRNA targeting the same gene) supplemented with four wells of non-targeting siCON, and two wells of siRNA directed against BRCA1 (positive control). Transfected cells were divided into six replica plates, half treated with DMSO vehicle alone and half with PARP inhibitor KU0058948 at 1 μM, the SF80 of CAL51. Cell viability was assessed after 5 days of KU0058948 exposure using CellTiter-Glo Luminescent Cell Viability Assay (Promega). ( B ) Reproducibility of HTS method. Correlation of the effect of siRNA on cell growth in vehicle-treated plates from two replicates of the entire screen. Spearman correlation coefficient, r =0.83. ( C ) Correlation of KU0058948 sensitivity Z -scores from two replicates of the entire screen. r =0.54. ( D ) Scatter plot of averaged Z -scores from PARP inhibitor sensitivity screen carried out in duplicate with KU0058948. Red line indicates −3 averaged Z -score significance threshold. Black, siRNA (SMARTPools) targeting 779 protein kinase genes; red, siCON and blue, siBRCA1. Reflecting the reproducibility and sensitivity of the screen, siCON and siBRCA1 Z -scores were widely separated, with a screen Z ′-factor ( <xref ref-type= Zhang et al , 1999 ) of 0.34. " title="... A ) HTS method. CAL51 cells plated in 96-well plates were transfected with siRNA. Each transfection plate ..." property="contentUrl" width="100%" height="100%"/>
    Figure Legend Snippet: PARP-inhibitor synthetic lethality screen with protein kinase siRNA library. ( A ) HTS method. CAL51 cells plated in 96-well plates were transfected with siRNA. Each transfection plate contained 80 experimental siRNAs (SMARTPools of four different siRNA targeting the same gene) supplemented with four wells of non-targeting siCON, and two wells of siRNA directed against BRCA1 (positive control). Transfected cells were divided into six replica plates, half treated with DMSO vehicle alone and half with PARP inhibitor KU0058948 at 1 μM, the SF80 of CAL51. Cell viability was assessed after 5 days of KU0058948 exposure using CellTiter-Glo Luminescent Cell Viability Assay (Promega). ( B ) Reproducibility of HTS method. Correlation of the effect of siRNA on cell growth in vehicle-treated plates from two replicates of the entire screen. Spearman correlation coefficient, r =0.83. ( C ) Correlation of KU0058948 sensitivity Z -scores from two replicates of the entire screen. r =0.54. ( D ) Scatter plot of averaged Z -scores from PARP inhibitor sensitivity screen carried out in duplicate with KU0058948. Red line indicates −3 averaged Z -score significance threshold. Black, siRNA (SMARTPools) targeting 779 protein kinase genes; red, siCON and blue, siBRCA1. Reflecting the reproducibility and sensitivity of the screen, siCON and siBRCA1 Z -scores were widely separated, with a screen Z ′-factor ( Zhang et al , 1999 ) of 0.34.

    Techniques Used: Transfection, Positive Control, Cell Viability Assay, Electron Microscopy

    10) Product Images from "hvTRA, a novel TRAIL receptor agonist, induces apoptosis and sustained growth retardation in melanoma"

    Article Title: hvTRA, a novel TRAIL receptor agonist, induces apoptosis and sustained growth retardation in melanoma

    Journal: Cell Death Discovery

    doi: 10.1038/cddiscovery.2016.81

    Sensitivity to hvTRA, vemurafenib and the combination in patient biopsies. Eleven biopsies from lymph node metastases from melanoma patients were made into single-cell suspensions, and seeded in 96-well round-bottom plates and exposed to hvTRA (5 μ g/ml), vemurafenib (5 μ M) or the combination. Cell viability was measured after 120 h using CellTiter-Glo Luminescent assay. Three technical replicates were performed, and the error bars represent±S.D. The luminescent signal from untreated control cells from each patient sample were set to 1, indicated by the dotted line. For DR5 expression 0; not expressed, 1; 0–10% of the cells express DR5, 2; 10–50% of the cells express DR5, 3; > 50% of the cells express DR5. Patient biopsies 294 and 328 have mutated NRAS. * P
    Figure Legend Snippet: Sensitivity to hvTRA, vemurafenib and the combination in patient biopsies. Eleven biopsies from lymph node metastases from melanoma patients were made into single-cell suspensions, and seeded in 96-well round-bottom plates and exposed to hvTRA (5 μ g/ml), vemurafenib (5 μ M) or the combination. Cell viability was measured after 120 h using CellTiter-Glo Luminescent assay. Three technical replicates were performed, and the error bars represent±S.D. The luminescent signal from untreated control cells from each patient sample were set to 1, indicated by the dotted line. For DR5 expression 0; not expressed, 1; 0–10% of the cells express DR5, 2; 10–50% of the cells express DR5, 3; > 50% of the cells express DR5. Patient biopsies 294 and 328 have mutated NRAS. * P

    Techniques Used: Luminescence Assay, Expressing

    11) Product Images from "SSTR5 P335L monoclonal antibody differentiates pancreatic neuroendocrine tumor patients with different SSTR5 genotypes"

    Article Title: SSTR5 P335L monoclonal antibody differentiates pancreatic neuroendocrine tumor patients with different SSTR5 genotypes

    Journal:

    doi: 10.1016/j.surg.2011.09.044

    CAPAN-1 and PANC-1 cells differentially respond to RPL-1980 in cell proliferation. CAPAN-1 and PANC-1 cells were seeded in 96-well plates (5 × 103 cells/well). After 24 h, the cells were treated with 10−5 M of RPL-1980 for 60 h before examining the cell proliferation by performing MTS assay. Absorbance at 492 nm was recorded. Data were expressed as the mean +/− standard deviation of triplicate values. * P < 0.05.
    Figure Legend Snippet: CAPAN-1 and PANC-1 cells differentially respond to RPL-1980 in cell proliferation. CAPAN-1 and PANC-1 cells were seeded in 96-well plates (5 × 103 cells/well). After 24 h, the cells were treated with 10−5 M of RPL-1980 for 60 h before examining the cell proliferation by performing MTS assay. Absorbance at 492 nm was recorded. Data were expressed as the mean +/− standard deviation of triplicate values. * P < 0.05.

    Techniques Used: MTS Assay, Standard Deviation

    12) Product Images from "Nanoparticles That Sense Thrombin Activity As Synthetic Urinary Biomarkers of Thrombosis"

    Article Title: Nanoparticles That Sense Thrombin Activity As Synthetic Urinary Biomarkers of Thrombosis

    Journal: ACS Nano

    doi: 10.1021/nn403550c

    Designing ligand-encoded reporters for detection by ELISA. (A) Schematic of ligand-encoded reporters R 1 and R 2 along with chemical structures of associated ligands. (B) Schematic of ELISA sandwich complex and photograph of developed 96-well plates showing specific detection of R 1 and R 2 spiked into control urine samples. (C) Absorbance values (λ = 450 nm) of wells coated with anti-Flsc antibodies used to detect serial dilutions of R 1 , R 1 + R 2 , and R 2 in urine ( n = 3 per condition, s.d.). (D) Quantification of the level of cleaved reporters (R 1 ) released from NWs after incubation with increasing concentrations of thrombin ( n = 3 per dose, s.d.).
    Figure Legend Snippet: Designing ligand-encoded reporters for detection by ELISA. (A) Schematic of ligand-encoded reporters R 1 and R 2 along with chemical structures of associated ligands. (B) Schematic of ELISA sandwich complex and photograph of developed 96-well plates showing specific detection of R 1 and R 2 spiked into control urine samples. (C) Absorbance values (λ = 450 nm) of wells coated with anti-Flsc antibodies used to detect serial dilutions of R 1 , R 1 + R 2 , and R 2 in urine ( n = 3 per condition, s.d.). (D) Quantification of the level of cleaved reporters (R 1 ) released from NWs after incubation with increasing concentrations of thrombin ( n = 3 per dose, s.d.).

    Techniques Used: Enzyme-linked Immunosorbent Assay, Incubation

    13) Product Images from "Nanoparticles That Sense Thrombin Activity As Synthetic Urinary Biomarkers of Thrombosis"

    Article Title: Nanoparticles That Sense Thrombin Activity As Synthetic Urinary Biomarkers of Thrombosis

    Journal: ACS Nano

    doi: 10.1021/nn403550c

    Designing ligand-encoded reporters for detection by ELISA. (A) Schematic of ligand-encoded reporters R 1 and R 2 along with chemical structures of associated ligands. (B) Schematic of ELISA sandwich complex and photograph of developed 96-well plates showing specific detection of R 1 and R 2 spiked into control urine samples. (C) Absorbance values (λ = 450 nm) of wells coated with anti-Flsc antibodies used to detect serial dilutions of R 1 , R 1 + R 2 , and R 2 in urine ( n = 3 per condition, s.d.). (D) Quantification of the level of cleaved reporters (R 1 ) released from NWs after incubation with increasing concentrations of thrombin ( n = 3 per dose, s.d.).
    Figure Legend Snippet: Designing ligand-encoded reporters for detection by ELISA. (A) Schematic of ligand-encoded reporters R 1 and R 2 along with chemical structures of associated ligands. (B) Schematic of ELISA sandwich complex and photograph of developed 96-well plates showing specific detection of R 1 and R 2 spiked into control urine samples. (C) Absorbance values (λ = 450 nm) of wells coated with anti-Flsc antibodies used to detect serial dilutions of R 1 , R 1 + R 2 , and R 2 in urine ( n = 3 per condition, s.d.). (D) Quantification of the level of cleaved reporters (R 1 ) released from NWs after incubation with increasing concentrations of thrombin ( n = 3 per dose, s.d.).

    Techniques Used: Enzyme-linked Immunosorbent Assay, Incubation

    14) Product Images from "In vitro Real-time Measurement of the Intra-bacterial Redox Potential"

    Article Title: In vitro Real-time Measurement of the Intra-bacterial Redox Potential

    Journal:

    doi:

    Example layout of a 96-well imaging plate In lane 1 all bacteria express roGFP2 while in lane 2 contains well with empty-vector containing bacteria to determine the background fluorescence (BG) for each condition. The two adjacent well e.g. A1 and A2 undergo an identical challenge with oxidizing agents. In row G, 100 mM of hydrogen peroxide is added and these results are used as maximum oxidized values for normalization. In row H, 10 mM DTT is added and these results are used as maximum reduced values for normalization.
    Figure Legend Snippet: Example layout of a 96-well imaging plate In lane 1 all bacteria express roGFP2 while in lane 2 contains well with empty-vector containing bacteria to determine the background fluorescence (BG) for each condition. The two adjacent well e.g. A1 and A2 undergo an identical challenge with oxidizing agents. In row G, 100 mM of hydrogen peroxide is added and these results are used as maximum oxidized values for normalization. In row H, 10 mM DTT is added and these results are used as maximum reduced values for normalization.

    Techniques Used: Imaging, Plasmid Preparation, Fluorescence

    15) Product Images from "Screening and identification of compounds with antiviral activity against hepatitis B virus using a safe compound library and novel real-time immune-absorbance PCR-based high throughput system"

    Article Title: Screening and identification of compounds with antiviral activity against hepatitis B virus using a safe compound library and novel real-time immune-absorbance PCR-based high throughput system

    Journal:

    doi: 10.1016/j.antiviral.2013.02.001

    Real-time IA-PCR specifically detects enveloped HBV DNA in G2.215 culture medium G2.215 cells were seeded in 96-well plates and incubated with lamivudine at different doses for 6 days. Cell culture medium was incubated with anti-HBs pre-immobilized on 96-well PCR plates at 4°C overnight. HBV DNA in captured virions was then amplified by real-time PCR. Medium from HepG2 cells and G2.215 cell medium incubated in wells coated with an anti-His antibody instead of anti-HBs were used as negative controls. A.) A dose-dependent increase in CT value was seen in medium from cells incubated with lamivudine. B.) The effect on CT levels was converted to reduction of HBV DNA by correlating to virus copy number using a standard curve of HBV genomic DNA. C). Serial CT dilution of HBV genomic DNA served as the standard curve for the real-time PCR assay.
    Figure Legend Snippet: Real-time IA-PCR specifically detects enveloped HBV DNA in G2.215 culture medium G2.215 cells were seeded in 96-well plates and incubated with lamivudine at different doses for 6 days. Cell culture medium was incubated with anti-HBs pre-immobilized on 96-well PCR plates at 4°C overnight. HBV DNA in captured virions was then amplified by real-time PCR. Medium from HepG2 cells and G2.215 cell medium incubated in wells coated with an anti-His antibody instead of anti-HBs were used as negative controls. A.) A dose-dependent increase in CT value was seen in medium from cells incubated with lamivudine. B.) The effect on CT levels was converted to reduction of HBV DNA by correlating to virus copy number using a standard curve of HBV genomic DNA. C). Serial CT dilution of HBV genomic DNA served as the standard curve for the real-time PCR assay.

    Techniques Used: IA, Polymerase Chain Reaction, Incubation, Cell Culture, Amplification, Real-time Polymerase Chain Reaction

    16) Product Images from "Paeoniflorin Inhibits Receptor Activator for Nuclear Factor κB (RANK) Ligand-Induced Osteoclast Differentiation In Vitro and Particle-Induced Osteolysis In Vivo"

    Article Title: Paeoniflorin Inhibits Receptor Activator for Nuclear Factor κB (RANK) Ligand-Induced Osteoclast Differentiation In Vitro and Particle-Induced Osteolysis In Vivo

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    doi: 10.12659/MSM.907739

    Paeoniflorin inhibits RANKL-induced osteoclast formation bone resorption. ( A ) Bone marrow macrophages (BMMs) were incubated with various concentrations of PF for 24 h. PF toxicity was measured using the cell counting kit 8 (CCK-8) assay. OD, optical density; ( B, C ) BMMs were incubated with RANKL (50 ng/mL) + murine colony stimulating factor (M-CSF) (30 ng/mL) in a 96-well plate along with various concentrations of paeoniflorin for 5 days. Tartrate-resistant acid phosphatase (TRAP)-positive cells in each well were counted; ( D, E ) BMMs were seeded in the osteo assay plate, cultured in the presence of RANKL (50 ng/ml) + M-CSF (30 ng/ml) and with the indicated concentration of PF. After 5 days of incubation, pit resorption areas were measured. All experiments were repeated 3 times. One-way ANOVA and Student-Newman-Keuls (SNK) tests were performed to measure significances between each group. * p < 0.05, ** p < 0.01, scale bar=100 μm.
    Figure Legend Snippet: Paeoniflorin inhibits RANKL-induced osteoclast formation bone resorption. ( A ) Bone marrow macrophages (BMMs) were incubated with various concentrations of PF for 24 h. PF toxicity was measured using the cell counting kit 8 (CCK-8) assay. OD, optical density; ( B, C ) BMMs were incubated with RANKL (50 ng/mL) + murine colony stimulating factor (M-CSF) (30 ng/mL) in a 96-well plate along with various concentrations of paeoniflorin for 5 days. Tartrate-resistant acid phosphatase (TRAP)-positive cells in each well were counted; ( D, E ) BMMs were seeded in the osteo assay plate, cultured in the presence of RANKL (50 ng/ml) + M-CSF (30 ng/ml) and with the indicated concentration of PF. After 5 days of incubation, pit resorption areas were measured. All experiments were repeated 3 times. One-way ANOVA and Student-Newman-Keuls (SNK) tests were performed to measure significances between each group. * p < 0.05, ** p < 0.01, scale bar=100 μm.

    Techniques Used: Incubation, Cell Counting, CCK-8 Assay, Cell Culture, Concentration Assay

    17) Product Images from "Identification of Two Subgroups of Type I IFNs in Perciforme Fish Large Yellow Croaker Larimichthys crocea Provides Novel Insights into Function and Regulation of Fish Type I IFNs"

    Article Title: Identification of Two Subgroups of Type I IFNs in Perciforme Fish Large Yellow Croaker Larimichthys crocea Provides Novel Insights into Function and Regulation of Fish Type I IFNs

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2016.00343

    Effects of IRF3 and IRF7 on induction of IFN promoter activity . (A) Full-length IFNd promoter (IFNdP1), (B) the IFNdP deletion constructs, (C) full-length IFNh promoter (IFNhP1), (D) the IFNhP deletion constructs. EPC cells (5 × 10 4 /well) were seeded in 96-well plates overnight and cotransfected with 50 ng of pGL3-IFNP plasmids, 50 ng of pCMV-HA-IRF3 or -IRF7, and 1 ng of pRL-TK. (E–G) Cooperative effect of IRF3 and IRF7 on IFN promoter. EPC cells were seeded in 96-well plates overnight and cotransfected with 50 ng of IFNdP1 (E) , the IFNdP deletion constructs (F) or IFNhP1 (G) , the indicated expression constructs (100 ng total at a ratio of 1:1), and 1 ng of pRL-TK. After 48 h of transfection, the cells were harvested for detection of luciferase activity. The empty vectors (EV) were used as controls. All data were obtained from three independent experiments with three replicates in each experiment. Error bars represent ±SEM. * p
    Figure Legend Snippet: Effects of IRF3 and IRF7 on induction of IFN promoter activity . (A) Full-length IFNd promoter (IFNdP1), (B) the IFNdP deletion constructs, (C) full-length IFNh promoter (IFNhP1), (D) the IFNhP deletion constructs. EPC cells (5 × 10 4 /well) were seeded in 96-well plates overnight and cotransfected with 50 ng of pGL3-IFNP plasmids, 50 ng of pCMV-HA-IRF3 or -IRF7, and 1 ng of pRL-TK. (E–G) Cooperative effect of IRF3 and IRF7 on IFN promoter. EPC cells were seeded in 96-well plates overnight and cotransfected with 50 ng of IFNdP1 (E) , the IFNdP deletion constructs (F) or IFNhP1 (G) , the indicated expression constructs (100 ng total at a ratio of 1:1), and 1 ng of pRL-TK. After 48 h of transfection, the cells were harvested for detection of luciferase activity. The empty vectors (EV) were used as controls. All data were obtained from three independent experiments with three replicates in each experiment. Error bars represent ±SEM. * p

    Techniques Used: Activity Assay, Construct, Hemagglutination Assay, Expressing, Transfection, Luciferase

    The structure and transcriptional activity of large yellow croaker IFN promoters . (A,D) Schematic representation of IFNd (A) and IFNh (D) promoters and a series of deletion constructs. (B,E) Transcriptional activity of IFN promoters. EPC cells (5 × 10 4 /well) were seeded in 96-well plates overnight and cotransfected with 100 ng of pGL3-IFNdP plasmid (B) or pGL3-IFNhP plasmid (E) and 2 ng of pRL-TK using the Fugene ® HD transfection reagent. Transcript levels were determined by real-time PCR. (C,F) Induction of large yellow croaker IFN promoter activity by poly(I:C). EPC cells (5 × 10 4 /well) were seeded in 96-well plates overnight and cotransfected with 50 ng of pGL3-IFNdP plasmid (C) or pGL3-IFNhP plasmid (F) , poly(I:C) (as indicated doses), and 1 ng of pRL-TK using the Fugene ® HD transfection reagent. After 48 h of transfection, the cells were harvested for detection of luciferase activity. All data were obtained from three independent experiments with three replicates in each experiment. Error bars represent ±SEM of three independent experiments. * p
    Figure Legend Snippet: The structure and transcriptional activity of large yellow croaker IFN promoters . (A,D) Schematic representation of IFNd (A) and IFNh (D) promoters and a series of deletion constructs. (B,E) Transcriptional activity of IFN promoters. EPC cells (5 × 10 4 /well) were seeded in 96-well plates overnight and cotransfected with 100 ng of pGL3-IFNdP plasmid (B) or pGL3-IFNhP plasmid (E) and 2 ng of pRL-TK using the Fugene ® HD transfection reagent. Transcript levels were determined by real-time PCR. (C,F) Induction of large yellow croaker IFN promoter activity by poly(I:C). EPC cells (5 × 10 4 /well) were seeded in 96-well plates overnight and cotransfected with 50 ng of pGL3-IFNdP plasmid (C) or pGL3-IFNhP plasmid (F) , poly(I:C) (as indicated doses), and 1 ng of pRL-TK using the Fugene ® HD transfection reagent. After 48 h of transfection, the cells were harvested for detection of luciferase activity. All data were obtained from three independent experiments with three replicates in each experiment. Error bars represent ±SEM of three independent experiments. * p

    Techniques Used: Activity Assay, Construct, Plasmid Preparation, Transfection, Real-time Polymerase Chain Reaction, Luciferase

    18) Product Images from ""

    Article Title:

    Journal:

    doi: 10.1074/jbc.M112.348151

    Effect of alanine substitutions on A2-8 cell attachment activity. 96-well plates were coated with various amounts of 12 Ala-substituted A2-8 peptides () and examined for cell attachment activity using HDFs. A–C , HDFs were added to the wells
    Figure Legend Snippet: Effect of alanine substitutions on A2-8 cell attachment activity. 96-well plates were coated with various amounts of 12 Ala-substituted A2-8 peptides () and examined for cell attachment activity using HDFs. A–C , HDFs were added to the wells

    Techniques Used: Cell Attachment Assay, Activity Assay

    Effect of 9 active peptides from short arm region and various integrin-binding peptides on the attachment of HDFs to rec-a2N. 96-well plates were coated with 10 μg/well of rec-a2N proteins. A , various amount of cell adhesive peptides from the
    Figure Legend Snippet: Effect of 9 active peptides from short arm region and various integrin-binding peptides on the attachment of HDFs to rec-a2N. 96-well plates were coated with 10 μg/well of rec-a2N proteins. A , various amount of cell adhesive peptides from the

    Techniques Used: Binding Assay

    Cell attachment to the peptide-coated plates and peptide-Sepharose beads. A and B , 96-well plates were coated with various amounts of synthetic peptides and examined for cell attachment activity using HDFs. The cells were added to the wells for 1 h. After
    Figure Legend Snippet: Cell attachment to the peptide-coated plates and peptide-Sepharose beads. A and B , 96-well plates were coated with various amounts of synthetic peptides and examined for cell attachment activity using HDFs. The cells were added to the wells for 1 h. After

    Techniques Used: Cell Attachment Assay, Activity Assay

    Effect of anti-integrin antibodies on cell attachment to A2-8 and effect of various peptides on cell attachment to rec-a2LN. A , 96-well plates were coated with 10 μg/well of A2-8. 10 and/or 30 μg/ml of anti-integrin antibodies were added
    Figure Legend Snippet: Effect of anti-integrin antibodies on cell attachment to A2-8 and effect of various peptides on cell attachment to rec-a2LN. A , 96-well plates were coated with 10 μg/well of A2-8. 10 and/or 30 μg/ml of anti-integrin antibodies were added

    Techniques Used: Cell Attachment Assay

    Neurite outgrowth of PC12 cells on the peptide-coated plates. A , 10 μg/well of peptides were coated on 96-well plates. PC12 cells (3.0 × 10 3 cells/well) were seeded in the wells and incubated for 24 h. After the cells were fixed and stained,
    Figure Legend Snippet: Neurite outgrowth of PC12 cells on the peptide-coated plates. A , 10 μg/well of peptides were coated on 96-well plates. PC12 cells (3.0 × 10 3 cells/well) were seeded in the wells and incubated for 24 h. After the cells were fixed and stained,

    Techniques Used: Incubation, Staining

    19) Product Images from "Mechanisms Underlying the Antiproliferative and Prodifferentiative Effects of Psoralen on Adult Neural Stem Cells via DNA Microarray"

    Article Title: Mechanisms Underlying the Antiproliferative and Prodifferentiative Effects of Psoralen on Adult Neural Stem Cells via DNA Microarray

    Journal: Evidence-based Complementary and Alternative Medicine : eCAM

    doi: 10.1155/2013/452948

    Effects of Psoralen on the differentiation of NSC. Single NSCs were seeded at a density of 50 cells/ μ L in PDL-coated 96-well plate in differentiation medium for 48 h without Psoralen (a, b) or with 100 nM Psoralen (c, d). Cells were subjected to primary antibodies of GFAP, TuJ1 and corresponding secondary antibodies and visualized with Alexa-conjugated 594 F(ab)′2 goat anti-rabit antibody and 488 Alexa-conjugated goat anti mouse IgG (H + L). The ratio of GFAP, TuJ1-positive cells against DAPI-stained cells was calculated. Psoralen significantly increased the GFAP-positive cells and decreased the TuJ1-positive cells (e). The same cells were performed using Western blotting. Psoralen significantly increased the GFAP, decreased the TuJ1 protein expression (f). Scale bars: 100 μ m. Results were expressed as mean ± S.D. from three independent experiments. * P
    Figure Legend Snippet: Effects of Psoralen on the differentiation of NSC. Single NSCs were seeded at a density of 50 cells/ μ L in PDL-coated 96-well plate in differentiation medium for 48 h without Psoralen (a, b) or with 100 nM Psoralen (c, d). Cells were subjected to primary antibodies of GFAP, TuJ1 and corresponding secondary antibodies and visualized with Alexa-conjugated 594 F(ab)′2 goat anti-rabit antibody and 488 Alexa-conjugated goat anti mouse IgG (H + L). The ratio of GFAP, TuJ1-positive cells against DAPI-stained cells was calculated. Psoralen significantly increased the GFAP-positive cells and decreased the TuJ1-positive cells (e). The same cells were performed using Western blotting. Psoralen significantly increased the GFAP, decreased the TuJ1 protein expression (f). Scale bars: 100 μ m. Results were expressed as mean ± S.D. from three independent experiments. * P

    Techniques Used: Staining, Western Blot, Expressing

    Effects of Psoralen on the proliferation of NSC. Single NSCs were plated at a density of 5000 cells per well in PDL-coated 96-well plate for 12 h. Then, cells were subjected to 10 nM EdU for 2 h, followed by addition of 100 nM OA (b) or not (a). Then Edu immunofluorescence analysis was performed. The cell nuclei were counterstained with DAPI. The percentage of EdU-positive cells in total of 1000 cells was calculated. As a result, Psoralen significantly inhibited the DNA incorporation (c). Scale bars: 100 μ m. Results were expressed as mean ± S.D. from four independent experiments. * P
    Figure Legend Snippet: Effects of Psoralen on the proliferation of NSC. Single NSCs were plated at a density of 5000 cells per well in PDL-coated 96-well plate for 12 h. Then, cells were subjected to 10 nM EdU for 2 h, followed by addition of 100 nM OA (b) or not (a). Then Edu immunofluorescence analysis was performed. The cell nuclei were counterstained with DAPI. The percentage of EdU-positive cells in total of 1000 cells was calculated. As a result, Psoralen significantly inhibited the DNA incorporation (c). Scale bars: 100 μ m. Results were expressed as mean ± S.D. from four independent experiments. * P

    Techniques Used: Immunofluorescence

    20) Product Images from "Hypoxic 3D in vitro culture models reveal distinct resistance processes to TKIs in renal cancer cells"

    Article Title: Hypoxic 3D in vitro culture models reveal distinct resistance processes to TKIs in renal cancer cells

    Journal: Cell & Bioscience

    doi: 10.1186/s13578-017-0197-8

    HKCSCs cells are resistant to sorafenib in hypoxia and to axitinib in normoxia. Hypoxia decreases the amount of colonies visible in 96-well plates. Representative images of colonies (performed in triplicate). Magnification 4×, scale bar = 200 µm. The scheme of the figure is shown below. Above 2D monolayer cell culture phenotypic view is presented for comparison. U untreated cells, N normoxia, H hypoxia, r resistant cells, s sensitized cells, A axitinib, S sorafenib. Resistant HKCSCs are marked with a red square
    Figure Legend Snippet: HKCSCs cells are resistant to sorafenib in hypoxia and to axitinib in normoxia. Hypoxia decreases the amount of colonies visible in 96-well plates. Representative images of colonies (performed in triplicate). Magnification 4×, scale bar = 200 µm. The scheme of the figure is shown below. Above 2D monolayer cell culture phenotypic view is presented for comparison. U untreated cells, N normoxia, H hypoxia, r resistant cells, s sensitized cells, A axitinib, S sorafenib. Resistant HKCSCs are marked with a red square

    Techniques Used: Cell Culture

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    Clone Assay:

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    Article Snippet: The following mAb clones were used: B220-PE-Cy7, CD4-PE-Cy7, CD8α-PerCP-Cy5.5, CD11b-PE-Cy7, CD11c-FITC, CD25–Alexa Fluor 647, CD44-FITC, CD45.1-FITC, CD45.2-BV605, CD62L-PE-Cy7, CD103-biotin (followed by streptavidin-BV711; BD), Foxp3-PE (eBioscience), Helios-APC, I-A(b)-PE, Ki-67–Alexa Fluor 647, and TCR-β–Pacific blue. .. Intracellular staining to detect Foxp3 expression was performed according to the manufacturer’s instructions, using the manufacturer’s protocol for Foxp3 staining in 96-well plates (eBioscience).

    Article Title: Isolation Efficiency of Mouse Pancreatic Stem Cells Is Age Dependent
    Article Snippet: Once the cells had attached and spread, cells with a fibroblast morphology (nonductal cells) were removed using a rubberscrapper (Life Technologies Japan). .. The “duct-like” cells were cultured in DMEM with 20% FBS in 96-well plates (Life Technologies Japan) and cloned by limiting dilution. .. After single cell cloning, the mouse pancreatic stem cells were maintained in specific culture condition with lot-limited FBS ( ) during the early studies (first study using a 0-week-old pancreas and first to fifth studies using 8-week-old pancreata) or in culture condition of mouse ES cells ( ) during the later studies (studies except first study using a 0-week-old pancreas and first to fifth studies using 8-week-old pancreata).

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    Article Snippet: Subconfluent cells were passaged after detachment with 0.25% trypsin-EDTA, and cell lines were established after > 60 passages. .. For cloning, one cell per well was plated in separate 96-well plates (Thermo Scientific, MA, USA). .. For measuring the growth curve and population doublings, the established cell lines were plated in 24-well plates (Thermo Scientific) at 5000 cells/well in 1 mL of Medium 199 containing 10% FBS.

    Luciferase:

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    Article Snippet: Paragraph title: Luciferase Activity Assay ... The EPC cells (5 × 104 /well) were seeded in 96-well plates (Thermo Fisher Scientific) overnight and cotransfected with 100 ng of pGL3-IFNP plasmid or pGL3-Basic plasmid (control) and 2 ng pRL-TK plasmid using the Fugene® HD transfection reagent.

    Filtration:

    Article Title: Uptake of HLA Alloantigens via CD89 and CD206 Does Not Enhance Antigen Presentation by Indirect Allorecognition
    Article Snippet: Monomers were purified by gel filtration HPLC, routinely tested, and validated. .. MaxiSorp flat-bottomed 96-well plates (Nunc, Thermo Scientific) were coated with 1 μ g of CD206-A/CD206 or CD89-A/CD89 (diluted in PBS) and incubated for 2 hours at 37°C.

    Stable Transfection:

    Article Title: MiR-16 regulates the pro-tumorigenic potential of lung fibroblasts through the inhibition of HGF production in an FGFR-1- and MEK1-dependent manner
    Article Snippet: As the large-scale screening experiment was not feasible with primary CAFs due to the limited cell number, fibroblasts (CAFs, AFs, and NFs) from different patients were transduced with retroviral particles to stably express human TERT (hTERT) and immortalize cells (see above). .. For the high-throughput screening, at day 1, CAF154-hTERT fibroblasts were reverse transfected with a library of human miRNA mimics composed of 988 mature miRNAs arrayed on 96-well plates (875 unique sequences, miRBase v.13.0, miRIDIAN technology, Dharmacon).

    Mouse Assay:

    Article Title: Isolation Efficiency of Mouse Pancreatic Stem Cells Is Age Dependent
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    Article Title: Constitutive phosphorylation of the mTORC2/Akt/4E-BP1 pathway in newly derived canine hemangiosarcoma cell lines
    Article Snippet: These tumor tissues were subcutaneously transplanted into the right and left dorsal area of the trunk of 3-week-old male KSN/Slc nude mice (Japan SLC, Inc., Hamamatsu, Japan), and xenograft models were established after > 5 passages. .. For cloning, one cell per well was plated in separate 96-well plates (Thermo Scientific, MA, USA).

    Cytometry:

    Article Title: Uptake of HLA Alloantigens via CD89 and CD206 Does Not Enhance Antigen Presentation by Indirect Allorecognition
    Article Snippet: “Avidinylated antibody” (referred to as CD206-A or CD89-A) specificity was confirmed by flow cytometry and compared with the non-avidinylated antibody (CD206 or CD89). .. MaxiSorp flat-bottomed 96-well plates (Nunc, Thermo Scientific) were coated with 1 μ g of CD206-A/CD206 or CD89-A/CD89 (diluted in PBS) and incubated for 2 hours at 37°C.

    Article Title: The lysophosphatidylserine receptor GPR174 constrains regulatory T cell development and function
    Article Snippet: Paragraph title: Flow cytometry. ... Intracellular staining to detect Foxp3 expression was performed according to the manufacturer’s instructions, using the manufacturer’s protocol for Foxp3 staining in 96-well plates (eBioscience).

    Construct:

    Article Title: Identification of Two Subgroups of Type I IFNs in Perciforme Fish Large Yellow Croaker Larimichthys crocea Provides Novel Insights into Function and Regulation of Fish Type I IFNs
    Article Snippet: For luciferase assays, the recombinant plasmids were constructed by inserting the promoter regions of two IFN genes and a series of their respective deleted fragments into the dual luciferase reporter plasmid pGL3-Basic (pGL3-IFNPs, primers in Table S1 in Supplementary Material; Promega). .. The EPC cells (5 × 104 /well) were seeded in 96-well plates (Thermo Fisher Scientific) overnight and cotransfected with 100 ng of pGL3-IFNP plasmid or pGL3-Basic plasmid (control) and 2 ng pRL-TK plasmid using the Fugene® HD transfection reagent.

    Enzyme-linked Immunosorbent Assay:

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    Article Snippet: HBV infection markers (HBsAg, anti-HBs, HBeAg, anti-HBe and anti-HBc) were quantified with Light Initiated Chemiluminescence Analyzing system (Bo Yang), while the HBV DNA viral load was determined by StepOnePlus system (KHB). .. For HBsAg detection by G12-based ELISA, 96-well plates (Thermo Fisher) were coated at 4°C for 16 hours with G12 diluted 1:10,000 in carbonate sodium buffer (pH9.6). .. The plates were washed with PBST, blocked at 37°C for 2 hours with 5% skimmed milk dissolved in PBST.

    Article Title: Uptake of HLA Alloantigens via CD89 and CD206 Does Not Enhance Antigen Presentation by Indirect Allorecognition
    Article Snippet: An ELISA was set up to confirm the formation of the monomeric HLA-A2 and “antibody” (Ab) complex. .. MaxiSorp flat-bottomed 96-well plates (Nunc, Thermo Scientific) were coated with 1 μ g of CD206-A/CD206 or CD89-A/CD89 (diluted in PBS) and incubated for 2 hours at 37°C.

    Incubation:

    Article Title:
    Article Snippet: The cell attachment assay using synthetic peptides and recombinant proteins was performed in 96-well plates (Thermo Fisher Scientific). .. The cell attachment assay using synthetic peptides and recombinant proteins was performed in 96-well plates (Thermo Fisher Scientific).

    Article Title: Mixed Fibronectin-Derived Peptides Conjugated to a Chitosan Matrix Effectively Promotes Biological Activities through Integrins, α4β1, α5β1, αvβ3, and Syndecan
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    Article Title:
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    Article Title: Uptake of HLA Alloantigens via CD89 and CD206 Does Not Enhance Antigen Presentation by Indirect Allorecognition
    Article Snippet: An ELISA was set up to confirm the formation of the monomeric HLA-A2 and “antibody” (Ab) complex. .. MaxiSorp flat-bottomed 96-well plates (Nunc, Thermo Scientific) were coated with 1 μ g of CD206-A/CD206 or CD89-A/CD89 (diluted in PBS) and incubated for 2 hours at 37°C. .. After each incubation step plates were washed extensively (at least 5x) with PBS containing 0.05% Tween-20 and 1% BSA (both from Sigma-Aldrich).

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    Article Title: Mechanisms Underlying the Antiproliferative and Prodifferentiative Effects of Psoralen on Adult Neural Stem Cells via DNA Microarray
    Article Snippet: In brief, single NSCs were grown in 96-well plates in DMEM/F12 medium containing 20 ng/mL hrFGF-2 and hrEGF (Invitrogen, CA, USA), Stempro NSC supplement (Invitrogen, CA, USA), 100 units/mL penicillin, and 100 μ g/mL streptomycin. .. EdU was added to the culture media in a final concentration of 10 μ M for 3 h. Cells were fixed in formaldehyde and penetrated with 0.5% Triton X-100.

    Article Title: hvTRA, a novel TRAIL receptor agonist, induces apoptosis and sustained growth retardation in melanoma
    Article Snippet: The cells were seeded in 96-well plates (Thermo Fisher Scientific, Rockford, IL, USA). .. The cells were seeded in 96-well plates (Thermo Fisher Scientific, Rockford, IL, USA).

    Article Title: Constitutive phosphorylation of the mTORC2/Akt/4E-BP1 pathway in newly derived canine hemangiosarcoma cell lines
    Article Snippet: For cloning, one cell per well was plated in separate 96-well plates (Thermo Scientific, MA, USA). .. For cloning, one cell per well was plated in separate 96-well plates (Thermo Scientific, MA, USA).

    Proliferation Assay:

    Article Title: Mechanisms Underlying the Antiproliferative and Prodifferentiative Effects of Psoralen on Adult Neural Stem Cells via DNA Microarray
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    Infection:

    Article Title: A human monoclonal antibody against small envelope protein of hepatitis B virus with potent neutralization effect
    Article Snippet: HBV infection markers (HBsAg, anti-HBs, HBeAg, anti-HBe and anti-HBc) were quantified with Light Initiated Chemiluminescence Analyzing system (Bo Yang), while the HBV DNA viral load was determined by StepOnePlus system (KHB). .. For HBsAg detection by G12-based ELISA, 96-well plates (Thermo Fisher) were coated at 4°C for 16 hours with G12 diluted 1:10,000 in carbonate sodium buffer (pH9.6).

    Expressing:

    Article Title: Anti-heparan Sulfate Peptides That Block Herpes Simplex Virus Infection in Vivo
    Article Snippet: A phage display library (PhDTM -12) expressing 12-mer peptides fused to a minor coat protein (pIII) of a non-lytic bacteriophage (M13) was purchased from New England Biolabs (Cambridge, MA). .. Both targets at a concentration of 10 μg/ml were used for overnight coating of wells of 96-well plates (Nalge Nunc International, Naperville, IL) in a humidifier chamber at 4 °C.

    Article Title: The lysophosphatidylserine receptor GPR174 constrains regulatory T cell development and function
    Article Snippet: Dead cells were excluded using Fixable Viability Dye eFluor780 (eBioscience). .. Intracellular staining to detect Foxp3 expression was performed according to the manufacturer’s instructions, using the manufacturer’s protocol for Foxp3 staining in 96-well plates (eBioscience). .. Cells were analyzed using an LSR-II flow cytometer (BD) equipped with 405-, 488-, 552-, and 640-nm lasers.

    Modification:

    Article Title: Anti-heparan Sulfate Peptides That Block Herpes Simplex Virus Infection in Vivo
    Article Snippet: Screening of the phage display library was accomplished by an affinity selection (or bio-panning) process during which phage populations were selected for their ability to bind HS and 3-OS HS (modified by 3-OST-3). .. Both targets at a concentration of 10 μg/ml were used for overnight coating of wells of 96-well plates (Nalge Nunc International, Naperville, IL) in a humidifier chamber at 4 °C.

    Article Title: Isolation Efficiency of Mouse Pancreatic Stem Cells Is Age Dependent
    Article Snippet: The tissue collagenase was digested (2 mg/ml) and cultured in Dulbecco’s modified Eagle’s medium (DMEM; Life Technologies Japan) with 20% lot-limited fetal bovine serum (FBS; BIO-WEST, Inc., Logan, UT, USA; S1560 Lot. .. The “duct-like” cells were cultured in DMEM with 20% FBS in 96-well plates (Life Technologies Japan) and cloned by limiting dilution.

    High Performance Liquid Chromatography:

    Article Title: Uptake of HLA Alloantigens via CD89 and CD206 Does Not Enhance Antigen Presentation by Indirect Allorecognition
    Article Snippet: Monomers were purified by gel filtration HPLC, routinely tested, and validated. .. MaxiSorp flat-bottomed 96-well plates (Nunc, Thermo Scientific) were coated with 1 μ g of CD206-A/CD206 or CD89-A/CD89 (diluted in PBS) and incubated for 2 hours at 37°C.

    Conjugation Assay:

    Article Title: Uptake of HLA Alloantigens via CD89 and CD206 Does Not Enhance Antigen Presentation by Indirect Allorecognition
    Article Snippet: Paragraph title: 2.4. Antibody Conjugation ... MaxiSorp flat-bottomed 96-well plates (Nunc, Thermo Scientific) were coated with 1 μ g of CD206-A/CD206 or CD89-A/CD89 (diluted in PBS) and incubated for 2 hours at 37°C.

    Flow Cytometry:

    Article Title: Uptake of HLA Alloantigens via CD89 and CD206 Does Not Enhance Antigen Presentation by Indirect Allorecognition
    Article Snippet: “Avidinylated antibody” (referred to as CD206-A or CD89-A) specificity was confirmed by flow cytometry and compared with the non-avidinylated antibody (CD206 or CD89). .. MaxiSorp flat-bottomed 96-well plates (Nunc, Thermo Scientific) were coated with 1 μ g of CD206-A/CD206 or CD89-A/CD89 (diluted in PBS) and incubated for 2 hours at 37°C.

    Article Title: The lysophosphatidylserine receptor GPR174 constrains regulatory T cell development and function
    Article Snippet: Paragraph title: Flow cytometry. ... Intracellular staining to detect Foxp3 expression was performed according to the manufacturer’s instructions, using the manufacturer’s protocol for Foxp3 staining in 96-well plates (eBioscience).

    Cell Culture:

    Article Title: Isolation Efficiency of Mouse Pancreatic Stem Cells Is Age Dependent
    Article Snippet: Once the cells had attached and spread, cells with a fibroblast morphology (nonductal cells) were removed using a rubberscrapper (Life Technologies Japan). .. The “duct-like” cells were cultured in DMEM with 20% FBS in 96-well plates (Life Technologies Japan) and cloned by limiting dilution. .. After single cell cloning, the mouse pancreatic stem cells were maintained in specific culture condition with lot-limited FBS ( ) during the early studies (first study using a 0-week-old pancreas and first to fifth studies using 8-week-old pancreata) or in culture condition of mouse ES cells ( ) during the later studies (studies except first study using a 0-week-old pancreas and first to fifth studies using 8-week-old pancreata).

    Article Title: Hypoxic 3D in vitro culture models reveal distinct resistance processes to TKIs in renal cancer cells
    Article Snippet: Cells were seeded on flat-bottom, 96-well plates (Thermo Fisher Scientific, Cat. No. 07-200-90) in the following concentrations to achieve their logarithmic phase of growth: HKCSCs—1000 cells/well, 769-P—1500 cells/well, Caki-1—2000 cells/well. .. Cells were seeded on flat-bottom, 96-well plates (Thermo Fisher Scientific, Cat. No. 07-200-90) in the following concentrations to achieve their logarithmic phase of growth: HKCSCs—1000 cells/well, 769-P—1500 cells/well, Caki-1—2000 cells/well.

    Article Title: Constitutive phosphorylation of the mTORC2/Akt/4E-BP1 pathway in newly derived canine hemangiosarcoma cell lines
    Article Snippet: Paragraph title: Cell culture ... For cloning, one cell per well was plated in separate 96-well plates (Thermo Scientific, MA, USA).

    Inhibition:

    Article Title: Mixed Fibronectin-Derived Peptides Conjugated to a Chitosan Matrix Effectively Promotes Biological Activities through Integrins, α4β1, α5β1, αvβ3, and Syndecan
    Article Snippet: The peptide-conjugated chitosan matrix was prepared in 96-well plates (Nalge Nunc, Inc., Rochester, NY) as described previously. .. The peptide-conjugated chitosan matrix was prepared in 96-well plates (Nalge Nunc, Inc., Rochester, NY) as described previously.

    Article Title:
    Article Snippet: 5 m m of peptide solution was added to the wells and incubated for 2 h. Then the plates were used for the cell attachment assay as described above. .. For inhibition of cell attachment to peptide-coated plates and to peptide-chitosan membranes, 96-well plates were prepared as described above. .. The cells were preincubated for 15 min at 37 °C in the presence of either 10 μg/ml heparin (Sigma-Aldrich), 5 m m EDTA (Wako Pure Chemicals), 10 μg/ml various cell adhesive peptides, and 10 or 30 μg/ml of integrin antibodies.

    Injection:

    Article Title: Isolation Efficiency of Mouse Pancreatic Stem Cells Is Age Dependent
    Article Snippet: Briefly, 2 ml of cold M199 medium (Life Technologies Japan, Tokyo, Japan) containing 2 mg/ml collagenase (Roche Boehringer Mannheim, Indianapolis, IN, USA) was injected into the cannulated common bile duct ( ). .. The “duct-like” cells were cultured in DMEM with 20% FBS in 96-well plates (Life Technologies Japan) and cloned by limiting dilution.

    Recombinant:

    Article Title:
    Article Snippet: The CD (J-720; JASCO, Tokyo, Japan) spectrum analysis did not show significant differences in rec-a2LN+ mutant proteins. .. The cell attachment assay using synthetic peptides and recombinant proteins was performed in 96-well plates (Thermo Fisher Scientific). .. 96-well plates were coated with various amounts of proteins in 50 μl of PBS overnight at 4 °C.

    Article Title: Identification of Two Subgroups of Type I IFNs in Perciforme Fish Large Yellow Croaker Larimichthys crocea Provides Novel Insights into Function and Regulation of Fish Type I IFNs
    Article Snippet: For luciferase assays, the recombinant plasmids were constructed by inserting the promoter regions of two IFN genes and a series of their respective deleted fragments into the dual luciferase reporter plasmid pGL3-Basic (pGL3-IFNPs, primers in Table S1 in Supplementary Material; Promega). .. The EPC cells (5 × 104 /well) were seeded in 96-well plates (Thermo Fisher Scientific) overnight and cotransfected with 100 ng of pGL3-IFNP plasmid or pGL3-Basic plasmid (control) and 2 ng pRL-TK plasmid using the Fugene® HD transfection reagent.

    Staining:

    Article Title: Mixed Fibronectin-Derived Peptides Conjugated to a Chitosan Matrix Effectively Promotes Biological Activities through Integrins, α4β1, α5β1, αvβ3, and Syndecan
    Article Snippet: The peptide-conjugated chitosan matrix was prepared in 96-well plates (Nalge Nunc, Inc., Rochester, NY) as described previously. .. Then, various concentrations of the peptides in a 0.1% TFA solution and a 1% NaHCO3 solution were added into the wells and incubated for 2 h. Then, the 96-well plates were blocked by the addition of 1% bovine serum albumin in DMEM for 1 h. Either HDFs or ARH-77 cells were added (100 μL, 2 × 104 cells) to each well and incubated at 37°C for 60 or 90 min in 5% CO2 .

    Article Title:
    Article Snippet: The cells were resuspended in DMEM/F-12 containing 30 n m Na2 SeO3 , 100 μg/ml transferrin, 20 n m progesterone (Sigma-Aldrich), 5 μg/ml insulin (Invitrogen), and 100 ng/ml NGF. .. The cells were added to 96-well plates at 3.0 × 103 cells/100 μl/well, incubated for 24 h, fixed with 20% formalin, and stained with 0.2% crystal violet. .. One hundred cells in each well were viewed under a BZ-8000, and the percentage of the active cells, which had neurites that extended twice the cell diameter in length and/or longer, was determined.

    Article Title: Mechanisms Underlying the Antiproliferative and Prodifferentiative Effects of Psoralen on Adult Neural Stem Cells via DNA Microarray
    Article Snippet: In brief, single NSCs were grown in 96-well plates in DMEM/F12 medium containing 20 ng/mL hrFGF-2 and hrEGF (Invitrogen, CA, USA), Stempro NSC supplement (Invitrogen, CA, USA), 100 units/mL penicillin, and 100 μ g/mL streptomycin. .. EdU was added to the culture media in a final concentration of 10 μ M for 3 h. Cells were fixed in formaldehyde and penetrated with 0.5% Triton X-100.

    Article Title: The lysophosphatidylserine receptor GPR174 constrains regulatory T cell development and function
    Article Snippet: Dead cells were excluded using Fixable Viability Dye eFluor780 (eBioscience). .. Intracellular staining to detect Foxp3 expression was performed according to the manufacturer’s instructions, using the manufacturer’s protocol for Foxp3 staining in 96-well plates (eBioscience). .. Cells were analyzed using an LSR-II flow cytometer (BD) equipped with 405-, 488-, 552-, and 640-nm lasers.

    Fluorescence:

    Article Title: MiR-16 regulates the pro-tumorigenic potential of lung fibroblasts through the inhibition of HGF production in an FGFR-1- and MEK1-dependent manner
    Article Snippet: For the high-throughput screening, at day 1, CAF154-hTERT fibroblasts were reverse transfected with a library of human miRNA mimics composed of 988 mature miRNAs arrayed on 96-well plates (875 unique sequences, miRBase v.13.0, miRIDIAN technology, Dharmacon). .. A549-green fluorescent protein (GFP) cells (3500 cells/well) were seeded at day 3 (48 h after fibroblast transfection) and co-cultured for further 48 h. The experiment was stopped by fixing the cells with 4% paraformaldehyde, and nuclei were counterstained with Hoechst 33342.

    Article Title: Mechanisms Underlying the Antiproliferative and Prodifferentiative Effects of Psoralen on Adult Neural Stem Cells via DNA Microarray
    Article Snippet: In brief, single NSCs were grown in 96-well plates in DMEM/F12 medium containing 20 ng/mL hrFGF-2 and hrEGF (Invitrogen, CA, USA), Stempro NSC supplement (Invitrogen, CA, USA), 100 units/mL penicillin, and 100 μ g/mL streptomycin. .. In brief, single NSCs were grown in 96-well plates in DMEM/F12 medium containing 20 ng/mL hrFGF-2 and hrEGF (Invitrogen, CA, USA), Stempro NSC supplement (Invitrogen, CA, USA), 100 units/mL penicillin, and 100 μ g/mL streptomycin.

    Isolation:

    Article Title: Anti-heparan Sulfate Peptides That Block Herpes Simplex Virus Infection in Vivo
    Article Snippet: A purified form of HS isolated from bovine kidney was purchased from Sigma. .. Both targets at a concentration of 10 μg/ml were used for overnight coating of wells of 96-well plates (Nalge Nunc International, Naperville, IL) in a humidifier chamber at 4 °C.

    Article Title: Isolation Efficiency of Mouse Pancreatic Stem Cells Is Age Dependent
    Article Snippet: Paragraph title: Isolation and Culture of Mouse Pancreatic Stem Cells and Islets ... The “duct-like” cells were cultured in DMEM with 20% FBS in 96-well plates (Life Technologies Japan) and cloned by limiting dilution.

    Transfection:

    Article Title: Identification of Two Subgroups of Type I IFNs in Perciforme Fish Large Yellow Croaker Larimichthys crocea Provides Novel Insights into Function and Regulation of Fish Type I IFNs
    Article Snippet: For luciferase assays, the recombinant plasmids were constructed by inserting the promoter regions of two IFN genes and a series of their respective deleted fragments into the dual luciferase reporter plasmid pGL3-Basic (pGL3-IFNPs, primers in Table S1 in Supplementary Material; Promega). .. The EPC cells (5 × 104 /well) were seeded in 96-well plates (Thermo Fisher Scientific) overnight and cotransfected with 100 ng of pGL3-IFNP plasmid or pGL3-Basic plasmid (control) and 2 ng pRL-TK plasmid using the Fugene® HD transfection reagent. .. After 48 h, the luciferase activity of total cell lysates was measured on a GloMax 20/20 luminometer (Promega) according to the Dual-Luciferase® Repoter Assay System (Promega).

    Article Title: MiR-16 regulates the pro-tumorigenic potential of lung fibroblasts through the inhibition of HGF production in an FGFR-1- and MEK1-dependent manner
    Article Snippet: As the large-scale screening experiment was not feasible with primary CAFs due to the limited cell number, fibroblasts (CAFs, AFs, and NFs) from different patients were transduced with retroviral particles to stably express human TERT (hTERT) and immortalize cells (see above). .. For the high-throughput screening, at day 1, CAF154-hTERT fibroblasts were reverse transfected with a library of human miRNA mimics composed of 988 mature miRNAs arrayed on 96-well plates (875 unique sequences, miRBase v.13.0, miRIDIAN technology, Dharmacon). .. Briefly, 15 μl miRNA (500 nM) was spotted per well, and a mix of 35 μl Opti-MEM containing RNAiMAX (Thermo Fisher Scientific) was added.

    Microscopy:

    Article Title: MiR-16 regulates the pro-tumorigenic potential of lung fibroblasts through the inhibition of HGF production in an FGFR-1- and MEK1-dependent manner
    Article Snippet: For the high-throughput screening, at day 1, CAF154-hTERT fibroblasts were reverse transfected with a library of human miRNA mimics composed of 988 mature miRNAs arrayed on 96-well plates (875 unique sequences, miRBase v.13.0, miRIDIAN technology, Dharmacon). .. A549-green fluorescent protein (GFP) cells (3500 cells/well) were seeded at day 3 (48 h after fibroblast transfection) and co-cultured for further 48 h. The experiment was stopped by fixing the cells with 4% paraformaldehyde, and nuclei were counterstained with Hoechst 33342.

    Article Title: Mechanisms Underlying the Antiproliferative and Prodifferentiative Effects of Psoralen on Adult Neural Stem Cells via DNA Microarray
    Article Snippet: In brief, single NSCs were grown in 96-well plates in DMEM/F12 medium containing 20 ng/mL hrFGF-2 and hrEGF (Invitrogen, CA, USA), Stempro NSC supplement (Invitrogen, CA, USA), 100 units/mL penicillin, and 100 μ g/mL streptomycin. .. In brief, single NSCs were grown in 96-well plates in DMEM/F12 medium containing 20 ng/mL hrFGF-2 and hrEGF (Invitrogen, CA, USA), Stempro NSC supplement (Invitrogen, CA, USA), 100 units/mL penicillin, and 100 μ g/mL streptomycin.

    Article Title: Constitutive phosphorylation of the mTORC2/Akt/4E-BP1 pathway in newly derived canine hemangiosarcoma cell lines
    Article Snippet: For cloning, one cell per well was plated in separate 96-well plates (Thermo Scientific, MA, USA). .. To examine the uptake of the acetylated low density lipoprotein (Ac-LDL) in HSA cell lines, subconfluent cells were incubated with 10 μg/mL DiI-Ac-LDL (Biomedical Technologies Inc., MA, USA) at 37 °C for 4 h in Medium 199 according to the manufacturer’s instructions.

    Avidin-Biotin Assay:

    Article Title: Uptake of HLA Alloantigens via CD89 and CD206 Does Not Enhance Antigen Presentation by Indirect Allorecognition
    Article Snippet: Antibodies targeting the MR (CD206, clone D547.3) or Fcα RI receptor (CD89, clone 2D11) were conjugated to avidin using the LL-avidin kit (Innova biosciences, UK) according to manufacturer's protocol. .. MaxiSorp flat-bottomed 96-well plates (Nunc, Thermo Scientific) were coated with 1 μ g of CD206-A/CD206 or CD89-A/CD89 (diluted in PBS) and incubated for 2 hours at 37°C.

    Cell Attachment Assay:

    Article Title:
    Article Snippet: The CD (J-720; JASCO, Tokyo, Japan) spectrum analysis did not show significant differences in rec-a2LN+ mutant proteins. .. The cell attachment assay using synthetic peptides and recombinant proteins was performed in 96-well plates (Thermo Fisher Scientific). .. 96-well plates were coated with various amounts of proteins in 50 μl of PBS overnight at 4 °C.

    Article Title: Mixed Fibronectin-Derived Peptides Conjugated to a Chitosan Matrix Effectively Promotes Biological Activities through Integrins, α4β1, α5β1, αvβ3, and Syndecan
    Article Snippet: Paragraph title: Cell attachment assay ... The peptide-conjugated chitosan matrix was prepared in 96-well plates (Nalge Nunc, Inc., Rochester, NY) as described previously.

    Article Title:
    Article Snippet: 5 m m of peptide solution was added to the wells and incubated for 2 h. Then the plates were used for the cell attachment assay as described above. .. For inhibition of cell attachment to peptide-coated plates and to peptide-chitosan membranes, 96-well plates were prepared as described above. .. The cells were preincubated for 15 min at 37 °C in the presence of either 10 μg/ml heparin (Sigma-Aldrich), 5 m m EDTA (Wako Pure Chemicals), 10 μg/ml various cell adhesive peptides, and 10 or 30 μg/ml of integrin antibodies.

    Article Title:
    Article Snippet: Each peptide was assayed using HDFs and C2C12 cells. .. Cell attachment assays using peptide-chitosan membranes were performed on 96-well plates as described previously ( , ). .. After 50 μl/well of maleimidobenzoyloxy-chitosan solution was added and dried, the plates were washed with 1% NaHCO3 and then washed with PBS.

    High Content Screening:

    Article Title: MiR-16 regulates the pro-tumorigenic potential of lung fibroblasts through the inhibition of HGF production in an FGFR-1- and MEK1-dependent manner
    Article Snippet: For the high-throughput screening, at day 1, CAF154-hTERT fibroblasts were reverse transfected with a library of human miRNA mimics composed of 988 mature miRNAs arrayed on 96-well plates (875 unique sequences, miRBase v.13.0, miRIDIAN technology, Dharmacon). .. A549-green fluorescent protein (GFP) cells (3500 cells/well) were seeded at day 3 (48 h after fibroblast transfection) and co-cultured for further 48 h. The experiment was stopped by fixing the cells with 4% paraformaldehyde, and nuclei were counterstained with Hoechst 33342.

    FACS:

    Article Title: The lysophosphatidylserine receptor GPR174 constrains regulatory T cell development and function
    Article Snippet: For cell surface staining, empirically determined dilutions of primary mAbs were used to stain single cell suspensions on ice for 20 min in FACS buffer (PBS with 2% FBS, 0.1% NaN3 , and 1 µM EDTA). .. Intracellular staining to detect Foxp3 expression was performed according to the manufacturer’s instructions, using the manufacturer’s protocol for Foxp3 staining in 96-well plates (eBioscience).

    Activated Clotting Time Assay:

    Article Title: Identification of Two Subgroups of Type I IFNs in Perciforme Fish Large Yellow Croaker Larimichthys crocea Provides Novel Insights into Function and Regulation of Fish Type I IFNs
    Article Snippet: The EPC cells (5 × 104 /well) were seeded in 96-well plates (Thermo Fisher Scientific) overnight and cotransfected with 100 ng of pGL3-IFNP plasmid or pGL3-Basic plasmid (control) and 2 ng pRL-TK plasmid using the Fugene® HD transfection reagent. .. The EPC cells (5 × 104 /well) were seeded in 96-well plates (Thermo Fisher Scientific) overnight and cotransfected with 100 ng of pGL3-IFNP plasmid or pGL3-Basic plasmid (control) and 2 ng pRL-TK plasmid using the Fugene® HD transfection reagent.

    Purification:

    Article Title: Uptake of HLA Alloantigens via CD89 and CD206 Does Not Enhance Antigen Presentation by Indirect Allorecognition
    Article Snippet: Monomers were purified by gel filtration HPLC, routinely tested, and validated. .. MaxiSorp flat-bottomed 96-well plates (Nunc, Thermo Scientific) were coated with 1 μ g of CD206-A/CD206 or CD89-A/CD89 (diluted in PBS) and incubated for 2 hours at 37°C.

    Article Title: Anti-heparan Sulfate Peptides That Block Herpes Simplex Virus Infection in Vivo
    Article Snippet: A purified form of HS isolated from bovine kidney was purchased from Sigma. .. Both targets at a concentration of 10 μg/ml were used for overnight coating of wells of 96-well plates (Nalge Nunc International, Naperville, IL) in a humidifier chamber at 4 °C.

    Viability Assay:

    Article Title: Hypoxic 3D in vitro culture models reveal distinct resistance processes to TKIs in renal cancer cells
    Article Snippet: Paragraph title: AlamarBlue® viability assay and growth curves ... Cells were seeded on flat-bottom, 96-well plates (Thermo Fisher Scientific, Cat. No. 07-200-90) in the following concentrations to achieve their logarithmic phase of growth: HKCSCs—1000 cells/well, 769-P—1500 cells/well, Caki-1—2000 cells/well.

    Plasmid Preparation:

    Article Title: Identification of Two Subgroups of Type I IFNs in Perciforme Fish Large Yellow Croaker Larimichthys crocea Provides Novel Insights into Function and Regulation of Fish Type I IFNs
    Article Snippet: For luciferase assays, the recombinant plasmids were constructed by inserting the promoter regions of two IFN genes and a series of their respective deleted fragments into the dual luciferase reporter plasmid pGL3-Basic (pGL3-IFNPs, primers in Table S1 in Supplementary Material; Promega). .. The EPC cells (5 × 104 /well) were seeded in 96-well plates (Thermo Fisher Scientific) overnight and cotransfected with 100 ng of pGL3-IFNP plasmid or pGL3-Basic plasmid (control) and 2 ng pRL-TK plasmid using the Fugene® HD transfection reagent. .. After 48 h, the luciferase activity of total cell lysates was measured on a GloMax 20/20 luminometer (Promega) according to the Dual-Luciferase® Repoter Assay System (Promega).

    Software:

    Article Title: The lysophosphatidylserine receptor GPR174 constrains regulatory T cell development and function
    Article Snippet: Intracellular staining to detect Foxp3 expression was performed according to the manufacturer’s instructions, using the manufacturer’s protocol for Foxp3 staining in 96-well plates (eBioscience). .. Cells were analyzed using an LSR-II flow cytometer (BD) equipped with 405-, 488-, 552-, and 640-nm lasers.

    Selection:

    Article Title: Anti-heparan Sulfate Peptides That Block Herpes Simplex Virus Infection in Vivo
    Article Snippet: Paragraph title: Selection of Phages against HS and 3-OS HS by Library Panning ... Both targets at a concentration of 10 μg/ml were used for overnight coating of wells of 96-well plates (Nalge Nunc International, Naperville, IL) in a humidifier chamber at 4 °C.

    Concentration Assay:

    Article Title: Anti-heparan Sulfate Peptides That Block Herpes Simplex Virus Infection in Vivo
    Article Snippet: Screening of the phage display library was accomplished by an affinity selection (or bio-panning) process during which phage populations were selected for their ability to bind HS and 3-OS HS (modified by 3-OST-3). .. Both targets at a concentration of 10 μg/ml were used for overnight coating of wells of 96-well plates (Nalge Nunc International, Naperville, IL) in a humidifier chamber at 4 °C. .. The following day, the plates were blocked for 1 h at room temperature with 5 mg/ml bovine serum albumin in 0.1 m NaHCO3 , pH 8.6, buffer.

    High Throughput Screening Assay:

    Article Title: MiR-16 regulates the pro-tumorigenic potential of lung fibroblasts through the inhibition of HGF production in an FGFR-1- and MEK1-dependent manner
    Article Snippet: As the large-scale screening experiment was not feasible with primary CAFs due to the limited cell number, fibroblasts (CAFs, AFs, and NFs) from different patients were transduced with retroviral particles to stably express human TERT (hTERT) and immortalize cells (see above). .. For the high-throughput screening, at day 1, CAF154-hTERT fibroblasts were reverse transfected with a library of human miRNA mimics composed of 988 mature miRNAs arrayed on 96-well plates (875 unique sequences, miRBase v.13.0, miRIDIAN technology, Dharmacon). .. Briefly, 15 μl miRNA (500 nM) was spotted per well, and a mix of 35 μl Opti-MEM containing RNAiMAX (Thermo Fisher Scientific) was added.

    Activity Assay:

    Article Title: Identification of Two Subgroups of Type I IFNs in Perciforme Fish Large Yellow Croaker Larimichthys crocea Provides Novel Insights into Function and Regulation of Fish Type I IFNs
    Article Snippet: Paragraph title: Luciferase Activity Assay ... The EPC cells (5 × 104 /well) were seeded in 96-well plates (Thermo Fisher Scientific) overnight and cotransfected with 100 ng of pGL3-IFNP plasmid or pGL3-Basic plasmid (control) and 2 ng pRL-TK plasmid using the Fugene® HD transfection reagent.

    Cell Viability Assay:

    Article Title: hvTRA, a novel TRAIL receptor agonist, induces apoptosis and sustained growth retardation in melanoma
    Article Snippet: The cells were seeded in 96-well plates (Thermo Fisher Scientific, Rockford, IL, USA). .. The cells were seeded in 96-well plates (Thermo Fisher Scientific, Rockford, IL, USA).

    other:

    Article Title:
    Article Snippet: Neurite outgrowth was performed in 96-well plates and assessed as described previously ( ).

    Article Title:
    Article Snippet: To evaluate the inhibitory effect of EDTA and heparin on cell adhesive peptides on the 96-well plates, A2-20 was conjugated with a chitosan membrane in the well.

    Article Title:
    Article Snippet: 96-well plates were coated with various amounts of proteins in 50 μl of PBS overnight at 4 °C.

    Article Title: A human monoclonal antibody against small envelope protein of hepatitis B virus with potent neutralization effect
    Article Snippet: Cells were grown in 96-well plates in William's E medium supplemented with cortisone and insulin.

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  • 99
    Thermo Fisher 96 well elisa plates
    Dose dependent detection of cleaved SNAP-25 specific for ( a ) Botulinum toxins (BoNT)/A and ( b ) BoNT/E toxins. SiMa cells were differentiated for 3 days on <t>96-well</t> tissue culture plates and treated with either purified BoNT/A or BoNT/E toxins in a range of concentrations between 1–1280 LD50/mL (~5 pg/mL to ~5 ng/mL). After 48 h exposure, cells were lysed and subjected to toxin specific capture <t>ELISA</t> for detection of either BoNT/A ( a ) or BoNT/E ( b ) cleaved SNAP-25. Dotted line indicates controls where cells were not exposed to toxins. Results are from one typical assay performed on at least three independent occasions and each data set is a mean from four individual wells ±SD. ( c ) Schematic overview of capture ELISA for BoNT/A and BoNT/E: BoNT/A cleaves SNAP-25 between amino acids 197 and 198 and the cleavage product is captured using a specific neo-epitope antibody raised against a peptide corresponding to amino acids 190–197 of SNAP-25 (SNAP-25 190–197 ). BoNT/E cleaves SNAP-25 between amino acids 180 and 181 and the cleavage product is captured using a specific neo-epitope antibody raised against a peptide corresponding to amino acids 173–180 of SNAP-25 (SNAP-25 173–180 ). The captured cleavage product is then detected using two polyclonal detection antibodies that bind to two distinct sites, SNAP-25 1–57 and SNAP-25 111–157 .
    96 Well Elisa Plates, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 79 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/96 well elisa plates/product/Thermo Fisher
    Average 99 stars, based on 79 article reviews
    Price from $9.99 to $1999.99
    96 well elisa plates - by Bioz Stars, 2020-01
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    84
    Thermo Fisher 96 well polystyrene microtiter plates
    Characterization of the DGC PA3177. (A) E. coli BL21-pET3177 and the vector control strain BL21-pET23a were grown in LB broth supplemented with 0.4 mM IPTG for 5 h at 37°C followed by nucleotide extraction and quantification of intracellular c-di-GMP by LC-MS. Levels of c-di-GMP were normalized to the total protein content of the respective sample and compared to c-di-GMP levels in empty vector control strain BL21-pET23a ( n = 6). The chromatogram shows c-di-GMP peaks of one representative measurement. (B) Attachment of P. aeruginosa during PA3177 overexpression was evaluated by crystal violet staining. PAO1-pJN3177 and empty vector control strain PAO1-pJN105 were incubated in <t>96-well</t> <t>microtiter</t> plates for 2 h at 37°C followed by biomass quantification. For recombinant gene expression, BM2 was supplemented with 0.1% (w/v) arabinose (+Ara). Cultures without arabinose (–Ara) served as negative controls. Experiments were carried out in triplicate, each with six wells per strain and condition ( n = 18). (C) Planktonic growth during 5 h at 37°C. (D) Overnight cultures of PA3177-overexpressing strain P. aeruginosa PA01-pJN3177 and vector control PAO1-pJN105 were diluted in LB broth supplemented with 0.1% (w/v) arabinose and grown for 5 h at 37°C followed by evaluation of swimming, swarming and twitching motility. Assays were carried out with three independent bacterial cultures and at least four agar plates per experiment ( n ≥ 12). In all experiments statistical significance was evaluated by the Mann-Whitney test ( ∗∗∗ p ≤ 0.001, ∗∗ p ≤ 0.01, n.s., not significant). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers.
    96 Well Polystyrene Microtiter Plates, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 84/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/96 well polystyrene microtiter plates/product/Thermo Fisher
    Average 84 stars, based on 5 article reviews
    Price from $9.99 to $1999.99
    96 well polystyrene microtiter plates - by Bioz Stars, 2020-01
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    75
    Thermo Fisher 96 well immuno micro titer plates
    Competitive ELISA titration curve of anti-Hn-33/A competing with Hn-33/A or BoNT/A complex. The 96 well <t>immuno-plate</t> was coated with 1ng/µL of BoNT/A complex, followed by a 2-fold serial dilution of BoNT/A complex or rHn-33/A with 2X titer of
    96 Well Immuno Micro Titer Plates, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 75/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/96 well immuno micro titer plates/product/Thermo Fisher
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    96 well immuno micro titer plates - by Bioz Stars, 2020-01
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    Image Search Results


    Dose dependent detection of cleaved SNAP-25 specific for ( a ) Botulinum toxins (BoNT)/A and ( b ) BoNT/E toxins. SiMa cells were differentiated for 3 days on 96-well tissue culture plates and treated with either purified BoNT/A or BoNT/E toxins in a range of concentrations between 1–1280 LD50/mL (~5 pg/mL to ~5 ng/mL). After 48 h exposure, cells were lysed and subjected to toxin specific capture ELISA for detection of either BoNT/A ( a ) or BoNT/E ( b ) cleaved SNAP-25. Dotted line indicates controls where cells were not exposed to toxins. Results are from one typical assay performed on at least three independent occasions and each data set is a mean from four individual wells ±SD. ( c ) Schematic overview of capture ELISA for BoNT/A and BoNT/E: BoNT/A cleaves SNAP-25 between amino acids 197 and 198 and the cleavage product is captured using a specific neo-epitope antibody raised against a peptide corresponding to amino acids 190–197 of SNAP-25 (SNAP-25 190–197 ). BoNT/E cleaves SNAP-25 between amino acids 180 and 181 and the cleavage product is captured using a specific neo-epitope antibody raised against a peptide corresponding to amino acids 173–180 of SNAP-25 (SNAP-25 173–180 ). The captured cleavage product is then detected using two polyclonal detection antibodies that bind to two distinct sites, SNAP-25 1–57 and SNAP-25 111–157 .

    Journal: Toxins

    Article Title: SiMa Cells for a Serotype Specific and Sensitive Cell-Based Neutralization Test for Botulinum Toxin A and E

    doi: 10.3390/toxins9070230

    Figure Lengend Snippet: Dose dependent detection of cleaved SNAP-25 specific for ( a ) Botulinum toxins (BoNT)/A and ( b ) BoNT/E toxins. SiMa cells were differentiated for 3 days on 96-well tissue culture plates and treated with either purified BoNT/A or BoNT/E toxins in a range of concentrations between 1–1280 LD50/mL (~5 pg/mL to ~5 ng/mL). After 48 h exposure, cells were lysed and subjected to toxin specific capture ELISA for detection of either BoNT/A ( a ) or BoNT/E ( b ) cleaved SNAP-25. Dotted line indicates controls where cells were not exposed to toxins. Results are from one typical assay performed on at least three independent occasions and each data set is a mean from four individual wells ±SD. ( c ) Schematic overview of capture ELISA for BoNT/A and BoNT/E: BoNT/A cleaves SNAP-25 between amino acids 197 and 198 and the cleavage product is captured using a specific neo-epitope antibody raised against a peptide corresponding to amino acids 190–197 of SNAP-25 (SNAP-25 190–197 ). BoNT/E cleaves SNAP-25 between amino acids 180 and 181 and the cleavage product is captured using a specific neo-epitope antibody raised against a peptide corresponding to amino acids 173–180 of SNAP-25 (SNAP-25 173–180 ). The captured cleavage product is then detected using two polyclonal detection antibodies that bind to two distinct sites, SNAP-25 1–57 and SNAP-25 111–157 .

    Article Snippet: For the ELISA, 96-well ELISA plates (Nunc, Maxisorb, Thermo Fisher Scientific, Hemel Hempstead, UK) were coated with 50 µL per well of 5–10 µg/mL of affinity purified capture antibody against SNAP-25190–197 (BoNT/A assay) or SNAP-25173–180 (BoNT/E assay) in carbonate coating buffer (pH 9.6), overnight at 4 °C.

    Techniques: Purification, Enzyme-linked Immunosorbent Assay

    Dose dependent inhibition of BoNT/A cleavage of SNAP-25 from SiMa cells by ( a ) reference polyclonal and ( b ) humanized recombinant monoclonal antibodies against BoNT/A. SiMa cells were differentiated for 3 days on 96-well tissue culture plates and treated with a mixture of purified BoNT/A toxin (200 LD50/mL or 40LD50 per well) and ( a ) reference antitoxin for BoNT/A (NIBSC product code 59/021) in the range of concentrations between 0.1 IU and 0.1 mIU or ( b ) humanized monoclonal antibodies targeting heavy chain (HC) or the light chain (LC) of BoNT/A [ 12 ]. After 48 h exposure to the corresponding mixtures, cells were lysed and subjected to capture ELISA for detection of BoNT/A cleaved SNAP-25. Results are from one representative assay where each data set is the mean from two individually treated wells ±SD. Reference antitoxin for BoNT/A (NIBSC product code 59/021) was also included as a negative control in the absence of BoNT/A ( a ).

    Journal: Toxins

    Article Title: SiMa Cells for a Serotype Specific and Sensitive Cell-Based Neutralization Test for Botulinum Toxin A and E

    doi: 10.3390/toxins9070230

    Figure Lengend Snippet: Dose dependent inhibition of BoNT/A cleavage of SNAP-25 from SiMa cells by ( a ) reference polyclonal and ( b ) humanized recombinant monoclonal antibodies against BoNT/A. SiMa cells were differentiated for 3 days on 96-well tissue culture plates and treated with a mixture of purified BoNT/A toxin (200 LD50/mL or 40LD50 per well) and ( a ) reference antitoxin for BoNT/A (NIBSC product code 59/021) in the range of concentrations between 0.1 IU and 0.1 mIU or ( b ) humanized monoclonal antibodies targeting heavy chain (HC) or the light chain (LC) of BoNT/A [ 12 ]. After 48 h exposure to the corresponding mixtures, cells were lysed and subjected to capture ELISA for detection of BoNT/A cleaved SNAP-25. Results are from one representative assay where each data set is the mean from two individually treated wells ±SD. Reference antitoxin for BoNT/A (NIBSC product code 59/021) was also included as a negative control in the absence of BoNT/A ( a ).

    Article Snippet: For the ELISA, 96-well ELISA plates (Nunc, Maxisorb, Thermo Fisher Scientific, Hemel Hempstead, UK) were coated with 50 µL per well of 5–10 µg/mL of affinity purified capture antibody against SNAP-25190–197 (BoNT/A assay) or SNAP-25173–180 (BoNT/E assay) in carbonate coating buffer (pH 9.6), overnight at 4 °C.

    Techniques: Inhibition, Recombinant, Purification, Liquid Chromatography, Enzyme-linked Immunosorbent Assay, Negative Control

    Dose dependent inhibition of BoNT/E cleavage of SNAP-25 from SiMa cells by polyclonal type E antitoxins. SiMa cells were differentiated for 3 days on 96-well tissue culture plates and treated with a mixture of purified BoNT/E toxin (200 LD50/mL or 40 LD50 per well) and reference antitoxin for BoNT/E (NIBSC product code 02/318), or two separate batches of polyclonal trivalent antitoxin (#079012A and #081021A, with assumed potency of > 50 IU/mL for antitoxin type E) diluted in the range between 0.1 IU/mL and 0.05 mIU/mL. Reference antitoxin for BoNT/A (NIBSC product code 59/021) was included as a negative control. After 48 h exposure to the corresponding mixtures, cells were lysed and subjected to capture ELISA for detection of BoNT/E cleaved SNAP-25. Results are from a single experiment where each data set is the mean of two individually treated wells ±SDs. SiMa cells were also treated with the antitoxins in the absence of BoNT/E and no signal was observed in the capture ELISA ( Supplementary Materials Figure S1 ).

    Journal: Toxins

    Article Title: SiMa Cells for a Serotype Specific and Sensitive Cell-Based Neutralization Test for Botulinum Toxin A and E

    doi: 10.3390/toxins9070230

    Figure Lengend Snippet: Dose dependent inhibition of BoNT/E cleavage of SNAP-25 from SiMa cells by polyclonal type E antitoxins. SiMa cells were differentiated for 3 days on 96-well tissue culture plates and treated with a mixture of purified BoNT/E toxin (200 LD50/mL or 40 LD50 per well) and reference antitoxin for BoNT/E (NIBSC product code 02/318), or two separate batches of polyclonal trivalent antitoxin (#079012A and #081021A, with assumed potency of > 50 IU/mL for antitoxin type E) diluted in the range between 0.1 IU/mL and 0.05 mIU/mL. Reference antitoxin for BoNT/A (NIBSC product code 59/021) was included as a negative control. After 48 h exposure to the corresponding mixtures, cells were lysed and subjected to capture ELISA for detection of BoNT/E cleaved SNAP-25. Results are from a single experiment where each data set is the mean of two individually treated wells ±SDs. SiMa cells were also treated with the antitoxins in the absence of BoNT/E and no signal was observed in the capture ELISA ( Supplementary Materials Figure S1 ).

    Article Snippet: For the ELISA, 96-well ELISA plates (Nunc, Maxisorb, Thermo Fisher Scientific, Hemel Hempstead, UK) were coated with 50 µL per well of 5–10 µg/mL of affinity purified capture antibody against SNAP-25190–197 (BoNT/A assay) or SNAP-25173–180 (BoNT/E assay) in carbonate coating buffer (pH 9.6), overnight at 4 °C.

    Techniques: Inhibition, Purification, Negative Control, Enzyme-linked Immunosorbent Assay

    Specificity of SiMa cell toxin neutralization assay for BoNT/A. SiMa cells were differentiated for 3 days on 96-well tissue culture plates and treated with a mixture of purified BoNT/A toxin (200 LD50/mL or 40 LD50 per well) and ( a ) a reference antitoxin for BoNT/E (NIBSC product code 02/318) in the range between 1 IU and 0.1 mIU or ( b ) humanized recombinant monoclonal antibody targeting the LC of BoNT/E (ELC18) [ 13 ]. After 48 h exposure to the corresponding mixtures, cells were lysed and subjected to capture ELISA for detection of BoNT/A cleaved SNAP-25. Results are from a representative experiment where each data set is the mean from two individually treated wells ±SD. Reference antitoxin for BoNT/E (NIBSC product code 02/318) and humanized antibody ELC18 were also incubated with SiMa cells in the absence of BoNT/A as negative controls and were from a single cell reading per dilution.

    Journal: Toxins

    Article Title: SiMa Cells for a Serotype Specific and Sensitive Cell-Based Neutralization Test for Botulinum Toxin A and E

    doi: 10.3390/toxins9070230

    Figure Lengend Snippet: Specificity of SiMa cell toxin neutralization assay for BoNT/A. SiMa cells were differentiated for 3 days on 96-well tissue culture plates and treated with a mixture of purified BoNT/A toxin (200 LD50/mL or 40 LD50 per well) and ( a ) a reference antitoxin for BoNT/E (NIBSC product code 02/318) in the range between 1 IU and 0.1 mIU or ( b ) humanized recombinant monoclonal antibody targeting the LC of BoNT/E (ELC18) [ 13 ]. After 48 h exposure to the corresponding mixtures, cells were lysed and subjected to capture ELISA for detection of BoNT/A cleaved SNAP-25. Results are from a representative experiment where each data set is the mean from two individually treated wells ±SD. Reference antitoxin for BoNT/E (NIBSC product code 02/318) and humanized antibody ELC18 were also incubated with SiMa cells in the absence of BoNT/A as negative controls and were from a single cell reading per dilution.

    Article Snippet: For the ELISA, 96-well ELISA plates (Nunc, Maxisorb, Thermo Fisher Scientific, Hemel Hempstead, UK) were coated with 50 µL per well of 5–10 µg/mL of affinity purified capture antibody against SNAP-25190–197 (BoNT/A assay) or SNAP-25173–180 (BoNT/E assay) in carbonate coating buffer (pH 9.6), overnight at 4 °C.

    Techniques: Neutralization, Purification, Recombinant, Liquid Chromatography, Enzyme-linked Immunosorbent Assay, Incubation

    Effect of alanine substitutions on A2-8 cell attachment activity. 96-well plates were coated with various amounts of 12 Ala-substituted A2-8 peptides () and examined for cell attachment activity using HDFs. A–C , HDFs were added to the wells

    Journal:

    Article Title:

    doi: 10.1074/jbc.M112.348151

    Figure Lengend Snippet: Effect of alanine substitutions on A2-8 cell attachment activity. 96-well plates were coated with various amounts of 12 Ala-substituted A2-8 peptides () and examined for cell attachment activity using HDFs. A–C , HDFs were added to the wells

    Article Snippet: The cell attachment assay using synthetic peptides and recombinant proteins was performed in 96-well plates (Thermo Fisher Scientific).

    Techniques: Cell Attachment Assay, Activity Assay

    Effect of 9 active peptides from short arm region and various integrin-binding peptides on the attachment of HDFs to rec-a2N. 96-well plates were coated with 10 μg/well of rec-a2N proteins. A , various amount of cell adhesive peptides from the

    Journal:

    Article Title:

    doi: 10.1074/jbc.M112.348151

    Figure Lengend Snippet: Effect of 9 active peptides from short arm region and various integrin-binding peptides on the attachment of HDFs to rec-a2N. 96-well plates were coated with 10 μg/well of rec-a2N proteins. A , various amount of cell adhesive peptides from the

    Article Snippet: The cell attachment assay using synthetic peptides and recombinant proteins was performed in 96-well plates (Thermo Fisher Scientific).

    Techniques: Binding Assay

    Cell attachment to the peptide-coated plates and peptide-Sepharose beads. A and B , 96-well plates were coated with various amounts of synthetic peptides and examined for cell attachment activity using HDFs. The cells were added to the wells for 1 h. After

    Journal:

    Article Title:

    doi: 10.1074/jbc.M112.348151

    Figure Lengend Snippet: Cell attachment to the peptide-coated plates and peptide-Sepharose beads. A and B , 96-well plates were coated with various amounts of synthetic peptides and examined for cell attachment activity using HDFs. The cells were added to the wells for 1 h. After

    Article Snippet: The cell attachment assay using synthetic peptides and recombinant proteins was performed in 96-well plates (Thermo Fisher Scientific).

    Techniques: Cell Attachment Assay, Activity Assay

    Effect of anti-integrin antibodies on cell attachment to A2-8 and effect of various peptides on cell attachment to rec-a2LN. A , 96-well plates were coated with 10 μg/well of A2-8. 10 and/or 30 μg/ml of anti-integrin antibodies were added

    Journal:

    Article Title:

    doi: 10.1074/jbc.M112.348151

    Figure Lengend Snippet: Effect of anti-integrin antibodies on cell attachment to A2-8 and effect of various peptides on cell attachment to rec-a2LN. A , 96-well plates were coated with 10 μg/well of A2-8. 10 and/or 30 μg/ml of anti-integrin antibodies were added

    Article Snippet: The cell attachment assay using synthetic peptides and recombinant proteins was performed in 96-well plates (Thermo Fisher Scientific).

    Techniques: Cell Attachment Assay

    Neurite outgrowth of PC12 cells on the peptide-coated plates. A , 10 μg/well of peptides were coated on 96-well plates. PC12 cells (3.0 × 10 3 cells/well) were seeded in the wells and incubated for 24 h. After the cells were fixed and stained,

    Journal:

    Article Title:

    doi: 10.1074/jbc.M112.348151

    Figure Lengend Snippet: Neurite outgrowth of PC12 cells on the peptide-coated plates. A , 10 μg/well of peptides were coated on 96-well plates. PC12 cells (3.0 × 10 3 cells/well) were seeded in the wells and incubated for 24 h. After the cells were fixed and stained,

    Article Snippet: The cell attachment assay using synthetic peptides and recombinant proteins was performed in 96-well plates (Thermo Fisher Scientific).

    Techniques: Incubation, Staining

    Characterization of the DGC PA3177. (A) E. coli BL21-pET3177 and the vector control strain BL21-pET23a were grown in LB broth supplemented with 0.4 mM IPTG for 5 h at 37°C followed by nucleotide extraction and quantification of intracellular c-di-GMP by LC-MS. Levels of c-di-GMP were normalized to the total protein content of the respective sample and compared to c-di-GMP levels in empty vector control strain BL21-pET23a ( n = 6). The chromatogram shows c-di-GMP peaks of one representative measurement. (B) Attachment of P. aeruginosa during PA3177 overexpression was evaluated by crystal violet staining. PAO1-pJN3177 and empty vector control strain PAO1-pJN105 were incubated in 96-well microtiter plates for 2 h at 37°C followed by biomass quantification. For recombinant gene expression, BM2 was supplemented with 0.1% (w/v) arabinose (+Ara). Cultures without arabinose (–Ara) served as negative controls. Experiments were carried out in triplicate, each with six wells per strain and condition ( n = 18). (C) Planktonic growth during 5 h at 37°C. (D) Overnight cultures of PA3177-overexpressing strain P. aeruginosa PA01-pJN3177 and vector control PAO1-pJN105 were diluted in LB broth supplemented with 0.1% (w/v) arabinose and grown for 5 h at 37°C followed by evaluation of swimming, swarming and twitching motility. Assays were carried out with three independent bacterial cultures and at least four agar plates per experiment ( n ≥ 12). In all experiments statistical significance was evaluated by the Mann-Whitney test ( ∗∗∗ p ≤ 0.001, ∗∗ p ≤ 0.01, n.s., not significant). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers.

    Journal: Frontiers in Microbiology

    Article Title: The Oxidative Stress Agent Hypochlorite Stimulates c-di-GMP Synthesis and Biofilm Formation in Pseudomonas aeruginosa

    doi: 10.3389/fmicb.2017.02311

    Figure Lengend Snippet: Characterization of the DGC PA3177. (A) E. coli BL21-pET3177 and the vector control strain BL21-pET23a were grown in LB broth supplemented with 0.4 mM IPTG for 5 h at 37°C followed by nucleotide extraction and quantification of intracellular c-di-GMP by LC-MS. Levels of c-di-GMP were normalized to the total protein content of the respective sample and compared to c-di-GMP levels in empty vector control strain BL21-pET23a ( n = 6). The chromatogram shows c-di-GMP peaks of one representative measurement. (B) Attachment of P. aeruginosa during PA3177 overexpression was evaluated by crystal violet staining. PAO1-pJN3177 and empty vector control strain PAO1-pJN105 were incubated in 96-well microtiter plates for 2 h at 37°C followed by biomass quantification. For recombinant gene expression, BM2 was supplemented with 0.1% (w/v) arabinose (+Ara). Cultures without arabinose (–Ara) served as negative controls. Experiments were carried out in triplicate, each with six wells per strain and condition ( n = 18). (C) Planktonic growth during 5 h at 37°C. (D) Overnight cultures of PA3177-overexpressing strain P. aeruginosa PA01-pJN3177 and vector control PAO1-pJN105 were diluted in LB broth supplemented with 0.1% (w/v) arabinose and grown for 5 h at 37°C followed by evaluation of swimming, swarming and twitching motility. Assays were carried out with three independent bacterial cultures and at least four agar plates per experiment ( n ≥ 12). In all experiments statistical significance was evaluated by the Mann-Whitney test ( ∗∗∗ p ≤ 0.001, ∗∗ p ≤ 0.01, n.s., not significant). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers.

    Article Snippet: Attachment assays in 96-well polystyrene microtiter plates (Nunc, Thermo Fisher Scientific, St. Leon-Rot, Germany) were performed as previously described ( ) with the following minor modifications.

    Techniques: Plasmid Preparation, Liquid Chromatography, Mass Spectrometry, Over Expression, Staining, Incubation, Recombinant, Expressing, Acetylene Reduction Assay, MANN-WHITNEY

    Implication of c-di-GMP in the response of P. aeruginosa PAO1 to NaClO. (A) P. aeruginosa PAO1 was incubated with NaClO (8 μg/ml, OD 600 = 1.0, BM2) for 1h followed by nucleotide extraction and quantification of intracellular c-di-GMP by LC-MS. Levels of c-di-GMP were normalized to the total protein content of the respective sample and compared to c-di-GMP levels in untreated controls ( n = 6). The chromatogram shows c-di-GMP peaks of one representative measurement. (B) Attachment of P. aeruginosa in response to NaClO during overexpression of the c-di-GMP-degrading PDE PA2133 was evaluated by crystal violet staining. PAO1-pJN2133 and the vector control strain PAO1-pJN105 were incubated in 96-well microtiter plates either in the presence or in the absence of NaClO (2 μg/ml) for 2 h at 37°C followed by the quantification of biofilm biomass. BM2 was supplemented with 0.1% (w/v) arabinose to induce recombinant gene expression. Experiments were repeated four times, each with six wells per strain and condition ( n = 24). Absorbance at 595nm (A 595 ) was determined and obtained values for NaClO-treated samples were normalized against the A 595 of respective untreated controls (=relative biomass). Statistical significance was evaluated by the Mann–Whitney test ( ∗∗∗ p ≤ 0.001, ∗∗ p ≤ 0.01, n.s., not significant). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers.

    Journal: Frontiers in Microbiology

    Article Title: The Oxidative Stress Agent Hypochlorite Stimulates c-di-GMP Synthesis and Biofilm Formation in Pseudomonas aeruginosa

    doi: 10.3389/fmicb.2017.02311

    Figure Lengend Snippet: Implication of c-di-GMP in the response of P. aeruginosa PAO1 to NaClO. (A) P. aeruginosa PAO1 was incubated with NaClO (8 μg/ml, OD 600 = 1.0, BM2) for 1h followed by nucleotide extraction and quantification of intracellular c-di-GMP by LC-MS. Levels of c-di-GMP were normalized to the total protein content of the respective sample and compared to c-di-GMP levels in untreated controls ( n = 6). The chromatogram shows c-di-GMP peaks of one representative measurement. (B) Attachment of P. aeruginosa in response to NaClO during overexpression of the c-di-GMP-degrading PDE PA2133 was evaluated by crystal violet staining. PAO1-pJN2133 and the vector control strain PAO1-pJN105 were incubated in 96-well microtiter plates either in the presence or in the absence of NaClO (2 μg/ml) for 2 h at 37°C followed by the quantification of biofilm biomass. BM2 was supplemented with 0.1% (w/v) arabinose to induce recombinant gene expression. Experiments were repeated four times, each with six wells per strain and condition ( n = 24). Absorbance at 595nm (A 595 ) was determined and obtained values for NaClO-treated samples were normalized against the A 595 of respective untreated controls (=relative biomass). Statistical significance was evaluated by the Mann–Whitney test ( ∗∗∗ p ≤ 0.001, ∗∗ p ≤ 0.01, n.s., not significant). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers.

    Article Snippet: Attachment assays in 96-well polystyrene microtiter plates (Nunc, Thermo Fisher Scientific, St. Leon-Rot, Germany) were performed as previously described ( ) with the following minor modifications.

    Techniques: Incubation, Liquid Chromatography, Mass Spectrometry, Over Expression, Staining, Plasmid Preparation, Recombinant, Expressing, MANN-WHITNEY

    Attachment and DGC expression in PAO1-PA3177Ω in response to NaClO. (A) Attachment of P. aeruginosa mutant strain PAO1-PA3177Ω and wildtype PAO1 after 2 h incubation with half-MIC concentrations of NaClO (2 μg/ml) compared to untreated controls was assayed in 96-well microtiter plates by crystal violet staining. To show alterations in attachment caused by NaClO, A 595 of treated samples was normalized to A 595 of untreated controls for each strain (=relative biomass). Statistical significance was evaluated by the Mann–Whitney test ( ∗∗∗ p ≤ 0.001, n ≥ 12). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers. (B) P. aeruginosa mutant strain PAO1-PA3177Ω was treated with 2 μg/ml NaClO and relative gene expression levels of genes encoding DGCs in comparison to untreated control cultures were analyzed by qRT-PCR. The figure shows mean averages and standard deviations calculated from three experiments, each analyzed in duplicate ( n = 6). Ct values were normalized against the expression of housekeeping genes rpoD and fabD . The red line corresponds to a relative gene expression level of 1, which means no change in gene expression. Blue bars indicate relative gene expression values ≥ 3.

    Journal: Frontiers in Microbiology

    Article Title: The Oxidative Stress Agent Hypochlorite Stimulates c-di-GMP Synthesis and Biofilm Formation in Pseudomonas aeruginosa

    doi: 10.3389/fmicb.2017.02311

    Figure Lengend Snippet: Attachment and DGC expression in PAO1-PA3177Ω in response to NaClO. (A) Attachment of P. aeruginosa mutant strain PAO1-PA3177Ω and wildtype PAO1 after 2 h incubation with half-MIC concentrations of NaClO (2 μg/ml) compared to untreated controls was assayed in 96-well microtiter plates by crystal violet staining. To show alterations in attachment caused by NaClO, A 595 of treated samples was normalized to A 595 of untreated controls for each strain (=relative biomass). Statistical significance was evaluated by the Mann–Whitney test ( ∗∗∗ p ≤ 0.001, n ≥ 12). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers. (B) P. aeruginosa mutant strain PAO1-PA3177Ω was treated with 2 μg/ml NaClO and relative gene expression levels of genes encoding DGCs in comparison to untreated control cultures were analyzed by qRT-PCR. The figure shows mean averages and standard deviations calculated from three experiments, each analyzed in duplicate ( n = 6). Ct values were normalized against the expression of housekeeping genes rpoD and fabD . The red line corresponds to a relative gene expression level of 1, which means no change in gene expression. Blue bars indicate relative gene expression values ≥ 3.

    Article Snippet: Attachment assays in 96-well polystyrene microtiter plates (Nunc, Thermo Fisher Scientific, St. Leon-Rot, Germany) were performed as previously described ( ) with the following minor modifications.

    Techniques: Expressing, Mutagenesis, Incubation, Staining, MANN-WHITNEY, Quantitative RT-PCR

    Attachment assays with PAO1 mutants. (A) Attachment of P. aeruginosa PAO1 wildtype (WT) or PAO1 mutant strains during overexpression of PA3177 was evaluated by crystal violet staining. Bacteria were incubated in 96-well microtiter plates for 2 h at 37°C prior to biomass quantification. For recombinant gene expression, L was supplemented with 0.1% (w/v) arabinose (+). Cultures without arabinose (-) served as negative controls. (B) Attachment of P. aeruginosa PAO1 WT or PAO1 deletion mutants during NaClO treatment (2 μg/ml in BM2, 2 h) was evaluated by crystal violet staining. All experiments were carried out in triplicate, each with six wells per strain and condition ( n = 18). Statistical significance was evaluated by the Mann–Whitney test ( ∗∗∗ p ≤ 0.001, ∗∗ p ≤ 0.01, n.s., not significant). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers.

    Journal: Frontiers in Microbiology

    Article Title: The Oxidative Stress Agent Hypochlorite Stimulates c-di-GMP Synthesis and Biofilm Formation in Pseudomonas aeruginosa

    doi: 10.3389/fmicb.2017.02311

    Figure Lengend Snippet: Attachment assays with PAO1 mutants. (A) Attachment of P. aeruginosa PAO1 wildtype (WT) or PAO1 mutant strains during overexpression of PA3177 was evaluated by crystal violet staining. Bacteria were incubated in 96-well microtiter plates for 2 h at 37°C prior to biomass quantification. For recombinant gene expression, L was supplemented with 0.1% (w/v) arabinose (+). Cultures without arabinose (-) served as negative controls. (B) Attachment of P. aeruginosa PAO1 WT or PAO1 deletion mutants during NaClO treatment (2 μg/ml in BM2, 2 h) was evaluated by crystal violet staining. All experiments were carried out in triplicate, each with six wells per strain and condition ( n = 18). Statistical significance was evaluated by the Mann–Whitney test ( ∗∗∗ p ≤ 0.001, ∗∗ p ≤ 0.01, n.s., not significant). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers.

    Article Snippet: Attachment assays in 96-well polystyrene microtiter plates (Nunc, Thermo Fisher Scientific, St. Leon-Rot, Germany) were performed as previously described ( ) with the following minor modifications.

    Techniques: Mutagenesis, Over Expression, Staining, Incubation, Recombinant, Expressing, MANN-WHITNEY

    Attachment of P. aeruginosa PAO1 in the presence of different oxidants. Attachment of P. aeruginosa PAO1 during 2 h incubation in the presence of sublethal NaClO concentrations (A–C) or half-MIC concentrations of oxidants NaClO (2 μg/ml), Ca(ClO) 2 (2 μg/ml), NH 2 Cl (equivalent to 2 μg/ml NaClO), H 2 O 2 (50 μg/ml) and paraquat (1 μg/ml) (D) . (A,B,D) Attachment was assayed in 96-well microtiter plates by crystal violet staining and subsequent measurement of A 595 . Obtained values for treated samples were normalized against A 595 of untreated controls (=relative biomass). Experiments were performed at least in triplicate, each with multiple wells per condition ( n ≥ 18). (C) P. aeruginosa PAO1 was incubated with sub-MIC concentrations of NaClO (2 μg/ml) in BM2 for 2 h in 50 ml reaction tubes containing glass microscope slides. Adhered bacteria were stained with the DNA-intercalating dye SYTO9 prior to visualization by fluorescence microscopy at 100× magnification (scale bar: 100 μm). Average surface coverages from 36 pictures per condition were calculated using ImageJ . Experiments were performed in triplicate. For all experiments, statistical significance was evaluated by the Mann-Whitney test ( ∗∗∗ p ≤ 0.001, n.s., not significant). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers.

    Journal: Frontiers in Microbiology

    Article Title: The Oxidative Stress Agent Hypochlorite Stimulates c-di-GMP Synthesis and Biofilm Formation in Pseudomonas aeruginosa

    doi: 10.3389/fmicb.2017.02311

    Figure Lengend Snippet: Attachment of P. aeruginosa PAO1 in the presence of different oxidants. Attachment of P. aeruginosa PAO1 during 2 h incubation in the presence of sublethal NaClO concentrations (A–C) or half-MIC concentrations of oxidants NaClO (2 μg/ml), Ca(ClO) 2 (2 μg/ml), NH 2 Cl (equivalent to 2 μg/ml NaClO), H 2 O 2 (50 μg/ml) and paraquat (1 μg/ml) (D) . (A,B,D) Attachment was assayed in 96-well microtiter plates by crystal violet staining and subsequent measurement of A 595 . Obtained values for treated samples were normalized against A 595 of untreated controls (=relative biomass). Experiments were performed at least in triplicate, each with multiple wells per condition ( n ≥ 18). (C) P. aeruginosa PAO1 was incubated with sub-MIC concentrations of NaClO (2 μg/ml) in BM2 for 2 h in 50 ml reaction tubes containing glass microscope slides. Adhered bacteria were stained with the DNA-intercalating dye SYTO9 prior to visualization by fluorescence microscopy at 100× magnification (scale bar: 100 μm). Average surface coverages from 36 pictures per condition were calculated using ImageJ . Experiments were performed in triplicate. For all experiments, statistical significance was evaluated by the Mann-Whitney test ( ∗∗∗ p ≤ 0.001, n.s., not significant). Boxes include median (thick horizontal line), 25th and 75th percentiles. Dots indicate extreme values considered as outliers.

    Article Snippet: Attachment assays in 96-well polystyrene microtiter plates (Nunc, Thermo Fisher Scientific, St. Leon-Rot, Germany) were performed as previously described ( ) with the following minor modifications.

    Techniques: Incubation, Staining, Microscopy, Fluorescence, MANN-WHITNEY

    Competitive ELISA titration curve of anti-Hn-33/A competing with Hn-33/A or BoNT/A complex. The 96 well immuno-plate was coated with 1ng/µL of BoNT/A complex, followed by a 2-fold serial dilution of BoNT/A complex or rHn-33/A with 2X titer of

    Journal:

    Article Title: Comparative Immunochemical Characteristics of Botulinum Neurotoxin Type A and its Associated Proteins

    doi: 10.1016/j.toxicon.2013.06.011

    Figure Lengend Snippet: Competitive ELISA titration curve of anti-Hn-33/A competing with Hn-33/A or BoNT/A complex. The 96 well immuno-plate was coated with 1ng/µL of BoNT/A complex, followed by a 2-fold serial dilution of BoNT/A complex or rHn-33/A with 2X titer of

    Article Snippet: 96-well immuno micro-titer plates (Thermo Fisher Scientific, Pittsburg, PA), dry milk, rabbit anti-BoNT/A complex IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-BoNT/A IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-Hn-33 IgG (Merdian Life Science, Inc., Saco, ME), goat anti-rabbit IgG alkaline phosphatase antibody (Sigma Aldrich, St. Louis, MO), anti-rabbit IgG-HRP conjugated secondary antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA), 2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) (Sigma Aldrich, St. Louis, MO), 3% hydrogen peroxide (H2 O2 ), Immun-Blot PVDF membrane (BioRad, Hercules, CA) and, 5-bromo-4-chloro-3 –indoyl phosphate (BCIP) and nitro-blue tetrazolium chloride (NBT) (BioRad, Hercules, CA).

    Techniques: Competitive ELISA, Titration, Serial Dilution

    Indirect ELISA titration curve of anti-BoNT/A complex binding to BoNT/A complex, BoNT/A, NAPs/A, and Hn-33/A. The 96 well immuno-plate was coated with 1ng/µL of corresponding antigen followed by a 2-fold serial dilution of anti-rabbit BoNT/A complex

    Journal:

    Article Title: Comparative Immunochemical Characteristics of Botulinum Neurotoxin Type A and its Associated Proteins

    doi: 10.1016/j.toxicon.2013.06.011

    Figure Lengend Snippet: Indirect ELISA titration curve of anti-BoNT/A complex binding to BoNT/A complex, BoNT/A, NAPs/A, and Hn-33/A. The 96 well immuno-plate was coated with 1ng/µL of corresponding antigen followed by a 2-fold serial dilution of anti-rabbit BoNT/A complex

    Article Snippet: 96-well immuno micro-titer plates (Thermo Fisher Scientific, Pittsburg, PA), dry milk, rabbit anti-BoNT/A complex IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-BoNT/A IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-Hn-33 IgG (Merdian Life Science, Inc., Saco, ME), goat anti-rabbit IgG alkaline phosphatase antibody (Sigma Aldrich, St. Louis, MO), anti-rabbit IgG-HRP conjugated secondary antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA), 2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) (Sigma Aldrich, St. Louis, MO), 3% hydrogen peroxide (H2 O2 ), Immun-Blot PVDF membrane (BioRad, Hercules, CA) and, 5-bromo-4-chloro-3 –indoyl phosphate (BCIP) and nitro-blue tetrazolium chloride (NBT) (BioRad, Hercules, CA).

    Techniques: Indirect ELISA, Titration, Binding Assay, Serial Dilution

    Indirect ELISA titration curve of anti-Hn-33/A binding to rHn-33/A and BoNT/A complex. The 96 well immuno-plate was coated with 1ng/µL of corresponding antigen followed by a 2-fold serial dilution of anti-rabbit Hn-33/A IgG and developed with

    Journal:

    Article Title: Comparative Immunochemical Characteristics of Botulinum Neurotoxin Type A and its Associated Proteins

    doi: 10.1016/j.toxicon.2013.06.011

    Figure Lengend Snippet: Indirect ELISA titration curve of anti-Hn-33/A binding to rHn-33/A and BoNT/A complex. The 96 well immuno-plate was coated with 1ng/µL of corresponding antigen followed by a 2-fold serial dilution of anti-rabbit Hn-33/A IgG and developed with

    Article Snippet: 96-well immuno micro-titer plates (Thermo Fisher Scientific, Pittsburg, PA), dry milk, rabbit anti-BoNT/A complex IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-BoNT/A IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-Hn-33 IgG (Merdian Life Science, Inc., Saco, ME), goat anti-rabbit IgG alkaline phosphatase antibody (Sigma Aldrich, St. Louis, MO), anti-rabbit IgG-HRP conjugated secondary antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA), 2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) (Sigma Aldrich, St. Louis, MO), 3% hydrogen peroxide (H2 O2 ), Immun-Blot PVDF membrane (BioRad, Hercules, CA) and, 5-bromo-4-chloro-3 –indoyl phosphate (BCIP) and nitro-blue tetrazolium chloride (NBT) (BioRad, Hercules, CA).

    Techniques: Indirect ELISA, Titration, Binding Assay, Serial Dilution

    Competitive ELISA titration curve of anti-BoNT/A complex competing with BoNT/A or BoNT/A complex. The 96 well immuno-plate was coated with 1ng/µL of BoNT/A, followed by a 2-fold serial dilution of BoNT/A or BoNT/A complex with 2X titer of anti-rabbit

    Journal:

    Article Title: Comparative Immunochemical Characteristics of Botulinum Neurotoxin Type A and its Associated Proteins

    doi: 10.1016/j.toxicon.2013.06.011

    Figure Lengend Snippet: Competitive ELISA titration curve of anti-BoNT/A complex competing with BoNT/A or BoNT/A complex. The 96 well immuno-plate was coated with 1ng/µL of BoNT/A, followed by a 2-fold serial dilution of BoNT/A or BoNT/A complex with 2X titer of anti-rabbit

    Article Snippet: 96-well immuno micro-titer plates (Thermo Fisher Scientific, Pittsburg, PA), dry milk, rabbit anti-BoNT/A complex IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-BoNT/A IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-Hn-33 IgG (Merdian Life Science, Inc., Saco, ME), goat anti-rabbit IgG alkaline phosphatase antibody (Sigma Aldrich, St. Louis, MO), anti-rabbit IgG-HRP conjugated secondary antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA), 2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) (Sigma Aldrich, St. Louis, MO), 3% hydrogen peroxide (H2 O2 ), Immun-Blot PVDF membrane (BioRad, Hercules, CA) and, 5-bromo-4-chloro-3 –indoyl phosphate (BCIP) and nitro-blue tetrazolium chloride (NBT) (BioRad, Hercules, CA).

    Techniques: Competitive ELISA, Titration, Serial Dilution

    Competitive ELISA titration curve of anti-Complex/A competing with Hn-33/A or BoNT/A Complex. The 96 well immuno-plate was coated with 1ng/µL of BoNT/A complex, followed by a 2-fold serial dilution of BoNT/A Complex or rHn-33/A with 2X titer of

    Journal:

    Article Title: Comparative Immunochemical Characteristics of Botulinum Neurotoxin Type A and its Associated Proteins

    doi: 10.1016/j.toxicon.2013.06.011

    Figure Lengend Snippet: Competitive ELISA titration curve of anti-Complex/A competing with Hn-33/A or BoNT/A Complex. The 96 well immuno-plate was coated with 1ng/µL of BoNT/A complex, followed by a 2-fold serial dilution of BoNT/A Complex or rHn-33/A with 2X titer of

    Article Snippet: 96-well immuno micro-titer plates (Thermo Fisher Scientific, Pittsburg, PA), dry milk, rabbit anti-BoNT/A complex IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-BoNT/A IgG (Merdian Life Science, Inc., Saco, ME), rabbit anti-Hn-33 IgG (Merdian Life Science, Inc., Saco, ME), goat anti-rabbit IgG alkaline phosphatase antibody (Sigma Aldrich, St. Louis, MO), anti-rabbit IgG-HRP conjugated secondary antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA), 2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) (Sigma Aldrich, St. Louis, MO), 3% hydrogen peroxide (H2 O2 ), Immun-Blot PVDF membrane (BioRad, Hercules, CA) and, 5-bromo-4-chloro-3 –indoyl phosphate (BCIP) and nitro-blue tetrazolium chloride (NBT) (BioRad, Hercules, CA).

    Techniques: Competitive ELISA, Titration, Serial Dilution