streptavidin horseradish peroxidase hrp conjugate  (Thermo Fisher)


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    Structured Review

    Thermo Fisher streptavidin horseradish peroxidase hrp conjugate
    Metabolic stability of the dimeric and monomeric forms of E-cadherin. (A) Surface proteins of AEcM cells grown in 5-cm-diameter dishes were biotinylated and then chased in regular media. The chase times (in hours) are indicated above the blots. Cells were immunoprecipitated with an excess of anti-myc antibody, and the immunoprecipitates were adjusted to 60 μl with SDS-polyacrylamide gel electrophoresis sample buffer. Five microliters of each immunoprecipitate was loaded. The blots were probed either with <t>streptavidin-HRP</t> (Str-HRP) or anti-myc antibody (Myc). Three different exposure times of the blot developed by Str-HRP are shown. After an exposure time of 10 s [Str-HRP (10)], only the Ec1M protein was visualized, allowing us to estimate the half-life of the monomeric E-cadherin. A longer, 60-s exposure [Str-HRP (60)] visualized coimmunoprecipitated endogenous cadherin (Ec). The blots at the bottom of panel A [Str-HRP (E)] show blots of different exposure times equilibrated on the Ec1M signals, which allowed us to demonstrate that the ratio of Ec1M to endogenous cadherin did not change during the chase periods. Panel B is identical to panel A except that cell lysates before anti-myc immunoprecipitation were subjected to sucrose gradient centrifugation either immediately after the biotinylation (blot 0) or after the 16-h chase (blot 16). The exposure time of blot 0 was shorter than that of blot 16, showing that during the 16-h chase the Ec1M/E-cadherin ratio did not change. Note that immunoprecipitation of Ec1M leads to coimmunoprecipitation of the endogenous E-cadherin (Ec) only in fractions 4 to 6. (C) Surface-biotinylated A-431 cells were dissociated by EGTA and either cocultured for additional 8 h with AEcM cells (lane 1) or cultivated separately and combined after lysis (lane 2). The latter served as a control, showing the absence of interactions between cadherin molecules in solution. The small black bars on the left of the blots indicate the positions of molecular mass markers of 116 and 97.4 kDa.
    Streptavidin Horseradish Peroxidase Hrp Conjugate, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/streptavidin horseradish peroxidase hrp conjugate/product/Thermo Fisher
    Average 90 stars, based on 11 article reviews
    Price from $9.99 to $1999.99
    streptavidin horseradish peroxidase hrp conjugate - by Bioz Stars, 2020-04
    90/100 stars

    Images

    1) Product Images from "Dynamic Interplay between Adhesive and Lateral E-Cadherin Dimers"

    Article Title: Dynamic Interplay between Adhesive and Lateral E-Cadherin Dimers

    Journal: Molecular and Cellular Biology

    doi: 10.1128/MCB.22.21.7449-7458.2002

    Metabolic stability of the dimeric and monomeric forms of E-cadherin. (A) Surface proteins of AEcM cells grown in 5-cm-diameter dishes were biotinylated and then chased in regular media. The chase times (in hours) are indicated above the blots. Cells were immunoprecipitated with an excess of anti-myc antibody, and the immunoprecipitates were adjusted to 60 μl with SDS-polyacrylamide gel electrophoresis sample buffer. Five microliters of each immunoprecipitate was loaded. The blots were probed either with streptavidin-HRP (Str-HRP) or anti-myc antibody (Myc). Three different exposure times of the blot developed by Str-HRP are shown. After an exposure time of 10 s [Str-HRP (10)], only the Ec1M protein was visualized, allowing us to estimate the half-life of the monomeric E-cadherin. A longer, 60-s exposure [Str-HRP (60)] visualized coimmunoprecipitated endogenous cadherin (Ec). The blots at the bottom of panel A [Str-HRP (E)] show blots of different exposure times equilibrated on the Ec1M signals, which allowed us to demonstrate that the ratio of Ec1M to endogenous cadherin did not change during the chase periods. Panel B is identical to panel A except that cell lysates before anti-myc immunoprecipitation were subjected to sucrose gradient centrifugation either immediately after the biotinylation (blot 0) or after the 16-h chase (blot 16). The exposure time of blot 0 was shorter than that of blot 16, showing that during the 16-h chase the Ec1M/E-cadherin ratio did not change. Note that immunoprecipitation of Ec1M leads to coimmunoprecipitation of the endogenous E-cadherin (Ec) only in fractions 4 to 6. (C) Surface-biotinylated A-431 cells were dissociated by EGTA and either cocultured for additional 8 h with AEcM cells (lane 1) or cultivated separately and combined after lysis (lane 2). The latter served as a control, showing the absence of interactions between cadherin molecules in solution. The small black bars on the left of the blots indicate the positions of molecular mass markers of 116 and 97.4 kDa.
    Figure Legend Snippet: Metabolic stability of the dimeric and monomeric forms of E-cadherin. (A) Surface proteins of AEcM cells grown in 5-cm-diameter dishes were biotinylated and then chased in regular media. The chase times (in hours) are indicated above the blots. Cells were immunoprecipitated with an excess of anti-myc antibody, and the immunoprecipitates were adjusted to 60 μl with SDS-polyacrylamide gel electrophoresis sample buffer. Five microliters of each immunoprecipitate was loaded. The blots were probed either with streptavidin-HRP (Str-HRP) or anti-myc antibody (Myc). Three different exposure times of the blot developed by Str-HRP are shown. After an exposure time of 10 s [Str-HRP (10)], only the Ec1M protein was visualized, allowing us to estimate the half-life of the monomeric E-cadherin. A longer, 60-s exposure [Str-HRP (60)] visualized coimmunoprecipitated endogenous cadherin (Ec). The blots at the bottom of panel A [Str-HRP (E)] show blots of different exposure times equilibrated on the Ec1M signals, which allowed us to demonstrate that the ratio of Ec1M to endogenous cadherin did not change during the chase periods. Panel B is identical to panel A except that cell lysates before anti-myc immunoprecipitation were subjected to sucrose gradient centrifugation either immediately after the biotinylation (blot 0) or after the 16-h chase (blot 16). The exposure time of blot 0 was shorter than that of blot 16, showing that during the 16-h chase the Ec1M/E-cadherin ratio did not change. Note that immunoprecipitation of Ec1M leads to coimmunoprecipitation of the endogenous E-cadherin (Ec) only in fractions 4 to 6. (C) Surface-biotinylated A-431 cells were dissociated by EGTA and either cocultured for additional 8 h with AEcM cells (lane 1) or cultivated separately and combined after lysis (lane 2). The latter served as a control, showing the absence of interactions between cadherin molecules in solution. The small black bars on the left of the blots indicate the positions of molecular mass markers of 116 and 97.4 kDa.

    Techniques Used: Immunoprecipitation, Polyacrylamide Gel Electrophoresis, Gradient Centrifugation, Lysis

    2) Product Images from "Platelet-Derived Short-Chain Polyphosphates Enhance the Inactivation of Tissue Factor Pathway Inhibitor by Activated Coagulation Factor XI"

    Article Title: Platelet-Derived Short-Chain Polyphosphates Enhance the Inactivation of Tissue Factor Pathway Inhibitor by Activated Coagulation Factor XI

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0165172

    Characterization of the interaction between polyphosphate and TFPIα. ( A ) 96-well plates were coated with 5 μg/ml TFPIα (○,●) or BSA (◻) and increasing concentrations of biotinylated-polyphosphate (bio-polyP) (◻, ○,●) was added to selected wells. Selected experiments were done in the presence of 100 μM SCP (●). Binding was detected with streptavidin-HRP. Data are mean ± SE (n = 3).
    Figure Legend Snippet: Characterization of the interaction between polyphosphate and TFPIα. ( A ) 96-well plates were coated with 5 μg/ml TFPIα (○,●) or BSA (◻) and increasing concentrations of biotinylated-polyphosphate (bio-polyP) (◻, ○,●) was added to selected wells. Selected experiments were done in the presence of 100 μM SCP (●). Binding was detected with streptavidin-HRP. Data are mean ± SE (n = 3).

    Techniques Used: Binding Assay

    Related Articles

    Centrifugation:

    Article Title: Inflammatory stimuli induce inhibitory S-nitrosylation of the deacetylase SIRT1 to increase acetylation and activation of p53 and p65
    Article Snippet: The cells were lysed in lysis buffer A, clarified by centrifugation, mixed with 1 volume of blocking buffer [PBS-HCl (pH 3.5), 150 mM NaCl, 1 mM EDTA, 1 mM DTPA, 2.5% SDS, 0.1 mM neocuproine, and 20 mM NEM], and incubated at 50°C for 20 min with vortex every 2 min. Lysates were biotinylated with NEM-PEG2 -biotin (0.2 mM) (Pierce) in the presence of ascorbate sodium (4 mM) for the detection of SNO or with DTT (4 mM) for the detection of both SNO and SOX ( ). .. Biotinylated SIRT1 was detected with horseradish peroxidase (HRP)–conjugated streptavidin (Pierce).

    Article Title: Heme and non-heme iron transporters in non-polarized and polarized cells
    Article Snippet: Then, cells were extracted in RIPA buffer [150 mM NaCl, 50 mM Tris (pH 8.0), 5 mM EDTA, 1% Nonidet P-40, 0.5% deoxycholate, and 0.1% SDS] and the extracts were clarified by centrifugation at 14,000 × g for 15 min. .. Biotinylated proteins were detected by ImmunoPure Avidin, Horseradish Peroxidase, Conjugated (Pierce Biotechnology, Rockford, IL).

    Blocking Assay:

    Article Title: Novel inter-hemispheric white matter connectivity in the BTBR mouse model of autism
    Article Snippet: .. The membranes were then washed with TBS-T, incubated with biotinylated goat-anti-mouse Ig G (1:10,000: Pierce Biotechnology Inc., Rockford, IL) for 2 hrs at RT, washed with TBS-T and incubated with streptavidin conjugated with HRP (1:5,000 in blocking buffer; Pierce Biotechonology) for 1 hr. .. The blot was developed with Super Signal/ECL chemiluminescent substrate (Pierce Biotechnology) and photographed with a LAS-1000plus (Fuji).

    Article Title: Topology and Accessibility of Germination Proteins in the Bacillus subtilis Spore Inner Membrane
    Article Snippet: Incubations with antisera were all carried out in 5% nonfat dry milk (Santa Cruz Biotech, Santa Cruz, CA) as the blocking agent. .. Biotinylated proteins were detected on PVDF membranes by incubation with streptavidin coupled to horseradish peroxidase (HRP) (Pierce) in 2.5% bovine serum albumin, and HRP on membranes was detected by chemiluminescence as described by the supplier.

    Article Title: Inflammatory stimuli induce inhibitory S-nitrosylation of the deacetylase SIRT1 to increase acetylation and activation of p53 and p65
    Article Snippet: The cells were lysed in lysis buffer A, clarified by centrifugation, mixed with 1 volume of blocking buffer [PBS-HCl (pH 3.5), 150 mM NaCl, 1 mM EDTA, 1 mM DTPA, 2.5% SDS, 0.1 mM neocuproine, and 20 mM NEM], and incubated at 50°C for 20 min with vortex every 2 min. Lysates were biotinylated with NEM-PEG2 -biotin (0.2 mM) (Pierce) in the presence of ascorbate sodium (4 mM) for the detection of SNO or with DTT (4 mM) for the detection of both SNO and SOX ( ). .. Biotinylated SIRT1 was detected with horseradish peroxidase (HRP)–conjugated streptavidin (Pierce).

    Article Title: Phosphorylation and regulation of a G protein-coupled receptor by protein kinase CK2
    Article Snippet: .. Samples were then separated by SDS-PAGE, electroblotted to nitrocellulose membranes, and, after blocking with TBST plus 5% milk powder for 1 h, incubated with 50 ng/ml streptavidin conjugated to horseradish peroxidase (Pierce Chemical Co.) for 30 min. Biotinylated proteins were then detected with chemiluminescence reagent (ECL plus; GE Healthcare). .. Generation of mouse M3 -muscarinic receptor antibody The region S344 -L462 of the mouse M3 -muscarinic receptor third intracellular loop was produced as an N-terminal tagged GST/receptor fusion protein, which was used to inoculate New Zealand white rabbits (Harlan Sera-Labs).

    Article Title: Engineering of a Bacillus subtilis Strain with Adjustable Levels of Intracellular Biotin for Secretory Production of Functional Streptavidin
    Article Snippet: .. After blocking with bovine serum albumin, wells were incubated with a mixture containing 25 ng of biotinylated HRP (Pierce) and different amounts of the purified SAV or commercial SAV from Roche Molecular Biochemicals. .. All PCRs (except colony PCR) were carried out using Vent DNA polymerase (New England BioLabs Canada).

    Adsorption:

    Article Title: Role of Nucleolin in Human Parainfluenza Virus Type 3 Infection of Human Lung Epithelial Cells
    Article Snippet: Biotinylated HPIV-3 (2 multiplicities of infection [MOI]) was added to A549 cells, and following 2 h of adsorption at 37°C the cells were washed extensively with 1× PBS and the cell lysate was subjected to immunoprecipitation with antinucleolin or anti-β-catenin antibody in the presence of protein A-Sepharose beads (Amersham-Pharmacia) as described previously ( ). .. The membrane was blotted with avidin conjugated to horseradish peroxidase (avidin-HRP) (Pierce) to visualize the biotinylated envelope proteins of HPIV-3.

    Electrophoresis:

    Article Title: Topology and Accessibility of Germination Proteins in the Bacillus subtilis Spore Inner Membrane
    Article Snippet: Western blot analyses were carried out following SDS-polyacrylamide gel (12%) electrophoresis (SDS-PAGE) and transfer of proteins to polyvinylidene difluoride (PVDF) membranes, and various germination proteins were detected as described previously ( , , , ). .. Biotinylated proteins were detected on PVDF membranes by incubation with streptavidin coupled to horseradish peroxidase (HRP) (Pierce) in 2.5% bovine serum albumin, and HRP on membranes was detected by chemiluminescence as described by the supplier.

    Incubation:

    Article Title: Novel inter-hemispheric white matter connectivity in the BTBR mouse model of autism
    Article Snippet: .. The membranes were then washed with TBS-T, incubated with biotinylated goat-anti-mouse Ig G (1:10,000: Pierce Biotechnology Inc., Rockford, IL) for 2 hrs at RT, washed with TBS-T and incubated with streptavidin conjugated with HRP (1:5,000 in blocking buffer; Pierce Biotechonology) for 1 hr. .. The blot was developed with Super Signal/ECL chemiluminescent substrate (Pierce Biotechnology) and photographed with a LAS-1000plus (Fuji).

    Article Title: Topology and Accessibility of Germination Proteins in the Bacillus subtilis Spore Inner Membrane
    Article Snippet: .. Biotinylated proteins were detected on PVDF membranes by incubation with streptavidin coupled to horseradish peroxidase (HRP) (Pierce) in 2.5% bovine serum albumin, and HRP on membranes was detected by chemiluminescence as described by the supplier. .. Dormant spores (10 ml) of various strains at an optical density at 600 nm (OD600 ) of ∼25 were decoated by incubation at 70°C for 2 h in 0.1 M NaOH, 0.1 M NaCl, 0.5% SDS, and 0.1 M DTT and washed 10 times with water as described previously ( ).

    Article Title: Inflammatory stimuli induce inhibitory S-nitrosylation of the deacetylase SIRT1 to increase acetylation and activation of p53 and p65
    Article Snippet: After precipitation with −20°C acetone, antibody recognizing FLAG M2 (Sigma) and protein G and protein A agarose suspension (#IP05, Calbiochem) were added, and the samples were incubated at room temperature for 1 hour or at 4°C overnight. .. Biotinylated SIRT1 was detected with horseradish peroxidase (HRP)–conjugated streptavidin (Pierce).

    Article Title: Activation of a Retroviral Membrane Fusion Protein: Soluble Receptor-induced Liposome Binding of the ALSV Envelope Glycoprotein
    Article Snippet: Anti-DAF antibody was added and incubated for 1 h at 4°C followed by another 1-h incubation after the addition of protein A–agarose ( Boehringer Mannheim Biochemicals , Indianapolis, IN). .. Proteins were transferred to nitrocellulose and probed with streptavidin coupled to HRP (strep-HRP; Pierce) to detect biotinylated proteins.

    Article Title: Role of Capsid Anchor in the Morphogenesis of Zika Virus
    Article Snippet: .. For SV5-tagged proteins, membranes were incubated for 1 h with anti-SV5 mAb (1 μg/ml), washed, and further probed with HRP-linked anti-mouse IgG goat antibodies (catalog no. 074-1809, 1:10,000; KPL, Gaithersburg, MA) for 1 h. 4G2 avibody was previously treated with biotinylated-HRP (ThermoFisher-Pierce, Rockford, IL, USA), incubated for 1 h, and washed. .. Mouse HRP-conjugated anti-actin (clone AC-15, 1:30,000; Sigma-Aldrich) was used as loading control.

    Article Title: Phosphorylation and regulation of a G protein-coupled receptor by protein kinase CK2
    Article Snippet: .. Samples were then separated by SDS-PAGE, electroblotted to nitrocellulose membranes, and, after blocking with TBST plus 5% milk powder for 1 h, incubated with 50 ng/ml streptavidin conjugated to horseradish peroxidase (Pierce Chemical Co.) for 30 min. Biotinylated proteins were then detected with chemiluminescence reagent (ECL plus; GE Healthcare). .. Generation of mouse M3 -muscarinic receptor antibody The region S344 -L462 of the mouse M3 -muscarinic receptor third intracellular loop was produced as an N-terminal tagged GST/receptor fusion protein, which was used to inoculate New Zealand white rabbits (Harlan Sera-Labs).

    Article Title: Synthesis and Characterization of PAMAM Dendrimer Based Multifunctional Nanodevices for targeting αvβ3 Integrins
    Article Snippet: At the end of this incubation period, plate was washed five times with binding buffer. .. The bound biotinylated NDs were detected by Immuno-Pure Avidin conjugated with horseradish peroxidase (product # 21123; Pierce, Rockford, IL), which was subsequently developed using o-Phenylenediamine and the absorbance at 450 nm recorded according to the manufacturer’s directions ( ).

    Article Title: Heme and non-heme iron transporters in non-polarized and polarized cells
    Article Snippet: After 2-h incubation, the beads were washed and the proteins were eluted with Laemmli buffer. .. Biotinylated proteins were detected by ImmunoPure Avidin, Horseradish Peroxidase, Conjugated (Pierce Biotechnology, Rockford, IL).

    Article Title: Engineering of a Bacillus subtilis Strain with Adjustable Levels of Intracellular Biotin for Secretory Production of Functional Streptavidin
    Article Snippet: .. After blocking with bovine serum albumin, wells were incubated with a mixture containing 25 ng of biotinylated HRP (Pierce) and different amounts of the purified SAV or commercial SAV from Roche Molecular Biochemicals. .. All PCRs (except colony PCR) were carried out using Vent DNA polymerase (New England BioLabs Canada).

    Article Title: Role of Nucleolin in Human Parainfluenza Virus Type 3 Infection of Human Lung Epithelial Cells
    Article Snippet: Following 12 h of incubation at 4°C, the beads were washed, the bound proteins were subjected to SDS-7.5% PAGE, and the separated proteins were transferred to the nitrocellulose membrane. .. The membrane was blotted with avidin conjugated to horseradish peroxidase (avidin-HRP) (Pierce) to visualize the biotinylated envelope proteins of HPIV-3.

    Cell Culture:

    Article Title: Phosphorylation and regulation of a G protein-coupled receptor by protein kinase CK2
    Article Snippet: Biotinylation Cells cultured on 6-well plates were incubated in buffer containing 1 mM biotin (Pierce Chemical Co.) for 30 min at 37°C. .. Samples were then separated by SDS-PAGE, electroblotted to nitrocellulose membranes, and, after blocking with TBST plus 5% milk powder for 1 h, incubated with 50 ng/ml streptavidin conjugated to horseradish peroxidase (Pierce Chemical Co.) for 30 min. Biotinylated proteins were then detected with chemiluminescence reagent (ECL plus; GE Healthcare).

    Expressing:

    Article Title: Novel inter-hemispheric white matter connectivity in the BTBR mouse model of autism
    Article Snippet: Paragraph title: 2.5. Total MBP and MAP expression ... The membranes were then washed with TBS-T, incubated with biotinylated goat-anti-mouse Ig G (1:10,000: Pierce Biotechnology Inc., Rockford, IL) for 2 hrs at RT, washed with TBS-T and incubated with streptavidin conjugated with HRP (1:5,000 in blocking buffer; Pierce Biotechonology) for 1 hr.

    Article Title: Activation of a Retroviral Membrane Fusion Protein: Soluble Receptor-induced Liposome Binding of the ALSV Envelope Glycoprotein
    Article Snippet: Biotinylation and Coimmunoprecipitation Binding Assay After induction with sodium butyrate (5 mM for API and 10 mM for Glu2-PI) for 16–18 h, cells expressing GPI-anchored proteins were labeled with the membrane-impermeant reagent NHS-LC-biotin (Pierce) and treated with PI-PLC as described previously ( ). sTva, s47, and s47WA proteins were biotinylated on ice for 45 min in PBS (pH 7.8) containing 0.5 mM MgCl2 and 1 mg/ml NHS-LC-biotin. .. Proteins were transferred to nitrocellulose and probed with streptavidin coupled to HRP (strep-HRP; Pierce) to detect biotinylated proteins.

    Western Blot:

    Article Title: Topology and Accessibility of Germination Proteins in the Bacillus subtilis Spore Inner Membrane
    Article Snippet: Paragraph title: Western blot analysis of germination proteins. ... Biotinylated proteins were detected on PVDF membranes by incubation with streptavidin coupled to horseradish peroxidase (HRP) (Pierce) in 2.5% bovine serum albumin, and HRP on membranes was detected by chemiluminescence as described by the supplier.

    Article Title: Role of Capsid Anchor in the Morphogenesis of Zika Virus
    Article Snippet: Paragraph title: Western blotting. ... For SV5-tagged proteins, membranes were incubated for 1 h with anti-SV5 mAb (1 μg/ml), washed, and further probed with HRP-linked anti-mouse IgG goat antibodies (catalog no. 074-1809, 1:10,000; KPL, Gaithersburg, MA) for 1 h. 4G2 avibody was previously treated with biotinylated-HRP (ThermoFisher-Pierce, Rockford, IL, USA), incubated for 1 h, and washed.

    Article Title: Role of Nucleolin in Human Parainfluenza Virus Type 3 Infection of Human Lung Epithelial Cells
    Article Snippet: The membrane was blotted with avidin conjugated to horseradish peroxidase (avidin-HRP) (Pierce) to visualize the biotinylated envelope proteins of HPIV-3. .. Similarly, the cell lysates obtained from the biotinylated HPIV-3-treated (incubated at 37 or 4°C) or VSV-treated (incubated at 37°C) cells were precipitated with avidin-agarose as described above and the bound proteins were subjected to Western blot analysis with antinucleolin antibody.

    Competitive Binding Assay:

    Article Title: Synthesis and Characterization of PAMAM Dendrimer Based Multifunctional Nanodevices for targeting αvβ3 Integrins
    Article Snippet: Competition assay was conducted adding 4, 40 and 400 fold excess of cRGD-peptide (Chemicon Int., Temecula, CA) simultaneously with the nanodevice. .. The bound biotinylated NDs were detected by Immuno-Pure Avidin conjugated with horseradish peroxidase (product # 21123; Pierce, Rockford, IL), which was subsequently developed using o-Phenylenediamine and the absorbance at 450 nm recorded according to the manufacturer’s directions ( ).

    Transfection:

    Article Title: Inflammatory stimuli induce inhibitory S-nitrosylation of the deacetylase SIRT1 to increase acetylation and activation of p53 and p65
    Article Snippet: To evaluate the role of CXXC motifs in S-nitrosylation of SIRT1 and assess S-nitrosylation and oxidative thiol modifications of SIRT1, COS-7 cells were transfected with FLAG-tagged wild-type SIRT1 or SIRT1M3 . .. Biotinylated SIRT1 was detected with horseradish peroxidase (HRP)–conjugated streptavidin (Pierce).

    Infection:

    Article Title: Role of Nucleolin in Human Parainfluenza Virus Type 3 Infection of Human Lung Epithelial Cells
    Article Snippet: Biotinylated HPIV-3 (2 multiplicities of infection [MOI]) was added to A549 cells, and following 2 h of adsorption at 37°C the cells were washed extensively with 1× PBS and the cell lysate was subjected to immunoprecipitation with antinucleolin or anti-β-catenin antibody in the presence of protein A-Sepharose beads (Amersham-Pharmacia) as described previously ( ). .. The membrane was blotted with avidin conjugated to horseradish peroxidase (avidin-HRP) (Pierce) to visualize the biotinylated envelope proteins of HPIV-3.

    Binding Assay:

    Article Title: Activation of a Retroviral Membrane Fusion Protein: Soluble Receptor-induced Liposome Binding of the ALSV Envelope Glycoprotein
    Article Snippet: Paragraph title: Biotinylation and Coimmunoprecipitation Binding Assay ... Proteins were transferred to nitrocellulose and probed with streptavidin coupled to HRP (strep-HRP; Pierce) to detect biotinylated proteins.

    Article Title: Synthesis and Characterization of PAMAM Dendrimer Based Multifunctional Nanodevices for targeting αvβ3 Integrins
    Article Snippet: Paragraph title: Nanodevice binding to αv β3 integrin in vitro ... The bound biotinylated NDs were detected by Immuno-Pure Avidin conjugated with horseradish peroxidase (product # 21123; Pierce, Rockford, IL), which was subsequently developed using o-Phenylenediamine and the absorbance at 450 nm recorded according to the manufacturer’s directions ( ).

    Article Title: Engineering of a Bacillus subtilis Strain with Adjustable Levels of Intracellular Biotin for Secretory Production of Functional Streptavidin
    Article Snippet: Paragraph title: Determination of number of biotin binding sites on SAV produced by WB800BIO(pSSAV-T cry ). ... After blocking with bovine serum albumin, wells were incubated with a mixture containing 25 ng of biotinylated HRP (Pierce) and different amounts of the purified SAV or commercial SAV from Roche Molecular Biochemicals.

    Radioactivity:

    Article Title: Phosphorylation and regulation of a G protein-coupled receptor by protein kinase CK2
    Article Snippet: Equivalent amounts of receptor were ensured by analysis of receptor levels using [3 H]-NMS radiolabeling. .. Samples were then separated by SDS-PAGE, electroblotted to nitrocellulose membranes, and, after blocking with TBST plus 5% milk powder for 1 h, incubated with 50 ng/ml streptavidin conjugated to horseradish peroxidase (Pierce Chemical Co.) for 30 min. Biotinylated proteins were then detected with chemiluminescence reagent (ECL plus; GE Healthcare).

    Avidin-Biotin Assay:

    Article Title: Synthesis and Characterization of PAMAM Dendrimer Based Multifunctional Nanodevices for targeting αvβ3 Integrins
    Article Snippet: .. The bound biotinylated NDs were detected by Immuno-Pure Avidin conjugated with horseradish peroxidase (product # 21123; Pierce, Rockford, IL), which was subsequently developed using o-Phenylenediamine and the absorbance at 450 nm recorded according to the manufacturer’s directions ( ). .. HDMECs were seeded (50,000 cells per well) and grown on chamber slides (Lab Tek) in a conditioned endothelial culture medium EBM Bullet kit 2 (Cambrex, Walkersville, MD).

    Article Title: Heme and non-heme iron transporters in non-polarized and polarized cells
    Article Snippet: .. Biotinylated proteins were detected by ImmunoPure Avidin, Horseradish Peroxidase, Conjugated (Pierce Biotechnology, Rockford, IL). .. Subcellular fractionation HEp-2 cells stably expressing HA-tagged HO-2 were grown to confluency in a 100-cm2 dish.

    Article Title: Role of Nucleolin in Human Parainfluenza Virus Type 3 Infection of Human Lung Epithelial Cells
    Article Snippet: .. The membrane was blotted with avidin conjugated to horseradish peroxidase (avidin-HRP) (Pierce) to visualize the biotinylated envelope proteins of HPIV-3. .. Similarly, the cell lysates obtained from the biotinylated HPIV-3-treated (incubated at 37 or 4°C) or VSV-treated (incubated at 37°C) cells were precipitated with avidin-agarose as described above and the bound proteins were subjected to Western blot analysis with antinucleolin antibody.

    Labeling:

    Article Title: Activation of a Retroviral Membrane Fusion Protein: Soluble Receptor-induced Liposome Binding of the ALSV Envelope Glycoprotein
    Article Snippet: Biotinylation and Coimmunoprecipitation Binding Assay After induction with sodium butyrate (5 mM for API and 10 mM for Glu2-PI) for 16–18 h, cells expressing GPI-anchored proteins were labeled with the membrane-impermeant reagent NHS-LC-biotin (Pierce) and treated with PI-PLC as described previously ( ). sTva, s47, and s47WA proteins were biotinylated on ice for 45 min in PBS (pH 7.8) containing 0.5 mM MgCl2 and 1 mg/ml NHS-LC-biotin. .. Proteins were transferred to nitrocellulose and probed with streptavidin coupled to HRP (strep-HRP; Pierce) to detect biotinylated proteins.

    Article Title: Heme and non-heme iron transporters in non-polarized and polarized cells
    Article Snippet: Paragraph title: Cell surface labeling ... Biotinylated proteins were detected by ImmunoPure Avidin, Horseradish Peroxidase, Conjugated (Pierce Biotechnology, Rockford, IL).

    Purification:

    Article Title: Synthesis and Characterization of PAMAM Dendrimer Based Multifunctional Nanodevices for targeting αvβ3 Integrins
    Article Snippet: An aliquot of 100 µL of purified αv β3 integrin receptor (Chemicon Int., Temecula, CA) 0.5 µg/mL in coating buffer (50 mM Tris, pH 7.4; 100 mM NaCl and 1 mM each of CaCl2 ,, MnCl2 , and MgCl2 ,) was immobilized in 96-well microtiter plate overnight at 4 °C. .. The bound biotinylated NDs were detected by Immuno-Pure Avidin conjugated with horseradish peroxidase (product # 21123; Pierce, Rockford, IL), which was subsequently developed using o-Phenylenediamine and the absorbance at 450 nm recorded according to the manufacturer’s directions ( ).

    Article Title: Engineering of a Bacillus subtilis Strain with Adjustable Levels of Intracellular Biotin for Secretory Production of Functional Streptavidin
    Article Snippet: .. After blocking with bovine serum albumin, wells were incubated with a mixture containing 25 ng of biotinylated HRP (Pierce) and different amounts of the purified SAV or commercial SAV from Roche Molecular Biochemicals. .. All PCRs (except colony PCR) were carried out using Vent DNA polymerase (New England BioLabs Canada).

    Polyacrylamide Gel Electrophoresis:

    Article Title: Role of Nucleolin in Human Parainfluenza Virus Type 3 Infection of Human Lung Epithelial Cells
    Article Snippet: Following 12 h of incubation at 4°C, the beads were washed, the bound proteins were subjected to SDS-7.5% PAGE, and the separated proteins were transferred to the nitrocellulose membrane. .. The membrane was blotted with avidin conjugated to horseradish peroxidase (avidin-HRP) (Pierce) to visualize the biotinylated envelope proteins of HPIV-3.

    Mouse Assay:

    Article Title: Novel inter-hemispheric white matter connectivity in the BTBR mouse model of autism
    Article Snippet: The membranes were then washed with TBS-T, incubated with biotinylated goat-anti-mouse Ig G (1:10,000: Pierce Biotechnology Inc., Rockford, IL) for 2 hrs at RT, washed with TBS-T and incubated with streptavidin conjugated with HRP (1:5,000 in blocking buffer; Pierce Biotechonology) for 1 hr. .. For gels n=4 BTBR and n = 4 C57Bl/6 mice were used per gel and gels were repeated in duplicate.

    SDS Page:

    Article Title: Novel inter-hemispheric white matter connectivity in the BTBR mouse model of autism
    Article Snippet: SDS-PAGE was performed with 4-20% agarose gels for 65 mins at 120 mV after which the gels were washed in transfer buffer and electroblotted using a semi-dry transfer apparatus onto Immobilon-P transfer membranes (Millipore Corporation, MA). .. The membranes were then washed with TBS-T, incubated with biotinylated goat-anti-mouse Ig G (1:10,000: Pierce Biotechnology Inc., Rockford, IL) for 2 hrs at RT, washed with TBS-T and incubated with streptavidin conjugated with HRP (1:5,000 in blocking buffer; Pierce Biotechonology) for 1 hr.

    Article Title: Topology and Accessibility of Germination Proteins in the Bacillus subtilis Spore Inner Membrane
    Article Snippet: Unless noted otherwise, protein samples were incubated for 1 h at 23°C in Laemmli's loading buffer (Bio-Rad, Hercules, CA) with 55 mM dithiothreitol (DTT) prior to SDS-PAGE. .. Biotinylated proteins were detected on PVDF membranes by incubation with streptavidin coupled to horseradish peroxidase (HRP) (Pierce) in 2.5% bovine serum albumin, and HRP on membranes was detected by chemiluminescence as described by the supplier.

    Article Title: Inflammatory stimuli induce inhibitory S-nitrosylation of the deacetylase SIRT1 to increase acetylation and activation of p53 and p65
    Article Snippet: The immunopurified complexes were washed five times with wash buffer [50 mM Hepes (pH 7.7), 150 mM NaCl, 0.1% NP-40, 1 mM EDTA] and separated by SDS-PAGE. .. Biotinylated SIRT1 was detected with horseradish peroxidase (HRP)–conjugated streptavidin (Pierce).

    Article Title: Activation of a Retroviral Membrane Fusion Protein: Soluble Receptor-induced Liposome Binding of the ALSV Envelope Glycoprotein
    Article Snippet: Samples were washed four times in lysis buffer (Hepes-buffered saline with 1% Nonidet P-40), boiled, and reduced in Laemmli sample buffer, and separated by SDS-PAGE. .. Proteins were transferred to nitrocellulose and probed with streptavidin coupled to HRP (strep-HRP; Pierce) to detect biotinylated proteins.

    Article Title: Role of Capsid Anchor in the Morphogenesis of Zika Virus
    Article Snippet: Briefly, samples were separated by nonreducing SDS-PAGE, transferred to polyvinylidene difluoride membranes (Millipore, Temecula, CA) and blocked with 5% milk solution in phosphate-buffered saline (PBS-milk). .. For SV5-tagged proteins, membranes were incubated for 1 h with anti-SV5 mAb (1 μg/ml), washed, and further probed with HRP-linked anti-mouse IgG goat antibodies (catalog no. 074-1809, 1:10,000; KPL, Gaithersburg, MA) for 1 h. 4G2 avibody was previously treated with biotinylated-HRP (ThermoFisher-Pierce, Rockford, IL, USA), incubated for 1 h, and washed.

    Article Title: Phosphorylation and regulation of a G protein-coupled receptor by protein kinase CK2
    Article Snippet: .. Samples were then separated by SDS-PAGE, electroblotted to nitrocellulose membranes, and, after blocking with TBST plus 5% milk powder for 1 h, incubated with 50 ng/ml streptavidin conjugated to horseradish peroxidase (Pierce Chemical Co.) for 30 min. Biotinylated proteins were then detected with chemiluminescence reagent (ECL plus; GE Healthcare). .. Generation of mouse M3 -muscarinic receptor antibody The region S344 -L462 of the mouse M3 -muscarinic receptor third intracellular loop was produced as an N-terminal tagged GST/receptor fusion protein, which was used to inoculate New Zealand white rabbits (Harlan Sera-Labs).

    Enzyme-linked Immunosorbent Assay:

    Article Title: Engineering of a Bacillus subtilis Strain with Adjustable Levels of Intracellular Biotin for Secretory Production of Functional Streptavidin
    Article Snippet: The number of biotin binding sites on the purified tetrameric SAV was determined using a competition ELISA method similar to the one established for the biotin assay but with modifications. .. After blocking with bovine serum albumin, wells were incubated with a mixture containing 25 ng of biotinylated HRP (Pierce) and different amounts of the purified SAV or commercial SAV from Roche Molecular Biochemicals.

    In Vitro:

    Article Title: Synthesis and Characterization of PAMAM Dendrimer Based Multifunctional Nanodevices for targeting αvβ3 Integrins
    Article Snippet: Paragraph title: Nanodevice binding to αv β3 integrin in vitro ... The bound biotinylated NDs were detected by Immuno-Pure Avidin conjugated with horseradish peroxidase (product # 21123; Pierce, Rockford, IL), which was subsequently developed using o-Phenylenediamine and the absorbance at 450 nm recorded according to the manufacturer’s directions ( ).

    Produced:

    Article Title: Engineering of a Bacillus subtilis Strain with Adjustable Levels of Intracellular Biotin for Secretory Production of Functional Streptavidin
    Article Snippet: Paragraph title: Determination of number of biotin binding sites on SAV produced by WB800BIO(pSSAV-T cry ). ... After blocking with bovine serum albumin, wells were incubated with a mixture containing 25 ng of biotinylated HRP (Pierce) and different amounts of the purified SAV or commercial SAV from Roche Molecular Biochemicals.

    Immunoprecipitation:

    Article Title: Activation of a Retroviral Membrane Fusion Protein: Soluble Receptor-induced Liposome Binding of the ALSV Envelope Glycoprotein
    Article Snippet: Proteins were transferred to nitrocellulose and probed with streptavidin coupled to HRP (strep-HRP; Pierce) to detect biotinylated proteins. .. For measuring sTva dissociation from API, unlabeled PI-PLC–released API was immunoprecipitated with anti-DAF/protein A–agarose as described above.

    Article Title: Phosphorylation and regulation of a G protein-coupled receptor by protein kinase CK2
    Article Snippet: The cells were then lysed, and the M3 -muscarinic receptor was immunoprecipitated as described for phosphorylation experiments. .. Samples were then separated by SDS-PAGE, electroblotted to nitrocellulose membranes, and, after blocking with TBST plus 5% milk powder for 1 h, incubated with 50 ng/ml streptavidin conjugated to horseradish peroxidase (Pierce Chemical Co.) for 30 min. Biotinylated proteins were then detected with chemiluminescence reagent (ECL plus; GE Healthcare).

    Article Title: Role of Nucleolin in Human Parainfluenza Virus Type 3 Infection of Human Lung Epithelial Cells
    Article Snippet: Biotinylated HPIV-3 (2 multiplicities of infection [MOI]) was added to A549 cells, and following 2 h of adsorption at 37°C the cells were washed extensively with 1× PBS and the cell lysate was subjected to immunoprecipitation with antinucleolin or anti-β-catenin antibody in the presence of protein A-Sepharose beads (Amersham-Pharmacia) as described previously ( ). .. The membrane was blotted with avidin conjugated to horseradish peroxidase (avidin-HRP) (Pierce) to visualize the biotinylated envelope proteins of HPIV-3.

    Lysis:

    Article Title: Inflammatory stimuli induce inhibitory S-nitrosylation of the deacetylase SIRT1 to increase acetylation and activation of p53 and p65
    Article Snippet: The cells were lysed in lysis buffer A, clarified by centrifugation, mixed with 1 volume of blocking buffer [PBS-HCl (pH 3.5), 150 mM NaCl, 1 mM EDTA, 1 mM DTPA, 2.5% SDS, 0.1 mM neocuproine, and 20 mM NEM], and incubated at 50°C for 20 min with vortex every 2 min. Lysates were biotinylated with NEM-PEG2 -biotin (0.2 mM) (Pierce) in the presence of ascorbate sodium (4 mM) for the detection of SNO or with DTT (4 mM) for the detection of both SNO and SOX ( ). .. Biotinylated SIRT1 was detected with horseradish peroxidase (HRP)–conjugated streptavidin (Pierce).

    Article Title: Activation of a Retroviral Membrane Fusion Protein: Soluble Receptor-induced Liposome Binding of the ALSV Envelope Glycoprotein
    Article Snippet: Samples were washed four times in lysis buffer (Hepes-buffered saline with 1% Nonidet P-40), boiled, and reduced in Laemmli sample buffer, and separated by SDS-PAGE. .. Proteins were transferred to nitrocellulose and probed with streptavidin coupled to HRP (strep-HRP; Pierce) to detect biotinylated proteins.

    Fluorescence In Situ Hybridization:

    Article Title: Novel inter-hemispheric white matter connectivity in the BTBR mouse model of autism
    Article Snippet: Nonspecific sites on the membrane were blocked by incubating with 5% fish gelatin in TBS-T overnight at 4°C. .. The membranes were then washed with TBS-T, incubated with biotinylated goat-anti-mouse Ig G (1:10,000: Pierce Biotechnology Inc., Rockford, IL) for 2 hrs at RT, washed with TBS-T and incubated with streptavidin conjugated with HRP (1:5,000 in blocking buffer; Pierce Biotechonology) for 1 hr.

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    Thermo Fisher horseradish peroxidase hrp conjugated streptavidin
    Comparison of receptor binding affinities to human- and avian-like receptors. Receptor binding avidities of recombinant viruses were measured using the solid-phase direct binding assay. The recombinant viruses were bound to fetuin-coated plates. Subsequently, serially diluted biotinylated avian-like receptor sialylglycopolymers (Neu5Acα2-3Galb1-4GlcNAcb-PAA-biotin [3′SLN-PAA]) and human-like receptor sialylglycopolymers (Neu5Acα2-6GalNAca-PAA-biotin [6′SLN-PAA]) were added. The plates were developed using an <t>HRP-conjugated</t> <t>streptavidin,</t> TMB substrate solution, and stop solution; the absorbance at 450 nm was measured by a microplate reader. (A) Receptor binding avidities of recombinant viruses to avian-like receptor. (B) Receptor binding avidities of recombinant viruses to human-like receptor. The data represents the average of three independent experiments ± SD . Statistical significance was analyzed by one-way ANOVA (compared to rH 2 N 2 , ∗ P
    Horseradish Peroxidase Hrp Conjugated Streptavidin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Comparison of receptor binding affinities to human- and avian-like receptors. Receptor binding avidities of recombinant viruses were measured using the solid-phase direct binding assay. The recombinant viruses were bound to fetuin-coated plates. Subsequently, serially diluted biotinylated avian-like receptor sialylglycopolymers (Neu5Acα2-3Galb1-4GlcNAcb-PAA-biotin [3′SLN-PAA]) and human-like receptor sialylglycopolymers (Neu5Acα2-6GalNAca-PAA-biotin [6′SLN-PAA]) were added. The plates were developed using an HRP-conjugated streptavidin, TMB substrate solution, and stop solution; the absorbance at 450 nm was measured by a microplate reader. (A) Receptor binding avidities of recombinant viruses to avian-like receptor. (B) Receptor binding avidities of recombinant viruses to human-like receptor. The data represents the average of three independent experiments ± SD . Statistical significance was analyzed by one-way ANOVA (compared to rH 2 N 2 , ∗ P

    Journal: Frontiers in Microbiology

    Article Title: Acquisition of Innate Inhibitor Resistance and Mammalian Pathogenicity During Egg Adaptation by the H9N2 Avian Influenza Virus

    doi: 10.3389/fmicb.2018.01939

    Figure Lengend Snippet: Comparison of receptor binding affinities to human- and avian-like receptors. Receptor binding avidities of recombinant viruses were measured using the solid-phase direct binding assay. The recombinant viruses were bound to fetuin-coated plates. Subsequently, serially diluted biotinylated avian-like receptor sialylglycopolymers (Neu5Acα2-3Galb1-4GlcNAcb-PAA-biotin [3′SLN-PAA]) and human-like receptor sialylglycopolymers (Neu5Acα2-6GalNAca-PAA-biotin [6′SLN-PAA]) were added. The plates were developed using an HRP-conjugated streptavidin, TMB substrate solution, and stop solution; the absorbance at 450 nm was measured by a microplate reader. (A) Receptor binding avidities of recombinant viruses to avian-like receptor. (B) Receptor binding avidities of recombinant viruses to human-like receptor. The data represents the average of three independent experiments ± SD . Statistical significance was analyzed by one-way ANOVA (compared to rH 2 N 2 , ∗ P

    Article Snippet: Finally, the plates were washed three times followed by incubation in horseradish peroxidase (HRP)-conjugated streptavidin (Thermo Fisher SCIENTIFIC, MA, United States) for 1 h at 4°C.

    Techniques: Binding Assay, Recombinant

    Comparison of receptor binding affinities to human- and avian-like receptors. Receptor binding avidities of recombinant viruses were measured using the solid-phase direct binding assay. The recombinant viruses were bound to fetuin-coated plates. Subsequently, serially diluted biotinylated avian-like receptor sialylglycopolymers (Neu5Acα2-3Galb1-4GlcNAcb-PAA-biotin [3′SLN-PAA]) and human-like receptor sialylglycopolymers (Neu5Acα2-6GalNAca-PAA-biotin [6′SLN-PAA]) were added. The plates were developed using an HRP-conjugated streptavidin, TMB substrate solution, and stop solution; the absorbance at 450 nm was measured by a microplate reader. (A) Receptor binding avidities of recombinant viruses to avian-like receptor. (B) Receptor binding avidities of recombinant viruses to human-like receptor. The data represents the average of three independent experiments ± SD . Statistical significance was analyzed by one-way ANOVA (compared to rH 2 N 2 , ∗ P

    Journal: Frontiers in Microbiology

    Article Title: Acquisition of Innate Inhibitor Resistance and Mammalian Pathogenicity During Egg Adaptation by the H9N2 Avian Influenza Virus

    doi: 10.3389/fmicb.2018.01939

    Figure Lengend Snippet: Comparison of receptor binding affinities to human- and avian-like receptors. Receptor binding avidities of recombinant viruses were measured using the solid-phase direct binding assay. The recombinant viruses were bound to fetuin-coated plates. Subsequently, serially diluted biotinylated avian-like receptor sialylglycopolymers (Neu5Acα2-3Galb1-4GlcNAcb-PAA-biotin [3′SLN-PAA]) and human-like receptor sialylglycopolymers (Neu5Acα2-6GalNAca-PAA-biotin [6′SLN-PAA]) were added. The plates were developed using an HRP-conjugated streptavidin, TMB substrate solution, and stop solution; the absorbance at 450 nm was measured by a microplate reader. (A) Receptor binding avidities of recombinant viruses to avian-like receptor. (B) Receptor binding avidities of recombinant viruses to human-like receptor. The data represents the average of three independent experiments ± SD . Statistical significance was analyzed by one-way ANOVA (compared to rH 2 N 2 , ∗ P

    Article Snippet: Finally, the plates were washed three times followed by incubation in horseradish peroxidase (HRP)-conjugated streptavidin (Thermo Fisher SCIENTIFIC, MA, United States) for 1 h at 4°C.

    Techniques: Binding Assay, Recombinant

    HIV-1-specific antibody responses following panobinostat treatment. (A) Concentrations of gp120-specific IgG antibodies pre- and post-panobinostat treatment. (B) ELISA to measure dimeric FcγRIIIa binding. Ninety-six-well ELISA plates were first coated with 50 ng/well of HIV-1 BaL gp120 prior to addition of diluted HIV-1 + plasma samples (containing HIV-1-specific IgG antibodies). Biotinylated recombinant dimeric FcγRIIIa was then added, followed by HRP-conjugated streptavidin. The wells were developed with TMB substrate, and absorbance was measured at 450 nm. (C) Levels of gp120-specific antibodies that bind dimeric FcγRIIIa pre- and post-panobinostat treatment. The cross-linking of recombinant FcγRIIIa was used as a surrogate measure of antibody Fc-mediated effector function. Statistical analyses between matched pairs were performed using the Wilcoxon signed-rank test.

    Journal: Journal of Virology

    Article Title: Anti-HIV-1 ADCC Antibodies following Latency Reversal and Treatment Interruption

    doi: 10.1128/JVI.00603-17

    Figure Lengend Snippet: HIV-1-specific antibody responses following panobinostat treatment. (A) Concentrations of gp120-specific IgG antibodies pre- and post-panobinostat treatment. (B) ELISA to measure dimeric FcγRIIIa binding. Ninety-six-well ELISA plates were first coated with 50 ng/well of HIV-1 BaL gp120 prior to addition of diluted HIV-1 + plasma samples (containing HIV-1-specific IgG antibodies). Biotinylated recombinant dimeric FcγRIIIa was then added, followed by HRP-conjugated streptavidin. The wells were developed with TMB substrate, and absorbance was measured at 450 nm. (C) Levels of gp120-specific antibodies that bind dimeric FcγRIIIa pre- and post-panobinostat treatment. The cross-linking of recombinant FcγRIIIa was used as a surrogate measure of antibody Fc-mediated effector function. Statistical analyses between matched pairs were performed using the Wilcoxon signed-rank test.

    Article Snippet: Biotinylated recombinant dimeric FcγRIIIa was added, followed by horseradish peroxidase (HRP)-conjugated streptavidin (Thermo Fisher Scientific).

    Techniques: Enzyme-linked Immunosorbent Assay, Binding Assay, Recombinant