hrp conjugated anti human igg  (Jackson Immuno)

 
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94
    Name:
    Peroxidase AffiniPure Goat Anti Human IgG
    Description:
    Whole IgG antibodies are isolated as intact molecules from antisera by immunoaffinity chromatography They have an Fc portion and two antigen binding Fab portions joined together by disulfide bonds and therefore they are divalent The average molecular weight is reported to be about 160 kDa The whole IgG form of antibodies is suitable for the majority of immunodetection procedures and is the most cost effective Based on immunoelectrophoresis and or ELISA the antibody reacts with whole molecule human IgG It also reacts with the light chains of other human immunoglobulins No antibody was detected against non immunoglobulin serum proteins The antibody may cross react with immunoglobulins from other species
    Catalog Number:
    109-035-003
    Price:
    109
    Purity:
    The antibody was purified from antisera by immunoaffinity chromatography using antigens coupled to agarose beads.
    Conjugate:
    Horseradish Peroxidase
    Size:
    ml
    Category:
    Secondary Antibody
    Source:
    Goat
    Quantity:
    2 0
    Buy from Supplier


    Structured Review

    Jackson Immuno hrp conjugated anti human igg
    Peroxidase AffiniPure Goat Anti Human IgG
    Whole IgG antibodies are isolated as intact molecules from antisera by immunoaffinity chromatography They have an Fc portion and two antigen binding Fab portions joined together by disulfide bonds and therefore they are divalent The average molecular weight is reported to be about 160 kDa The whole IgG form of antibodies is suitable for the majority of immunodetection procedures and is the most cost effective Based on immunoelectrophoresis and or ELISA the antibody reacts with whole molecule human IgG It also reacts with the light chains of other human immunoglobulins No antibody was detected against non immunoglobulin serum proteins The antibody may cross react with immunoglobulins from other species
    https://www.bioz.com/result/hrp conjugated anti human igg/product/Jackson Immuno
    Average 94 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    hrp conjugated anti human igg - by Bioz Stars, 2020-08
    94/100 stars

    Images

    1) Product Images from "Direct recognition of the mycobacterial glycolipid, trehalose dimycolate, by C-type lectin Mincle"

    Article Title: Direct recognition of the mycobacterial glycolipid, trehalose dimycolate, by C-type lectin Mincle

    Journal: The Journal of Experimental Medicine

    doi: 10.1084/jem.20091750

    Purified TDM is recognized by Mincle. (A) Chemical structure of TDM. α-Mycolate (shown), methoxy-mycolate, and keto-mycolate are the major subclasses of mycolate found in M. tuberculosis TDM. (B and C) Reporter cells were stimulated with the indicated amount of plate-coated TDM (B) or TDB (C). (D) Reporter cells were stimulated with the indicated amount of TDM, methyl α-mycolate (α-mycolate), or methyl keto-mycolate (keto-mycolate). (E) Reporter cells were stimulated with the indicated amount of TDM and TMM. (F) ELISA-based detection of TDM by Mincle-Ig. hIgG1-Fc (Ig), Mincle-Ig, and Dectin2-Ig were incubated with 0.1 nmol/0.32 cm 2 of plate-coated TDM. Bound protein was detected with anti–hIgG-HRP followed by the addition of colorimetric substrate. (G) Effect of trehalose (100 µg/ml), EDTA (10 mM), rat IgG (10 µg/ml), and anti-Mincle mAb (10 µg/ml) on TDM recognition by Mincle-Ig. ELISA-based detection was performed as in E. (H) Reporter cells were stimulated with TDM, which was treated with trehalase as described in Materials and methods. Cells were also stimulated with plate-coated anti-Mincle mAb treated with trehalase as a negative control. All data are means ± SD for triplicate assays and representative results from three independent experiments with similar results are shown.
    Figure Legend Snippet: Purified TDM is recognized by Mincle. (A) Chemical structure of TDM. α-Mycolate (shown), methoxy-mycolate, and keto-mycolate are the major subclasses of mycolate found in M. tuberculosis TDM. (B and C) Reporter cells were stimulated with the indicated amount of plate-coated TDM (B) or TDB (C). (D) Reporter cells were stimulated with the indicated amount of TDM, methyl α-mycolate (α-mycolate), or methyl keto-mycolate (keto-mycolate). (E) Reporter cells were stimulated with the indicated amount of TDM and TMM. (F) ELISA-based detection of TDM by Mincle-Ig. hIgG1-Fc (Ig), Mincle-Ig, and Dectin2-Ig were incubated with 0.1 nmol/0.32 cm 2 of plate-coated TDM. Bound protein was detected with anti–hIgG-HRP followed by the addition of colorimetric substrate. (G) Effect of trehalose (100 µg/ml), EDTA (10 mM), rat IgG (10 µg/ml), and anti-Mincle mAb (10 µg/ml) on TDM recognition by Mincle-Ig. ELISA-based detection was performed as in E. (H) Reporter cells were stimulated with TDM, which was treated with trehalase as described in Materials and methods. Cells were also stimulated with plate-coated anti-Mincle mAb treated with trehalase as a negative control. All data are means ± SD for triplicate assays and representative results from three independent experiments with similar results are shown.

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

    2) Product Images from "A broad and potent H1-specific human monoclonal antibody produced in plants prevents influenza virus infection and transmission in guinea pigs"

    Article Title: A broad and potent H1-specific human monoclonal antibody produced in plants prevents influenza virus infection and transmission in guinea pigs

    Journal: bioRxiv

    doi: 10.1101/2020.01.13.902841

    In vitro characterization of KPF1-Antx hMAb. A-B) Purity and mobility of the KPF1-HEK and KPF1-Antx hMAbs: Equal amount of KPF1 hMAbs (2.500, 1.250, 0.625 and 0.313 μg) generated from either HEK293T cells (KPF1-HEK) or tobacco plant (KPF1-Antx) were separated by SDS-PAGE and evaluated by Coomassie blue staining ( A ) or Western blot using an anti-human IgG HRP-conjugated Ab ( B ). C) Recombinant HA binding of KPF1-HEK and the KPF1-Antx hMAbs: Binding to 1 μg /ml of indicated HA proteins was determined for serially diluted hMAbs by ELISA. n.s.: No significance. D) Avidity: Binding of hMAbs in the presence of increasing concentrations of urea was determined by ELISA. The statistical analysis between KPF1-HEK and KPF1-Antx (markers above the lines; between 1 μg/ml of KPF1-HEK and KPF1-Antx, marker below the lines: between 0.1 µg/ml of KPF1-HEK and KPF1-Antx), * p
    Figure Legend Snippet: In vitro characterization of KPF1-Antx hMAb. A-B) Purity and mobility of the KPF1-HEK and KPF1-Antx hMAbs: Equal amount of KPF1 hMAbs (2.500, 1.250, 0.625 and 0.313 μg) generated from either HEK293T cells (KPF1-HEK) or tobacco plant (KPF1-Antx) were separated by SDS-PAGE and evaluated by Coomassie blue staining ( A ) or Western blot using an anti-human IgG HRP-conjugated Ab ( B ). C) Recombinant HA binding of KPF1-HEK and the KPF1-Antx hMAbs: Binding to 1 μg /ml of indicated HA proteins was determined for serially diluted hMAbs by ELISA. n.s.: No significance. D) Avidity: Binding of hMAbs in the presence of increasing concentrations of urea was determined by ELISA. The statistical analysis between KPF1-HEK and KPF1-Antx (markers above the lines; between 1 μg/ml of KPF1-HEK and KPF1-Antx, marker below the lines: between 0.1 µg/ml of KPF1-HEK and KPF1-Antx), * p

    Techniques Used: In Vitro, Generated, SDS Page, Staining, Western Blot, Recombinant, Binding Assay, Enzyme-linked Immunosorbent Assay, Marker

    Related Articles

    Enzyme-linked Immunosorbent Assay:

    Article Title: An antidote approach to reduce risk and broaden utility of antibody-based therapeutics
    Article Snippet: .. Washed plates were next blocked with 5% blocking reagent (Bio-Rad catalog no. 1706404) for 2 h. This was followed by the introduction of serial antibody dilutions in 2.5% blocking reagent for 1 h. After washes to remove non-bound proteins, AffiniPure goat anti-human IgG1 (H+L) HRP antibody (Jackson ImmunoResearch catalog no. 109-035-088) was added at a 1:10,000 dilution in 2.5% blocking reagent for 1 h. For detection of the bound antibodies, the washed plates were incubated with Pierce Supersignal ELISA Pico Chemiluminescent substrate (Thermo Fisher catalog no. 37069) for 5 min at room temperature followed by chemiluminescent detection using an EnVision plate reader (PerkinElmer Life Sciences). ..

    Incubation:

    Article Title: Unique binding modes for the broad neutralizing activity of single-chain variable fragments (scFv) targeting CD4-induced epitopes
    Article Snippet: .. After incubation at 37 °C for 1 h, peroxidase-conjugated AffiniPure F(ab′)2 Fragment Goat Anti-Human IgG (H+L) (1:2500 dilution; Jackson ImmunoResearch) was added to each well, and incubated at RT for 1 h. Finally, the wells were reacted with 100 µl ABTS [2,2′-azinobis-(3-ethylbenzthiazoline sulfonic acid)] solution (Roche Molecular Biochemicals) and the absorbance at 405 nm was measured using a microplate reader. ..

    Article Title: A human monoclonal antibody targeting the stem cell factor receptor (c-Kit) blocks tumor cell signaling and inhibits tumor growth
    Article Snippet: .. Goat anti-human IgG antibody HRP conjugate (Jackson ImmunoResearch Laboratories; 109-035-088) was added to each well at 1:6000 dilution in 2% milk/PBS/T and incubated for 1 h at room temperature. .. After washing the plate, color was developed by adding 100 μL TMB peroxidase substrate from KPL and stopped with 100 μL of 2 N H2 SO4 stop solution (R & D Systems).

    Article Title: An antidote approach to reduce risk and broaden utility of antibody-based therapeutics
    Article Snippet: .. Washed plates were next blocked with 5% blocking reagent (Bio-Rad catalog no. 1706404) for 2 h. This was followed by the introduction of serial antibody dilutions in 2.5% blocking reagent for 1 h. After washes to remove non-bound proteins, AffiniPure goat anti-human IgG1 (H+L) HRP antibody (Jackson ImmunoResearch catalog no. 109-035-088) was added at a 1:10,000 dilution in 2.5% blocking reagent for 1 h. For detection of the bound antibodies, the washed plates were incubated with Pierce Supersignal ELISA Pico Chemiluminescent substrate (Thermo Fisher catalog no. 37069) for 5 min at room temperature followed by chemiluminescent detection using an EnVision plate reader (PerkinElmer Life Sciences). ..

    Article Title: Designed Ankyrin Repeat Protein (DARPin) Neutralizers of TcdB from Clostridium difficile Ribotype 027
    Article Snippet: .. After blocking and thorough washing, CSPG4-EC-GFP (1 nM), a GFP-tagged extracellular domain of CSPG4 ( ); FZD2-Fc (R & D Systems catalog no. 1307-FZ-050) (4 nM); or PVRL3 (Sino Biological catalog no. 10852-H08H) (100 nM) was added to each well alone or in the presence of the relevant DARPin (250 nM) followed by incubation at room temperature for 2 h. After thorough washing, the amount of bound CSPG4-EC-GFP was determined using rabbit anti-GFP antibody (Proteintech catalog no. 50430-2-AP) (0.08 μg/ml) plus HRP-conjugated goat anti-rabbit antibody (Santa Cruz Biotechnology catalog no. SC-2004) (0.8 μg/ml), and the bound FZD2-Fc amount was determined using HRP-conjugated goat anti-human antibody (Jackson Immuno Research catalog no. 109-035-088) (0.2 μg/ml), while the bound PVRL3 was detected using a goat anti-PVRL3 antibody (R & D Systems catalog no. AF3064) plus HRP-conjugated donkey anti-goat antibody (Santa Cruz Biotechnology catalog no. SC-2020) followed by color development using TMB. .. A homology model of TcdBUK1 Frizzled binding domain (FBD; amino acids 1284 to 1803) was constructed using SWISS-MODEL and was overlaid onto the FBD of TcdBVPI (PDB code: 6C0B ).

    Article Title: A Neuronal Culture System to Detect Prion Synaptotoxicity
    Article Snippet: .. Membranes were blocked for 1 h in 5% (w/v) non-fat dry milk in Tris-buffered saline containing 0.1% Tween 20, followed by incubation with anti-PrP antibody D18 (a human chimeric monoclonal[ ]), and then with HRP-conjugated anti-human secondary antibody (Jackson ImmunoResearch, cat. no. 109-035-088). .. Signals were revealed using HRP substrate (Millipore, cat. no. WBKLS0500), and were visualized using a BioRad XRS image scanner.

    other:

    Article Title: Direct recognition of the mycobacterial glycolipid, trehalose dimycolate, by C-type lectin Mincle
    Article Snippet: HRP-conjugated anti–human IgG (109–035-088) was from Jackson ImmunoResearch Laboratories.

    Blocking Assay:

    Article Title: An antidote approach to reduce risk and broaden utility of antibody-based therapeutics
    Article Snippet: .. Washed plates were next blocked with 5% blocking reagent (Bio-Rad catalog no. 1706404) for 2 h. This was followed by the introduction of serial antibody dilutions in 2.5% blocking reagent for 1 h. After washes to remove non-bound proteins, AffiniPure goat anti-human IgG1 (H+L) HRP antibody (Jackson ImmunoResearch catalog no. 109-035-088) was added at a 1:10,000 dilution in 2.5% blocking reagent for 1 h. For detection of the bound antibodies, the washed plates were incubated with Pierce Supersignal ELISA Pico Chemiluminescent substrate (Thermo Fisher catalog no. 37069) for 5 min at room temperature followed by chemiluminescent detection using an EnVision plate reader (PerkinElmer Life Sciences). ..

    Article Title: Designed Ankyrin Repeat Protein (DARPin) Neutralizers of TcdB from Clostridium difficile Ribotype 027
    Article Snippet: .. After blocking and thorough washing, CSPG4-EC-GFP (1 nM), a GFP-tagged extracellular domain of CSPG4 ( ); FZD2-Fc (R & D Systems catalog no. 1307-FZ-050) (4 nM); or PVRL3 (Sino Biological catalog no. 10852-H08H) (100 nM) was added to each well alone or in the presence of the relevant DARPin (250 nM) followed by incubation at room temperature for 2 h. After thorough washing, the amount of bound CSPG4-EC-GFP was determined using rabbit anti-GFP antibody (Proteintech catalog no. 50430-2-AP) (0.08 μg/ml) plus HRP-conjugated goat anti-rabbit antibody (Santa Cruz Biotechnology catalog no. SC-2004) (0.8 μg/ml), and the bound FZD2-Fc amount was determined using HRP-conjugated goat anti-human antibody (Jackson Immuno Research catalog no. 109-035-088) (0.2 μg/ml), while the bound PVRL3 was detected using a goat anti-PVRL3 antibody (R & D Systems catalog no. AF3064) plus HRP-conjugated donkey anti-goat antibody (Santa Cruz Biotechnology catalog no. SC-2020) followed by color development using TMB. .. A homology model of TcdBUK1 Frizzled binding domain (FBD; amino acids 1284 to 1803) was constructed using SWISS-MODEL and was overlaid onto the FBD of TcdBVPI (PDB code: 6C0B ).

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 91
    Jackson Immuno hrp conjugated goat anti human igg fcγ antibodies
    Removal of inhibitory domains from recombinant pro-antibodies by MMP-2. Unmodified anti-EGFR antibody ( A ), CBa-anti-EGFR antibody ( B ), C2b-anti-EGFR antibody ( C ) or LAP-anti-EGFR antibody ( D ) were digested by MMP-2 for the indicated time. Digested antibodies were resolved by reducing SDS-PAGE and analyzed by western blot using <t>HRP-conjugated</t> goat anti <t>human-IgG</t> <t>Fcγ</t> antibodies. Note that amounts of cut (mature form) antibodies increased as digestion was prolonged from 1 minute to 60 minutes. ID-HC: inhibitory domain-heavy chain. HC: heavy chain. with MMP: with MMP-2 incubation. The original blots image for panel A–D are presented in Supplementary Figure S5 .
    Hrp Conjugated Goat Anti Human Igg Fcγ Antibodies, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hrp conjugated goat anti human igg fcγ antibodies/product/Jackson Immuno
    Average 91 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    hrp conjugated goat anti human igg fcγ antibodies - by Bioz Stars, 2020-08
    91/100 stars
      Buy from Supplier

    94
    Jackson Immuno hrp conjugated anti human igg
    Purified TDM is recognized by Mincle. (A) Chemical structure of TDM. α-Mycolate (shown), methoxy-mycolate, and keto-mycolate are the major subclasses of mycolate found in M. tuberculosis TDM. (B and C) Reporter cells were stimulated with the indicated amount of plate-coated TDM (B) or TDB (C). (D) Reporter cells were stimulated with the indicated amount of TDM, methyl α-mycolate (α-mycolate), or methyl keto-mycolate (keto-mycolate). (E) Reporter cells were stimulated with the indicated amount of TDM and TMM. (F) ELISA-based detection of TDM by Mincle-Ig. hIgG1-Fc (Ig), Mincle-Ig, and Dectin2-Ig were incubated with 0.1 nmol/0.32 cm 2 of plate-coated TDM. Bound protein was detected with <t>anti–hIgG-HRP</t> followed by the addition of colorimetric substrate. (G) Effect of trehalose (100 µg/ml), EDTA (10 mM), rat <t>IgG</t> (10 µg/ml), and anti-Mincle mAb (10 µg/ml) on TDM recognition by Mincle-Ig. ELISA-based detection was performed as in E. (H) Reporter cells were stimulated with TDM, which was treated with trehalase as described in Materials and methods. Cells were also stimulated with plate-coated anti-Mincle mAb treated with trehalase as a negative control. All data are means ± SD for triplicate assays and representative results from three independent experiments with similar results are shown.
    Hrp Conjugated Anti Human Igg, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 94/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hrp conjugated anti human igg/product/Jackson Immuno
    Average 94 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    hrp conjugated anti human igg - by Bioz Stars, 2020-08
    94/100 stars
      Buy from Supplier

    92
    Jackson Immuno horseradish peroxidase hrp conjugated anti human igg antibodies
    Optimization of ELISA for α-syn AAb detection Dilute serum was applied to 384-well ELISA plates either coated with recombinant α-syn or not coated. (A) Plates were blocked with either 1% cold fish gelatin (top row) or 5% bovine serum albumin (middle row) overnight at 4 °C. Binding was detected using anti-human <t>IgG</t> <t>HRP-conjugates</t> raised in rabbit (left column), donkey (middle column), or goat (right column). (B) Plates were blocked with 1% cold fish gelatin at 4°C for either 6 days (left), 4 days (middle), or 1 day (right). Additional blocking solutions (1% bovine serum albumin, 5% milk in PBS, several commercial blocking solutions) did not demonstrate any improvement over 1% cold fish gelatin. Each data point represents three technical replicates (n=3 wells). Wells coated with α-syn denoted by closed circles. Uncoated wells denoted by closed triangles. BSA = bovine serum albumin; CFG = cold fish gelatin; HRP = horseradish peroxidase conjugate.
    Horseradish Peroxidase Hrp Conjugated Anti Human Igg Antibodies, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/horseradish peroxidase hrp conjugated anti human igg antibodies/product/Jackson Immuno
    Average 92 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    horseradish peroxidase hrp conjugated anti human igg antibodies - by Bioz Stars, 2020-08
    92/100 stars
      Buy from Supplier

    85
    Jackson Immuno pe conjugated mouse anti human igg4
    ALBIA (Luminex) <t>IgG.</t> Panel A: The median of ALBIA fluorescence units of anti-PLA 2 R positive IMN samples were compared to IMN samples negative for anti-PLA 2 R antibodies and control samples (normal healthy controls and unrelated inflammatory disease controls).Fluorescence values of anti-PLA 2 R positive samples were significantly higher than values of anti-PLA 2 R negative samples and of controls (p-values
    Pe Conjugated Mouse Anti Human Igg4, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pe conjugated mouse anti human igg4/product/Jackson Immuno
    Average 85 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pe conjugated mouse anti human igg4 - by Bioz Stars, 2020-08
    85/100 stars
      Buy from Supplier

    Image Search Results


    Removal of inhibitory domains from recombinant pro-antibodies by MMP-2. Unmodified anti-EGFR antibody ( A ), CBa-anti-EGFR antibody ( B ), C2b-anti-EGFR antibody ( C ) or LAP-anti-EGFR antibody ( D ) were digested by MMP-2 for the indicated time. Digested antibodies were resolved by reducing SDS-PAGE and analyzed by western blot using HRP-conjugated goat anti human-IgG Fcγ antibodies. Note that amounts of cut (mature form) antibodies increased as digestion was prolonged from 1 minute to 60 minutes. ID-HC: inhibitory domain-heavy chain. HC: heavy chain. with MMP: with MMP-2 incubation. The original blots image for panel A–D are presented in Supplementary Figure S5 .

    Journal: Scientific Reports

    Article Title: Selective antibody activation through protease-activated pro-antibodies that mask binding sites with inhibitory domains

    doi: 10.1038/s41598-017-11886-7

    Figure Lengend Snippet: Removal of inhibitory domains from recombinant pro-antibodies by MMP-2. Unmodified anti-EGFR antibody ( A ), CBa-anti-EGFR antibody ( B ), C2b-anti-EGFR antibody ( C ) or LAP-anti-EGFR antibody ( D ) were digested by MMP-2 for the indicated time. Digested antibodies were resolved by reducing SDS-PAGE and analyzed by western blot using HRP-conjugated goat anti human-IgG Fcγ antibodies. Note that amounts of cut (mature form) antibodies increased as digestion was prolonged from 1 minute to 60 minutes. ID-HC: inhibitory domain-heavy chain. HC: heavy chain. with MMP: with MMP-2 incubation. The original blots image for panel A–D are presented in Supplementary Figure S5 .

    Article Snippet: After the cells were washed with DMEM (200 μL/well) three times to remove unbound antibodies, 1 μg/ml of HRP-conjugated goat anti-human IgG Fcγ antibodies (Jackson ImmunoResearch Laboratories) were added to the cells (50 μL/well) at room temperature for 1 hour.

    Techniques: Recombinant, Crocin Bleaching Assay, SDS Page, Western Blot, Incubation

    Recovery of binding activities of the recombinant antibodies after removal of inhibitory domains by MMP-2. CBa-anti-EGFR antibody ( A ), C2b-anti-EGFR antibody ( B ) or LAP-anti-EGFR antibody ( C ) that were digested by MMP-2 for indicated time were added to microtiter plates pre-coated with MDA-MB-468 cells. Degree of antibody binding to the cells was compared to the binding of the cells by the unmodified anti-EGFR antibody (set as 100%). The LAP domain reduced antibody binding activity by 53.8%. The CBa and C2b domains masked antibody binding sites ineffectively, only marginally reducing the antibody binding activity by 9.3% and 21%, respectively. Note that after MMP-2 digestion of the LAP-anti-EGFR antibody, the binding activity rose progressively from 46.2% to 100%. ( D ) The binding activities of MMP-2 substrate peptide-linked anti-EGFR antibody (GPLGVR-anti-EGFR Ab) and MMP-2 substrate peptide-removed anti-EGFR antibody (GPLGVR-anti-EGFR Ab + MMP) showed the MMP-2 substrate peptide does not affect the binding activity of the antibody. ( E ) MMP-2 digested antibodies were resolved by reducing SDS-PAGE and analyzed by western blot using HRP-conjugated goat anti human-IgG Fcγ antibodies. GPLGVR peptide was indeed removed from the heavy chains of anti-EGFR antibodies by MMP-2, as indicated by reduction of molecular size. GPLGVR-HC: MMP-2 substrate peptide-linked heavy chain. HC: heavy chain. The original blot image for panel E is presented in Supplementary Figure S6 .

    Journal: Scientific Reports

    Article Title: Selective antibody activation through protease-activated pro-antibodies that mask binding sites with inhibitory domains

    doi: 10.1038/s41598-017-11886-7

    Figure Lengend Snippet: Recovery of binding activities of the recombinant antibodies after removal of inhibitory domains by MMP-2. CBa-anti-EGFR antibody ( A ), C2b-anti-EGFR antibody ( B ) or LAP-anti-EGFR antibody ( C ) that were digested by MMP-2 for indicated time were added to microtiter plates pre-coated with MDA-MB-468 cells. Degree of antibody binding to the cells was compared to the binding of the cells by the unmodified anti-EGFR antibody (set as 100%). The LAP domain reduced antibody binding activity by 53.8%. The CBa and C2b domains masked antibody binding sites ineffectively, only marginally reducing the antibody binding activity by 9.3% and 21%, respectively. Note that after MMP-2 digestion of the LAP-anti-EGFR antibody, the binding activity rose progressively from 46.2% to 100%. ( D ) The binding activities of MMP-2 substrate peptide-linked anti-EGFR antibody (GPLGVR-anti-EGFR Ab) and MMP-2 substrate peptide-removed anti-EGFR antibody (GPLGVR-anti-EGFR Ab + MMP) showed the MMP-2 substrate peptide does not affect the binding activity of the antibody. ( E ) MMP-2 digested antibodies were resolved by reducing SDS-PAGE and analyzed by western blot using HRP-conjugated goat anti human-IgG Fcγ antibodies. GPLGVR peptide was indeed removed from the heavy chains of anti-EGFR antibodies by MMP-2, as indicated by reduction of molecular size. GPLGVR-HC: MMP-2 substrate peptide-linked heavy chain. HC: heavy chain. The original blot image for panel E is presented in Supplementary Figure S6 .

    Article Snippet: After the cells were washed with DMEM (200 μL/well) three times to remove unbound antibodies, 1 μg/ml of HRP-conjugated goat anti-human IgG Fcγ antibodies (Jackson ImmunoResearch Laboratories) were added to the cells (50 μL/well) at room temperature for 1 hour.

    Techniques: Binding Assay, Recombinant, Crocin Bleaching Assay, Multiple Displacement Amplification, Activity Assay, SDS Page, Western Blot

    Purified TDM is recognized by Mincle. (A) Chemical structure of TDM. α-Mycolate (shown), methoxy-mycolate, and keto-mycolate are the major subclasses of mycolate found in M. tuberculosis TDM. (B and C) Reporter cells were stimulated with the indicated amount of plate-coated TDM (B) or TDB (C). (D) Reporter cells were stimulated with the indicated amount of TDM, methyl α-mycolate (α-mycolate), or methyl keto-mycolate (keto-mycolate). (E) Reporter cells were stimulated with the indicated amount of TDM and TMM. (F) ELISA-based detection of TDM by Mincle-Ig. hIgG1-Fc (Ig), Mincle-Ig, and Dectin2-Ig were incubated with 0.1 nmol/0.32 cm 2 of plate-coated TDM. Bound protein was detected with anti–hIgG-HRP followed by the addition of colorimetric substrate. (G) Effect of trehalose (100 µg/ml), EDTA (10 mM), rat IgG (10 µg/ml), and anti-Mincle mAb (10 µg/ml) on TDM recognition by Mincle-Ig. ELISA-based detection was performed as in E. (H) Reporter cells were stimulated with TDM, which was treated with trehalase as described in Materials and methods. Cells were also stimulated with plate-coated anti-Mincle mAb treated with trehalase as a negative control. All data are means ± SD for triplicate assays and representative results from three independent experiments with similar results are shown.

    Journal: The Journal of Experimental Medicine

    Article Title: Direct recognition of the mycobacterial glycolipid, trehalose dimycolate, by C-type lectin Mincle

    doi: 10.1084/jem.20091750

    Figure Lengend Snippet: Purified TDM is recognized by Mincle. (A) Chemical structure of TDM. α-Mycolate (shown), methoxy-mycolate, and keto-mycolate are the major subclasses of mycolate found in M. tuberculosis TDM. (B and C) Reporter cells were stimulated with the indicated amount of plate-coated TDM (B) or TDB (C). (D) Reporter cells were stimulated with the indicated amount of TDM, methyl α-mycolate (α-mycolate), or methyl keto-mycolate (keto-mycolate). (E) Reporter cells were stimulated with the indicated amount of TDM and TMM. (F) ELISA-based detection of TDM by Mincle-Ig. hIgG1-Fc (Ig), Mincle-Ig, and Dectin2-Ig were incubated with 0.1 nmol/0.32 cm 2 of plate-coated TDM. Bound protein was detected with anti–hIgG-HRP followed by the addition of colorimetric substrate. (G) Effect of trehalose (100 µg/ml), EDTA (10 mM), rat IgG (10 µg/ml), and anti-Mincle mAb (10 µg/ml) on TDM recognition by Mincle-Ig. ELISA-based detection was performed as in E. (H) Reporter cells were stimulated with TDM, which was treated with trehalase as described in Materials and methods. Cells were also stimulated with plate-coated anti-Mincle mAb treated with trehalase as a negative control. All data are means ± SD for triplicate assays and representative results from three independent experiments with similar results are shown.

    Article Snippet: HRP-conjugated anti–human IgG (109–035-088) was from Jackson ImmunoResearch Laboratories.

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

    Optimization of ELISA for α-syn AAb detection Dilute serum was applied to 384-well ELISA plates either coated with recombinant α-syn or not coated. (A) Plates were blocked with either 1% cold fish gelatin (top row) or 5% bovine serum albumin (middle row) overnight at 4 °C. Binding was detected using anti-human IgG HRP-conjugates raised in rabbit (left column), donkey (middle column), or goat (right column). (B) Plates were blocked with 1% cold fish gelatin at 4°C for either 6 days (left), 4 days (middle), or 1 day (right). Additional blocking solutions (1% bovine serum albumin, 5% milk in PBS, several commercial blocking solutions) did not demonstrate any improvement over 1% cold fish gelatin. Each data point represents three technical replicates (n=3 wells). Wells coated with α-syn denoted by closed circles. Uncoated wells denoted by closed triangles. BSA = bovine serum albumin; CFG = cold fish gelatin; HRP = horseradish peroxidase conjugate.

    Journal: Journal of neurochemistry

    Article Title: Measurements of auto-antibodies to α-synuclein in the serum and cerebral spinal fluids of patients with Parkinson’s disease

    doi: 10.1111/jnc.14330

    Figure Lengend Snippet: Optimization of ELISA for α-syn AAb detection Dilute serum was applied to 384-well ELISA plates either coated with recombinant α-syn or not coated. (A) Plates were blocked with either 1% cold fish gelatin (top row) or 5% bovine serum albumin (middle row) overnight at 4 °C. Binding was detected using anti-human IgG HRP-conjugates raised in rabbit (left column), donkey (middle column), or goat (right column). (B) Plates were blocked with 1% cold fish gelatin at 4°C for either 6 days (left), 4 days (middle), or 1 day (right). Additional blocking solutions (1% bovine serum albumin, 5% milk in PBS, several commercial blocking solutions) did not demonstrate any improvement over 1% cold fish gelatin. Each data point represents three technical replicates (n=3 wells). Wells coated with α-syn denoted by closed circles. Uncoated wells denoted by closed triangles. BSA = bovine serum albumin; CFG = cold fish gelatin; HRP = horseradish peroxidase conjugate.

    Article Snippet: Secondary antibodies used were horseradish peroxidase (HRP)-conjugated anti-human IgG antibodies raised in rabbit (RRID:AB_2339653, Jackson Immunoresearch, cat. 309-035-082, West Grove, PA), donkey (RRID:AB_2340495, Jackson Immunoresearch, cat. 709-035-149), or goat (RRID:AB_257868, Sigma-Aldrich, cat. A0170, St. Louis, MO).

    Techniques: Enzyme-linked Immunosorbent Assay, Recombinant, Fluorescence In Situ Hybridization, Binding Assay, Blocking Assay

    ALBIA (Luminex) IgG. Panel A: The median of ALBIA fluorescence units of anti-PLA 2 R positive IMN samples were compared to IMN samples negative for anti-PLA 2 R antibodies and control samples (normal healthy controls and unrelated inflammatory disease controls).Fluorescence values of anti-PLA 2 R positive samples were significantly higher than values of anti-PLA 2 R negative samples and of controls (p-values

    Journal: PLoS ONE

    Article Title: An Anti-Phospholipase A2 Receptor Quantitative Immunoassay and Epitope Analysis in Membranous Nephropathy Reveals Different Antigenic Domains of the Receptor

    doi: 10.1371/journal.pone.0061669

    Figure Lengend Snippet: ALBIA (Luminex) IgG. Panel A: The median of ALBIA fluorescence units of anti-PLA 2 R positive IMN samples were compared to IMN samples negative for anti-PLA 2 R antibodies and control samples (normal healthy controls and unrelated inflammatory disease controls).Fluorescence values of anti-PLA 2 R positive samples were significantly higher than values of anti-PLA 2 R negative samples and of controls (p-values

    Article Snippet: 40 µl of PE conjugated goat anti-human IgG (1∶50/HRP sample diluent, Jackson ImmunoResearch) or 40 µl of PE conjugated mouse anti-human IgG4 (1∶25 in HRP sample diluents, Jackson ImmunoResearch) was then added and the plate was incubated for an additional 30 min at room temperature.

    Techniques: Luminex, Fluorescence, Proximity Ligation Assay

    Epitope Mapping of PLA 2 R Peptides with anti-human IgG 4 . Grey scale heat map representation of results from SPOT used to detect PLA 2 R epitopes. Peptide membranes were probed with 10 randomly selected IMN samples that had previously been tested by IIF-CBA for anti-PLA 2 R antibodies (7 positive (IMN+) and 3 negative (IMN-)), as well as 5 normal healthy controls (NHC). The positive control rabbit antibody to PLA 2 R reacted with the expected peptide used as the immunogen. Consensus epitopes and their respective PLA 2 R domains derived from this analysis are illustrated in Figure 3 and summarized in Table 1 .

    Journal: PLoS ONE

    Article Title: An Anti-Phospholipase A2 Receptor Quantitative Immunoassay and Epitope Analysis in Membranous Nephropathy Reveals Different Antigenic Domains of the Receptor

    doi: 10.1371/journal.pone.0061669

    Figure Lengend Snippet: Epitope Mapping of PLA 2 R Peptides with anti-human IgG 4 . Grey scale heat map representation of results from SPOT used to detect PLA 2 R epitopes. Peptide membranes were probed with 10 randomly selected IMN samples that had previously been tested by IIF-CBA for anti-PLA 2 R antibodies (7 positive (IMN+) and 3 negative (IMN-)), as well as 5 normal healthy controls (NHC). The positive control rabbit antibody to PLA 2 R reacted with the expected peptide used as the immunogen. Consensus epitopes and their respective PLA 2 R domains derived from this analysis are illustrated in Figure 3 and summarized in Table 1 .

    Article Snippet: 40 µl of PE conjugated goat anti-human IgG (1∶50/HRP sample diluent, Jackson ImmunoResearch) or 40 µl of PE conjugated mouse anti-human IgG4 (1∶25 in HRP sample diluents, Jackson ImmunoResearch) was then added and the plate was incubated for an additional 30 min at room temperature.

    Techniques: Proximity Ligation Assay, Crocin Bleaching Assay, Positive Control, Derivative Assay