Rabbit anti human igg hrp

Manufactured by Agilent Technologies

Sourced in Denmark

Rabbit anti-human IgG-HRP is a horseradish peroxidase (HRP)-conjugated secondary antibody that binds to human immunoglobulin G (IgG) antibodies. It is commonly used in various immunoassay and immunodetection techniques to facilitate the detection and quantification of human IgG proteins.

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Lab products found in correlation

Eia ria plate, by Corning (2 mentions) Tmb solution, by Thermo Fisher Scientific (2 mentions) Multiskan microplate spectrophotometer, by Thermo Fisher Scientific (2 mentions) Maxisorp plate, by Thermo Fisher Scientific (2 mentions) Human reference serum, by Fortis Life Sciences (1 mentions) Igκ clone g20 193, by BD (1 mentions) Bond epitope retrieval solution 2, by Leica (1 mentions) Bond polymer refine detection, by Leica (1 mentions) Immunolon plates, by Thermo Fisher Scientific (1 mentions) Superblock, by Thermo Fisher Scientific (1 mentions) Bovine serum albumin (bsa), by Merck Group (1 mentions) Glomax discovery system, by Promega (1 mentions) Goat anti mouse igg hrp, by Agilent Technologies (1 mentions) Supersignal elisa pico chemiluminescent substrate, by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

Rabbit anti-human IgG (13 citations) HRP-conjugated rabbit anti-human IgG (9 citations) Anti-human IgG-HRP (5 citations) HRP-conjugated polyclonal rabbit anti-human IgG (3 citations) HRP-conjugated rabbit anti-human IgG antibody (3 citations) HRP-labelled rabbit anti-human IgG antibodies (2 citations) Polyclonal rabbit anti-human IgG/HRP (2 citations) HRP rabbit anti-human IgG secondary antibody (2 citations) HRP)-conjugated polyclonal rabbit anti-human-IgG antibody (2 citations) Anti-TM (1 citations)

10 protocols using rabbit anti human igg hrp

Assessing Light Chain Composition of ACPA in RA

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The presence of light chains (LC) in ACPA-IgG was assessed by incubation of isolated ACPA or ACPA-depleted serum (n = 87 patients) and SF (n = 21 patients) on ELISA plates coated with mouse anti-human immunoglobulin lambda LC (Igλ; clone JDC-12, BD Pharmingen) or mouse anti-human immunoglobulin kappa LC (Igκ; clone G20-193, BD Pharmingen) monoclonal antibodies. ACPA bound by the respective anti-LC antibodies were detected with polyclonal rabbit anti-human IgG-HRP (DAKO). Human reference serum (Bethyl Laboratories) was used as standard in all ELISA experiments and for data normalization.
ACPA-Ig levels in citrullinated antigen-binding B-cell cultures were based on reactivity towards the CCP2-peptide. Biotinylated CCP2-peptides were coupled to streptavidin-coated ELISA plates, followed by incubation with 1:2 diluted supernatant from cultures. ACPA-Ig levels were detected with secondary antibodies for IgG (rabbit anti-human IgG-HRP, DAKO), IgM (goat anti-human IgM-HRP, Millipore) or IgA (goat anti-human IgA-HRP, Invitrogen) to identify Ig-isotypes. The presence of tetanus toxoid (TT)-specific antibodies in supernatants of TT-specific B-cell cultures was determined using TT-coated (Staten Serum Institute) plates. Detection of TT-specific antibodies was identical to Ig-detection of ACPA.

Slot L.M., Vergroesen R.D., Kerkman P.F., Staudinger E., Reijm S., van Dooren H.J., van der Voort E.I., Huizinga T.W., Toes R.E, & Scherer H.U. (2021). Light chain skewing in autoantibodies and B-cell receptors of the citrullinated antigen-binding B-cell response in rheumatoid arthritis. PLoS ONE, 16(3), e0247847.

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Immunohistochemical analysis of skin biopsies

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Skin punch biopsies were placed in 4% formaldehyde for 24 hours at room temperature, paraffin embedded and sectioned. All skin biopsies were stained with Masson’s Trichrome (MT) for the evaluation of epidermis. In addition, biopsies were stained by immunohistochemistry (IHC) with either rabbit anti-human CD3 (polyclonal, DAKO, 3 μg/ml), rabbit anti-KU80 (clone C48E7, Cell Signaling, 0.0355 μg/ml), rabbit anti-human IgG/HRP (polyclonal, DAKO, 13 μg/ml) or mouse anti-human ki67 (MIB-1, DAKO, 0.31 μg/ml). Prior to the immunohistochemical stainings, antigen-retrieval was performed over night at 60°C in BOND Epitope retrieval solution 2 (Leica Biosystems, Nussloch, Germany) followed by incubation in BondTM Wash Solution (Leica Biosystems, Nussloch, Germany) for 5 minutes at room temperature. Rabbit anti-human CD3, rabbit anti-K80 and mouse anti-human ki67 were detected with BOND Polymere Refine RED Detection (Leica Biosystems, Nussloch, Germany) with Fast Red. BrightVision Poly-AP-Anti Rabbit IgG (Immunologic, Duiven, Netherlands) was used as secondary antibody for rabbit anti-KU80. The rabbit anti-human IgG/HRP were detected with BOND Polymer Refine Detection (Leica Biosystems, Nussloch, Germany). The slides were mounted with DXP mountant (LEO Pharma, Ballerup, Denmark). Relevant blocking, positive, negative and isotype controls were used for all stainings.

Christensen P.K., Hansen A.K., Skov S., Engkilde K., Larsen J., Høyer-Hansen M.H, & Koch J. (2023). Sustaining the T-cell activity in xenografted psoriasis skin. PLOS ONE, 18(1), e0278390.

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Plasma HA-Specific IgG Quantification

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Plasma‐specific HA IgG was measured by ELISA. Briefly, EIA/RIA plates (Costar, St Louis, MO) were coated with HA antigen (2 μg mL^–1), blocked with 2% BSA and then incubated with diluted plasma samples (1:50). HA‐specific IgG was detected by adding rabbit anti‐human IgG HRP (Dako, Glostrup, Denmark). ELISA plates were developed using TMB solution (Life Technologies, Carlsbad, CA), and the reaction was stopped with 1 m HCl. Absorbance (OD450 nm) was measured using a Multiskan Microplate Spectrophotometer (Thermo Fisher). Serial dilutions of recombinant human IgG in separate wells on the same plate were performed for quantification of specific IgG.

Hartley G.E., Edwards E.S., Bosco J.J., Ojaimi S., Stirling R.G., Cameron P.U., Flanagan K., Plebanski M., Hogarth P.M., O’Hehir R.E, & van Zelm M.C. (2020). Influenza‐specific IgG1+ memory B‐cell numbers increase upon booster vaccination in healthy adults but not in patients with predominantly antibody deficiency. Clinical & Translational Immunology, 9(10), e1199.

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SARS-CoV-2 Antibody Detection by ELISA

Cited 1 time

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EIA/RIA plates (Costar, St Louis, MO) were coated with 2 μg/ml recombinant SARS-CoV-2 NCP or RBD or with hemagglutinin (HA) from influenza A/Michigan/08/2015 (AM15) overnight at 4°C (28 (link)). Plates were subsequently blocked with 3% BSA in PBS and incubated with diluted plasma samples, 1:30 for RBD and NCP and 1:50 for AM15. Antigen-specific IgG was detected by adding rabbit anti-human IgG HRP (Dako, Glostrup, Denmark). ELISA plates were developed using TMB solution (Life Technologies, Carlsbad, CA) and the reaction was stopped with 1 M HCl. Absorbance (OD450nm) was measured using a Multiskan Microplate Spectrophotometer (Thermo Fisher). Serial dilutions of recombinant human IgG (in-house made human Rituximab) in separate wells on the same plate were performed for quantification of specific IgG.

Hartley G.E., Edwards E.S., Aui P.M., Varese N., Stojanovic S., McMahon J., Peleg A.Y., Boo I., Drummer H.E., Hogarth P.M., O’Hehir R.E, & van Zelm M.C. (2020). Rapid generation of durable B cell memory to SARS-CoV-2 spike and nucleocapsid proteins in COVID-19 and convalescence. Science Immunology, 5(54), eabf8891.

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ELISA for PfEMP1 DBLβ IgG Reactivity

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ELISA was used to analyze IgG reactivity to the above PfEMP1 DBLβ domains. Briefly, Immunolon plates (Thermo Scientific) were coated with 2.5µg/mL of the recombinant proteins PF3D7_1150400 (PF11_0521) and PF3D7_0425800 (PFD1235w) diluted in ELISA carbonate coating buffer (Thermo Scientific)and incubated overnight at 4°C. The plates were then washed in PBST (1X PBS with 0.05% Tween 20) followed by blocking with PBST and 1% skimmed milk for 1h at room temperature. After washing with PBST, plasma samples (1:100 in blocking buffer) were added and incubated for 1h at room temperature. Binding was detected using rabbit anti-human IgG-HRP (Dako, 1:5000 in blocking buffer). Malaria naïve plasma samples were used as negative controls.

Kimingi H.W., Kinyua A.W., Achieng N.A., Wambui K.M., Mwangi S., Nguti R., Kivisi C.A., Jensen A.T., Bejon P., Kapulu M.C., Abdi A.I, & Kinyanjui S.M. (2022). Breadth of Antibodies to Plasmodium falciparum Variant Surface Antigens Is Associated With Immunity in a Controlled Human Malaria Infection Study. Frontiers in Immunology, 13, 894770.

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ICAM-1 Binding Assay with DBLβ

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ICAM-1 binding was assessed by ELISA using DBLβ domain-coated Maxisorp plates, with detection using ICAM-1^D1D5-Fc and rabbit anti-human IgG-HRP (Dako). A similar assay was used to assess the reactivity of human plasma samples to the DBLβ domains or to assess their efficacy at blocking ICAM-1 binding.

Lennartz F., Adams Y., Bengtsson A., Olsen R.W., Turner L., Ndam N.T., Ecklu-Mensah G., Moussiliou A., Ofori M.F., Gamain B., Lusingu J.P., Petersen J.E., Wang C.W., Nunes-Silva S., Jespersen J.S., Lau C.K., Theander T.G., Lavstsen T., Hviid L., Higgins M.K, & Jensen A.T. (2017). Structure-Guided Identification of a Family of Dual Receptor-Binding PfEMP1 that Is Associated with Cerebral Malaria. Cell Host & Microbe, 21(3), 403-414.

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Quantification of Serum Anti-Factor H IgG

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Serum anti-FH IgG level was determined by an ELISA method described previously (1 (link)), with some modifications applied as follows. Nunc MaxiSorp plates (Nunc, Roskilde, Denmark) were coated with 1 μg/ml purified human FH (Calbiochem, San Diego, CA, USA). Serum samples were added at a dilution of 1:200 following blocking with PBS-1% BSA. As secondary antibody a rabbit anti-human IgG-HRP was used (Dako, Glostrup, Denmark) followed by the detection of bound IgG using 3,3′,5,5′-tetramethylbenzidine (TMB) with optical density (OD) read at λ = 450 nm (reference at λ = 620 nm). The assay was calibrated to a sample obtained as a kind gift from Dr. Dragon-Durey, and the cutoff value (>110 AU/ml) was determined according to the mean + 2 SD of 80 healthy individuals.

Trojnár E., Józsi M., Uray K., Csuka D., Szilágyi Á., Milosevic D., Stojanović V.D., Spasojević B., Rusai K., Müller T., Arbeiter K., Kelen K., Szabó A.J., Reusz G.S., Hyvärinen S., Jokiranta T.S, & Prohászka Z. (2017). Analysis of Linear Antibody Epitopes on Factor H and CFHR1 Using Sera of Patients with Autoimmune Atypical Hemolytic Uremic Syndrome. Frontiers in Immunology, 8, 302.

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ELISA Protocol for gp350 Antibody Titration

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Immulon 4HBX High-binding 96-well plates (ThermoFisher Scientific) were coated with 0.25 μg/ml recombinant purified gp350 (suspension CHO cell line produced)19 (link) in PBS overnight at 4 °C (all subsequent steps were performed at room temperature). The plates were washed 4 times with PBST (PBS with 0.05% Tween-20) and blocked with Superblock (ThermoFisher Scientific) for 1 hour. Human serum samples along with positive and negative control serum were serially diluted 1:5 in dilution buffer containing 0.1% Superblock in PBST (1:10 to 1:97,656,250 for sample, 1:10 to 1:781,250 for positive control, and 1:10 to 1:1250 for negative control) at 100 μL/well and incubated for 1 hour. After washing, HRP-conjugated secondary antibody (Rabbit anti-human IgG HRP, Dako# P0214) was diluted to 1: 7,500 in dilution buffer and incubated for 1 hour, followed by 4 washes in PBST. TMB substrate (SIGMA) was added at 100 μL/well for colorimetric reading. After adding 1N HCl at 100μL/well to stop the reaction, the absorption was measured at OD450 on a SpectraMax plate reader. The antiserum endpoint titer was quantified as the reciprocal dilution factor using SoftmaxPro to calculate the 4-fold rise above the ELISA assay background. The lower limit of detection (LOD) of this assay is 10. Negative samples were assigned an artificial titer of 0.1 for graphing purpose.

Liu H., Gemmell L., Lin R., Zuo F., Balfour HH J.r., Woo J.C, & Hayes G.M. (2020). Development of an Improved Epstein-Barr Virus (EBV) Neutralizing Antibody Assay to Facilitate Development of a Prophylactic gp350-Subunit EBV Vaccine. Mediterranean Journal of Hematology and Infectious Diseases, 12(1), e2020016.

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Optimizing Antibody Conjugate Signals

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We tested different conjugates in varying concentrations to obtain the best signal of bound α-PLG on the plate coated with PLG. The conjugates tested were: Sigma goat anti-human IgG HRP 1/10000 and 1/20000, Sigma goat anti-human IgG AP 1/5000 and 1/10000, Dako rabbit anti-human IgG HRP 1/30000 and 1/50000.

Göçeroğlu A., Grenmyr E., Berden A.E., Hagen E.C., Bunch D., Sommarin Y., Bruijn J.A., Bajema I.M, & Wieslander J. (2018). Anti-plasminogen antibodies in ANCA-associated vasculitis: An optimized anti-plasminogen assay. PLoS ONE, 13(11), e0207064.

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ELISA for IgG Binding to E. coli O157:H7

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IgG binding to E. coli O157:H7 proteins and lipopolysaccharide was detected by enzyme-linked immunosorbent assay. To detect IgG binding to E. coli O157:H7 proteins, E. coli O157:H7 culture filtrates (concentration 10, 5, 2.5 and 1.25%), Shiga toxin 2 (1 μg/mL, Phoenix Lab, Tufts Medical Center, Boston, MA) or O157LPS (1 μg/mL, a gift from R. Johnson, Public Health Agency, Guelph, ON, Canada), diluted in PBS (pH 7.4) were coated on a white 96-well Maxisorp plate (Nunc, Roskilde, Denmark) overnight at 4°C. The plate was washed three times with PBS-Tween (Medicago, Uppsala, Sweden) and blocked with 1% bovine serum albumin (BSA) (Sigma-Aldrich) in PBS. For antibody binding to E. coli O157:H7 proteins, purified mouse or human IgG (described above, 1 μg/mL diluted in 1% BSA) or 1% BSA (negative control), were added to the plate and incubated for 1h at RT. The plate was washed as above and incubated for 1 h at RT with goat anti-mouse IgG HRP (1:1000, Dako, Glostrup, Denmark) or rabbit anti-human IgG HRP (1:1000, Dako). The plate was washed as above and IgG binding was detected using Super signal ELISA pico chemiluminescent substrate (Thermo Fisher Scientific) and measured using Glomax Discovery system (Promega, Madison, WI).

Tontanahal A., Sperandio V., Kovbasnjuk O., Loos S., Kristoffersson A.C., Karpman D, & Arvidsson I. (2022). IgG Binds Escherichia coli Serine Protease EspP and Protects Mice From E. coli O157:H7 Infection. Frontiers in Immunology, 13, 807959.

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