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Hrp conjugated anti human igg fc

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HRP-conjugated anti-human IgG Fc is a laboratory reagent used for the detection and quantification of human IgG Fc in various immunoassay applications. The product consists of horseradish peroxidase (HRP) enzyme conjugated to antibodies specific to the Fc region of human immunoglobulin G (IgG).

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Lab products found in correlation

High binding flat bottom 96 well plates, by Corning (4 mentions) Opteia substrate solution, by BD (4 mentions) Spectramax id3 microplate reader, by Molecular Devices (4 mentions)

Spelling variants of this product

Anti-human IgG-HRP (13 citations) HRP conjugated anti-human IgG (7 citations) Anti-human IgG Fc HRP (4 citations)

4 protocols using hrp conjugated anti human igg fc

1

SARS-CoV-2 RBD-ACE2 Inhibition Assay

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The hACE2-RBD inhibition assay modified a previously existing protocol 47 , 54 (link). Briefly, high-binding flat-bottom 96-well plates (Corning) were coated with 100 ng per well recombinant human ACE2 (hACE2) (Sigma-Aldrich) in PBS, incubated overnight at 4°C, washed three times with 0.05% Tween 20 PBS, and blocked with 1% BSA PBS for 1 hour at room temperature. Each serum sample was diluted 1:80, pre-incubated with 3 ng of RBD-Fc in 1% BSA PBS for 1 hour at room temperature, and then transferred to the hACE2-coated plate. RBD-Fc without pre-incubation with serum samples was added as a positive control, and 1% BSA PBS without serum pre-incubation was added as a negative control. Plates were then washed three times and incubated with HRP-conjugated anti-human IgG Fc (Southern Biotech) for 1 hour at room temperature. Plates were washed five times and developed with tetramethylbenzidine (BD OptEIA Substrate Solution, BD Biosciences) for 5 min, then stopped with 2 N H2SO4. The optical density was read at 450 nm with a SpectraMax iD3 microplate reader (Molecular Devices). Percentage inhibition of RBD binding to hACE2 was calculated with the following formula: Inhibition (%) = [1 – (Sample OD value – Negative Control OD value)/(Positive Control OD value – Negative Control OD value)] x 100.
O’Meara T.R., Nanishi E., McGrath M.E., Barman S., Dong D., Dillen C., Menon M., Seo H.S., Dhe-Paganon S., Ernst R.K., Levy O., Frieman M.B, & Dowling D.J. (2022). Reduced SARS-CoV-2 mRNA vaccine immunogenicity and protection in mice with diet-induced obesity and insulin resistance. bioRxiv.
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2

SARS-CoV-2 RBD-ACE2 Inhibition Assay

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The hACE2-RBD inhibition assay employed a modification of a previously published protocol(84 (link)). Briefly, high-binding flat-bottom 96-well plates (Corning) were coated with 100 ng per well recombinant human ACE2 (hACE2) (Sigma-Aldrich) in PBS, incubated overnight at 4°C, washed three times with 0.05% Tween 20 PBS, and blocked with 1% BSA PBS for 1 hour at room temperature. Each serum sample was diluted 1:160, pre-incubated with 3 ng of RBD-Fc in 1% BSA PBS for 1 hour at room temperature, and then transferred to the hACE2-coated plate. RBD-Fc without pre-incubation with serum samples was added as a positive control, and 1% BSA PBS without serum pre-incubation was added as a negative control. Plates were then washed three times and incubated with HRP-conjugated anti-human IgG Fc (Southern Biotech) for 1 hour at room temperature. Plates were washed five times and developed with tetramethylbenzidine (BD OptEIA Substrate Solution, BD Biosciences) for 5 min, then stopped with 2 N H2SO4. The optical density was read at 450 nm with SpectraMax iD3 microplate reader (Molecular Devices). Percentage inhibition of RBD binding to hACE2 was calculated with the following formula: Inhibition (%) = [1 – (Sample OD value – Negative Control OD value)/(Positive Control OD value – Negative Control OD value)] x 100.
Nanishi E., Borriello F., O’Meara T.R., McGrath M.E., Saito Y., Haupt R.E., Seo H.S., van Haren S.D., Cavazzoni C.B., Brook B., Barman S., Chen J., Diray-Arce J., Doss-Gollin S., De Leon M., Prevost-Reilly A., Chew K., Menon M., Song K., Xu A.Z., Caradonna T.M., Feldman J., Hauser B.M., Schmidt A.G., Sherman A.C., Baden L.R., Ernst R.K., Dillen C., Weston S.M., Johnson R.M., Hammond H.L., Mayer R., Burke A., Bottazzi M.E., Hotez P.J., Strych U., Chang A., Yu J., Sage P.T., Barouch D.H., Dhe-Paganon S., Zanoni I., Ozonoff A., Frieman M.B., Levy O, & Dowling D.J. (2022). An aluminum hydroxide:CpG adjuvant enhances protection elicited by a SARS-CoV-2 receptor-binding domain vaccine in aged mice. Science translational medicine, 14(629), eabj5305.
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3

hACE2-RBD Inhibition Assay Protocol

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The hACE2-RBD inhibition assay used a modification of a previously published protocol29 (link). Briefly, high-binding flat-bottom 96- well plates (Corning) were coated with recombinant hACE2 (100 ng per well) (Sigma-Aldrich) in PBS, incubated overnight at 4 °C, washed three times with 0.05% Tween 20 PBS, and blocked with 1% BSA PBS for 1 h at RT. Serum samples were diluted 1:160, preincubated with 3 ng of wildtype RBD-Fc in 1% BSA PBS for 1 h at RT, and then transferred to the hACE2-coated plate. RBD-Fc without preincubation with serum samples was added as a positive control, and 1% BSA PBS without serum preincubation was added as a negative control. Plates were then washed three times and incubated with HRP-conjugated anti-human IgG Fc (SouthernBiotech) for 1 h at RT. Plates were washed five times and developed with tetramethylbenzidine (BD OptEIA Substrate Solution, BD Biosciences) for 5 min and then stopped with 2 N H2SO4. The OD was read at 450 nm with a SpectraMax iD3 microplate reader (Molecular Devices). Percentage inhibition of RBD binding to hACE2 was calculated as: Inhibition (%) = [1 − (Sample OD value − Negative Control OD value)/(Positive Control OD value − Negative Control OD value)] × 100.
Nanishi E., McGrath M.E., O’Meara T.R., Barman S., Yu J., Wan H., Dillen C.A., Menon M., Seo H.S., Song K., Xu A.Z., Sebastian L., Brook B., Bosco A.N., Borriello F., Ernst R.K., Barouch D.H., Dhe-Paganon S., Levy O., Frieman M.B, & Dowling D.J. (2022). mRNA booster vaccination protects aged mice against the SARS-CoV-2 Omicron variant. Communications Biology, 5, 790.
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4

ACE2-RBD Inhibition Assay

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We performed sVNT to measure the degree of hACE2/RBD inhibition by immune sera. High-binding flat-bottom 96-well plates (Corning, NY) were coated with 100 ng/well recombinant human ACE2 (hACE2) (Sigma-Aldrich) in PBS, incubated overnight at 4 °C, washed three times with PBS-T, and blocked with 1% BSA PBS for 1 h at RT. Each serum sample was diluted at 1:160, pre-incubated with 3 ng of RBD-Fc in 1% BSA PBS for 1 h at RT, and then transferred to the hACE2-coated plate. RBD-Fc without pre-incubation with serum samples was added as a positive control, and 1% BSA PBS without serum pre-incubation was added as a negative control. Plates were then washed three times and incubated with HRP-conjugated anti-human IgG Fc (Southern Biotech) for 1 h at RT. Plates were washed five times and developed with tetramethylbenzidine (BD OptEIA Substrate Solution, BD Biosciences) for 5 min, then stopped with 2 N H2SO4. The optical density was read at 450 nm with a SpectraMax iD3 microplate reader (Molecular Devices). Percentage inhibition of RBD binding to hACE2 was calculated with the following formula: Inhibition (%) = [1 – (Sample OD value – Negative Control OD value)/(Positive Control OD value – Negative Control OD value)] × 100.
Nanishi E., Borriello F., Seo H.S., O’Meara T.R., McGrath M.E., Saito Y., Chen J., Diray-Arce J., Song K., Xu A.Z., Barman S., Menon M., Dong D., Caradonna T.M., Feldman J., Hauser B.M., Schmidt A.G., Baden L.R., Ernst R.K., Dillen C., Yu J., Chang A., Hilgers L., Platenburg P.P., Dhe-Paganon S., Barouch D.H., Ozonoff A., Zanoni I., Frieman M.B., Dowling D.J, & Levy O. (2023). Carbohydrate fatty acid monosulphate: oil-in-water adjuvant enhances SARS-CoV-2 RBD nanoparticle-induced immunogenicity and protection in mice. NPJ Vaccines, 8, 18.
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