Recombinant SARS-CoV-2 Spike protein was coated onto high-binding 96-well plates (Corning, 3690) at 2 μg/mL, overnight at 4 °C. Wash steps were done using 1xPBS containing 0.05% Tween-20 (Sigma, P1379) and all incubations were done at 25 °C. Plates were blocked with 3% nonfat dry milk (BioRad, P1379) in 1xPBS for 1 hour. Serum from COVID-19 convalescent patients was serially diluted in 1XPBS, containing 1% nonfat dry milk and 0.1% Tween-20, then added to plates and incubated for 2 hours. Following plate washing, serum reactivity to the Spike protein was measured using an HRP-conjugated goat anti-human IgG (H+L) (Invitrogen, 31410) at a 1:3000 dilution for 1 hour. After a final wash, Ultra-TMB substrate (ThermoFisher, 34029) was added to the plates and incubated for 5 minutes, followed by quenching of the reaction by addition of an equal volume of 0.5M H2SO4. Absorption was measured at OD450 using a Synergy4 plate reader (Biotek), and data was analyzed using GraphPad Prism7.0 to calculate IC50 values.
Hrp conjugated goat anti human igg h l
Manufactured by Thermo Fisher Scientific
HRP-conjugated goat anti-human IgG (H+L) is a secondary antibody used to detect and quantify human immunoglobulin G (IgG) in various immunoassays. The antibody is conjugated with horseradish peroxidase (HRP), which serves as a reporter enzyme for colorimetric or chemiluminescent detection.
Automatically generated - may contain errors
2 protocols using hrp conjugated goat anti human igg h l
1
SARS-CoV-2 Spike Protein Antibody Assay
2
SARS-CoV-2 Spike Protein Antibody Assay
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Recombinant
SARS-CoV-2 Spike
protein was coated onto high-binding 96-well plates (Corning, 3690)
at 2 μg/mL, overnight at 4 °C. Wash steps were done using
1× PBS containing 0.05% Tween-20 (Sigma, P1379), and all incubations
were done at 25 °C. Plates were blocked with 3% nonfat dry milk
(Bio-Rad, P1379) in 1× PBS for 1 h. Serum from COVID-19 convalescent
patients was serially diluted in 1× PBS, containing 1% nonfat
dry milk and 0.1% Tween-20, and then added to plates and incubated
for 2 h. Following plate washing, serum reactivity to the Spike protein
was measured using an HRP-conjugated goat anti-human IgG (H + L) (Invitrogen,
31410) at a 1:3000 dilution for 1 h. After a final wash, the Ultra-TMB
substrate (Thermo Fisher, 34029) was added to the plates and incubated
for 5 min followed by quenching of the reaction by addition of an
equal volume of 0.5 M H2SO4. Absorption was
measured at OD450 using a Synergy4 plate reader (Biotek),
and data was analyzed using GraphPad Prism7.0 to calculate IC50 values.
SARS-CoV-2 Spike
protein was coated onto high-binding 96-well plates (Corning, 3690)
at 2 μg/mL, overnight at 4 °C. Wash steps were done using
1× PBS containing 0.05% Tween-20 (Sigma, P1379), and all incubations
were done at 25 °C. Plates were blocked with 3% nonfat dry milk
(Bio-Rad, P1379) in 1× PBS for 1 h. Serum from COVID-19 convalescent
patients was serially diluted in 1× PBS, containing 1% nonfat
dry milk and 0.1% Tween-20, and then added to plates and incubated
for 2 h. Following plate washing, serum reactivity to the Spike protein
was measured using an HRP-conjugated goat anti-human IgG (H + L) (Invitrogen,
31410) at a 1:3000 dilution for 1 h. After a final wash, the Ultra-TMB
substrate (Thermo Fisher, 34029) was added to the plates and incubated
for 5 min followed by quenching of the reaction by addition of an
equal volume of 0.5 M H2SO4. Absorption was
measured at OD450 using a Synergy4 plate reader (Biotek),
and data was analyzed using GraphPad Prism7.0 to calculate IC50 values.
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