Anti-Qβ development was assessed using an enzyme-linked immunosorbent assay. 96-well plates (Corning, product 3590) were coated with vidutolimod. Serum samples and monoclonal anti-Qβ (clone Qb5) standard were incubated overnight at 4°C, and human antibodies were detected using goat anti-human Ig-HRP (SouthernBiotech, catalog no. 2010-05). Plates were developed with 3,3′,5,5′-tetramethylbenzidine (Sigma-Aldrich, catalog no. T8665), and anti-Qβ concentrations were calculated from the standard curve generated with monoclonal anti-Qβ antibody (clone Qb5l ref. 14 (link)).
Goat anti human ig hrp
Manufactured by Southern Biotech
Goat anti-human Ig-HRP is a laboratory reagent used for the detection and quantification of human immunoglobulins (Ig) in various immunoassay applications. It is a conjugate of goat-derived anti-human Ig antibodies and the enzyme horseradish peroxidase (HRP).
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Lab products found in correlation
Immulon 2hb microtiter plates, by Thermo Fisher Scientific (1 mentions)
Spectramax m2 plate reader, by Molecular Devices (1 mentions)
Calf thymus dna, by Merck Group (1 mentions)
Fraction 5, by Thermo Fisher Scientific (1 mentions)
Spectramax i3x plate reader, by Molecular Devices (1 mentions)
Maxisorp microtiter plate, by Thermo Fisher Scientific (1 mentions)
96 well plate, by Corning (1 mentions)
5 protocols using goat anti human ig hrp
1
Quantification of Anti-Qβ Antibodies
2
Quantitative SARS-CoV RBD IgG ELISA
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Immulon 2HB microtiter plates (Thermo Scientific) were coated with 50 ng/well of SARS-CoV-1 or SARS-CoV-2 RBD at 4 °C overnight. Plates were washed 4X with phosphate-buffered saline (PBS) with 0.02% Tween-20 (wash buffer). Plates were blocked with 250 μL of 10% non-fat milk and 0.02% Tween-20 in PBS (blocking buffer) for 1 h at 37 °C. After washing 4× with wash buffer, purified IgG was added at an initial concentration of 20 μg/mL and diluted in tenfold serial dilutions in blocking buffer and incubated for 1 h at 37 °C. Plates were washed 4× in wash buffer and the secondary antibody Goat anti-Human Ig-HRP (Southern Biotech, catalog number 2010-05) was added at a 1 : 3000 dilution and incubated at 37 °C for 1 h. After a final 4× wash, 50 μL of SureBlue Reserve TMB Peroxidase Substrate (SeraCare, catalog number 5120-0081) was added and incubated for 4 min followed by addition of 100 μL of 1 N H2SO4 to stop the reaction. The optical density at 450 nm was measured using a SpectraMax M2 plate reader (Molecular Devices). All wash steps were performed using a BioTek 405 Select Microplate Washer.
3
Measuring Anti-dsDNA and Anti-Sm/RNP Autoantibodies in SLE
4
Streptavidin-Coated ELISA for Peptide-mAb Binding
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MaxiSorp microtiter plates (Thermo Scientific Cat#464718) were coated with 300 ng/well of streptavidin (New England Biolabs Catalog #: N7021S) overnight at room temperature. Plates were washed 4X with PBS with 0.02% Tween-20 (wash buffer), then incubated with 60 μL/well of 3% BSA and 0.02% Tween-20 in PBS (blocking buffer) for 1 hr at 37 °C. After washing 4X with wash buffer, 380 ng/well of biotinylated peptides diluted in blocking buffer were incubated for 1 hr at 37 °C. Plates were washed 4X in wash buffer and then mAbs were serially diluted in blocking buffer, added to the plate and incubated for 1 hr at 37 °C. Plates were washed 4X in wash buffer and the secondary antibody Goat anti-Human Ig-HRP (Southern Biotech, Cat# 2010-05), was added and incubated at 37 °C for 1 hr. Plates were washed 4X wash buffer, and then 30 μL/well of SureBlue Reserve TMB Peroxidase Substrate (Seracare KPL, Cat# 5120-0080) was added and incubated for 3 min followed by addition of 30 μL of 1 N H2SO4 to stop the reaction. The optical density at 450 nm was measured using a SpectraMax i3x plate reader (Molecular Devices). All wash steps were performed using a BioTek 405/TSMicroplate Washer.
5
Enzyme-Linked Immunosorbent Assay for Anti-Qβ
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