Alkaline phosphatase conjugated anti human igg clone mt78

Manufactured by Mabtech

Alkaline phosphatase-conjugated anti-human IgG (clone MT78) is a laboratory reagent. It is an antibody conjugated to the enzyme alkaline phosphatase.

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Lab products found in correlation

P nitrophenyl phosphate substrate, by Thermo Fisher Scientific (2 mentions) Nunc maxisorp 96 well plate, by Thermo Fisher Scientific (2 mentions) Nunc maxisorp 96 well elisa plate, by Thermo Fisher Scientific (2 mentions)

4 protocols using alkaline phosphatase conjugated anti human igg clone mt78

ELISA-based Quantification of Antigen-specific Antibodies

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Concentrations of Ag-specific serum Abs and MPAbs were measured by ELISA as previously described (22 (link)). Briefly, Nunc MaxiSorp 96-well plates (Thermo Fisher, Waltham, MA) were coated overnight with optimized concentrations of antigens. Serially diluted samples were added to blocked plates and incubated for 2 h at room temperature. Alkaline phosphatase-conjugated anti-human IgG (clone MT78; Mabtech, Stockholm, Sweden) and p-nitrophenyl phosphate substrate (Thermo Fisher) were subsequently added to detect bound antigen-specific Abs. Absorbance was read at 405 nm after color development. A weight-based concentration method was used to quantify antigen-specific Ab levels in test samples as described previously (22 (link), 35 (link)). Sera from healthy donors and convalescent subjects with high titers for test antigens were used to establish human serum standards. The cutoff for assay positivity was set at approximately 2× the mean optical density (OD) value for negative wells.

Nguyen-Contant P., Embong A.K., Kanagaiah P., Chaves F.A., Yang H., Branche A.R., Topham D.J, & Sangster M.Y. (2020). S Protein-Reactive IgG and Memory B Cell Production after Human SARS-CoV-2 Infection Includes Broad Reactivity to the S2 Subunit. mBio, 11(5), e01991-20.

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ELISA for SARS-CoV-2 Antibody Quantification

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Serum IgG titers specific for SARS-CoV-2 proteins and selected non-coronavirus proteins were determined by ELISA as described previously.^{23 (link)} In brief, NUNC MaxiSorp 96-well ELISA plates (Invitrogen, Carlsbad, CA) were coated with optimized concentrations of coating reagents at least one day prior to the assay. After blocking plates with 3% BSA/PBS for 1 h, serial 3-fold dilutions of serum samples in ELISA diluent (0.5% BSA/0.05% Tween-20/PBS) were added and incubated for 2 h. Antigen-specific IgG was detected by addition of alkaline phosphatase-conjugated anti-human IgG (clone MT78; Mabtech, Cincinnati, OH), followed by p-nitrophenyl phosphate substrate. Well absorbance was read at 405 nm after color development. Human serum standards were used to assign weight-based concentrations of antigen-specific IgG as previously described, with the limit of assay sensitivity set at 0.5 μg/mL for all antigens.^{23 (link),24 (link)}

Steiner D.J., Cognetti J.S., Luta E.P., Klose A.M., Bucukovski J., Bryan M.R., Schmuke J.J., Nguyen-Contant P., Sangster M.Y., Topham D.J, & Miller B.L. (2020). Array-based analysis of SARS-CoV-2, other coronaviruses, and influenza antibodies in convalescent COVID-19 patients. Biosensors & Bioelectronics, 169, 112643.

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SARS-CoV-2 IgG Antibody Quantification

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Serum IgG titers specific for SARS-CoV-2 proteins and selected non-coronavirus proteins were determined by ELISA as described previously (Tesini et al., 2019 ). In brief, NUNC MaxiSorp 96-well ELISA plates (Invitrogen, Carlsbad, CA) were coated with optimized concentrations of coating reagents at least one day prior to the assay. After blocking plates with 3% BSA/PBS for 1 h, serial 3-fold dilutions of serum samples in ELISA diluent (0.5% BSA/0.05% Tween-20/PBS) were added and incubated for 2 h. Antigen-specific IgG was detected by addition of alkaline phosphatase-conjugated anti-human IgG (clone MT78; Mabtech, Cincinnati, OH), followed by p-nitrophenyl phosphate substrate. Well absorbance was read at 405 nm after color development. Human serum standards were used to assign weight-based concentrations of antigen-specific IgG as previously described, with the limit of assay sensitivity set at 0.5 μg/mL for all antigens. (Tesini et al., 2019 )., (Quataert et al., 1995 (link))

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Milk-borne Antibodies against SARS-CoV-2 and Seasonal Coronaviruses

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Concentrations of milk-borne IgA and IgG reactive to the SARS-CoV-2 spike (both S2 subunit and RBD) and nucleocapsid (N) proteins and spike proteins of seasonal coronaviruses 229E and OC43 were measured in milk samples by ELISA as previously described.^{31 (link)} Briefly, Nunc MaxiSorp 96-well plates (Thermo Fisher, Waltham, MA) were coated with optimized concentrations of antigens (1–5 μg/mL) overnight at 4°C. Coated plates were blocked for 1 h before the addition of serial 2-fold dilutions of samples. After a 2 h incubation at room temperature, plates were washed and bound IgG and IgA were detected with alkaline phosphatase-conjugated anti-human IgG (clone MT78; Mabtech, Stockholm, Sweden) and anti-human IgA. Bound antigen-specific antibodies were detected by adding p-nitrophenyl phosphate substrate (Thermo Fisher). Absorbance was read at 405 nm after color development. A weight-based concentration method was used to assign antigen-specific antibody titers in test samples.^{31 (link),32 (link)}

Pace R.M., Williams J.E., Järvinen K.M., Belfort M.B., Pace C.D., Lackey K.A., Gogel A.C., Nguyen-Contant P., Kanagaiah P., Fitzgerald T., Ferri R., Young B., Rosen-Carole C., Diaz N., Meehan C.L., Caffe B., Sangster M.Y., Topham D., McGuire M.A., Seppo A, & McGuire M.K. (2020). COVID-19 and human milk: SARS-CoV-2, antibodies, and neutralizing capacity. medRxiv.

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