Mt91 145

Manufactured by Mabtech

The MT91/145 is a lab equipment product from Mabtech. It functions as a pre-coated 96-well ELISPOT plate for the detection and enumeration of cytokine-secreting cells.

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Lab products found in correlation

Mt78 145, by Mabtech (4 mentions) Nbt bcip, by Thermo Fisher Scientific (1 mentions) Streptavidin alp, by Mabtech (1 mentions) Phosphate buffered saline (pbs), by Thermo Fisher Scientific (1 mentions) Elispot plate, by Merck Group (1 mentions) Streptavidin hrp, by BD (1 mentions) Tmb substrate, by Mabtech (1 mentions) Streptavidin alkaline phosphatase, by Mabtech (1 mentions) Bcip nbt, by Mabtech (1 mentions) Rhil 2, by Mabtech (1 mentions) Maips4510, by Merck Group (1 mentions) Immunospot reader, by Cellular Technology (1 mentions)

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Anti-human IgG MT91/145 (2 citations)

6 protocols using mt91 145

ELISPOT Quantification of IgG and IgA

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Briefly, ELISPOT plates (MSIPSW10, Millipore Sigma) were activated with 70% EtOH before being washed 4x with sterile dH₂O. Plates were coated overnight with purified anti-human IgG (MT91/145, Mabtech), anti-monkey IgA (polyclonal, Rockland), or anti-human IgA (MT57, Mabtech). Cells to be assayed were plated in duplicate and serially diluted down the ELISPOT plate before overnight incubation at 37 °C in a 5% CO₂ incubator. The following day, cells were lysed by incubation with dH₂O before being washed 4× with PBS-T. Secreted IgG or IgA were detected using biotinylated anti-human IgG (MT78/145, Mabtech) or biotinylated anti-human IgA (MT57, Mabtech) followed by incubation with streptavidin-ALP (Mabtech). ELISPOT plates were developed with NBT/BCIP (ThermoFisher Scientific) before being quenched with dH₂O. All washes between incubation steps were performed using PBS-T. Plates were read and quantified on an Immunospot S6 (Cellular Technology Limited (CTL)) using CTL Immunocapture (v7.0.16.1, CTL) and CTL Biospot (v7.0.3.4, CTL). Manual inspection of automated counting results was performed as quality control during analysis.

Staupe R.P., Lodge K.E., Thambi N., Toole D., Tamburino A.M., Chang D., Howell B.J., Hazuda D.J., Vora K.A, & Sullivan N.L. (2022). Single cell multi-omic reference atlases of non-human primate immune tissues reveals CD102 as a biomarker for long-lived plasma cells. Communications Biology, 5, 1399.

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Human IgG and IgA ELISPOT Assay

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Human total-IgG and total-IgA ELISPOTs were performed as described above for rhesus macaque ELISPOTs with the following exception. Plates were coated overnight with purified anti-human IgG (MT91/145, Mabtech) or anti-human IgA (MT57, Mabtech). All other detection reagents, wash steps, and data acquisition and analysis were performed as for rhesus macaque samples.

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Quantifying Dengue Virus-Specific IgG Responses

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Millipore ELISpot plates (cat. # MAIPSWU10) were pre-wetted with 70% ethanol and washed 5 times with distilled water. Wells were coated with 1×10⁶ PFU of unlabeled DENV. To detect total IgG, wells were coated with 100μl of anti-human IgG (15μg/ml) (Mabtech, MT91/145). Plates were stored at 4°C overnight, washed with PBS (Gibco) and blocked with 100μl RPMI 1640/10% FBS for 30 minutes at room temperature (RT). The cells were washed to remove any bound IgG, counted (AF-DENV⁺ and total B cells) and 100 μl of cells were added to each well. Cell concentrations varied depending on the cell recovery. The plate was incubated at 37°C overnight. After washing, 100 μl of biotinylated monoclonal antibody directed against human IgG (1 μg/ml in PBS/0.5 % FCS; MT78/145, Mabtech) was added. Plates were incubated for 2 h at RT, washed and Streptavidin HRP (100μl of 1:1000 dilution in PBS/0.5% FBS) (BD Pharmingen) was added. Plates were incubated at RT for 1 h, developed with TMB substrate (100μl/well) (Mabtech) and analyzed using an ELISpot plate reader (CTL Immunospot reader).

Woda M, & Mathew A. (2014). Fluorescently labeled dengue viruses as probes to identify antigen-specific memory B cells by multiparametric flow cytometry. Journal of immunological methods, 416, 167-177.

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IgG Monoclonal Antibody Detection

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Anti-human IgG monocloning antibodies (MT91/145, Mabtech) were diluted with PBS (PH7.4) and coated onto a 96-well polyvinylidene fluoride plate (Millipore) treated with 70% ethanol for 2 min at a concentration of 15 µg/mL (100 µL/well) overnight at 4°C. The plates were washed with PBS for five times (300 µL/well) and blocked using 10% bovine serum (200 µL/well) at room temperature for 1 hour. MB cells were pre-stimulated with a mixture of R848 at 1 µg/mL and rhIL-2 at 10 ng/mL (Mabtech) for 24 hours. Activated MB cells and CD8⁺ T cells were inoculated into the plates at a ratio of 1:4 and cultured at 37°C and 5% CO₂ for 48 hours. After washing the plate five times with PBS (300 µL/well), 100 µL (1 µg/mL) of biotin-anti human IgG mAb (MT78/145, Mabtech) was added to the plates and incubated at RT for 2 hours. After another round of washing, 100 µL streptavidin-Alkaline phosphatase (1:1000, Mabtech) was added and incubated for 1 hour at RT. 100 µL substrate solution (BCIP/NBT, Mabtech) were added and developed until the distinct spots emerge. The plates were analysed using a CTL reader (ImmunoSpot, Cleveland, Ohio, USA).

Wang K., Zhao J., Feng X., He S., Li J., Sun F., Xu Z., Yang H., Ye J., Cao L, & Ye S. (2024). PD-1/PD-L1 governed cross-talk of exhausted CD8+ T and memory B cells in systemic lupus erythematosus. RMD Open, 10(1), e003503.

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ELISpot Assay for Antibody-Secreting Cells

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ELISpot was performed according to manufactures instructions (Human IgG ELISpotBASIC HRP, Mabtech). Briefly, PVDF ELISpot plates (MAIPS4510, Millipore) were Ethanol treated and coated with capture antibody (MT91/145, Mabtech) at a concentration of 15 µg/ml in PBS and incubated overnight at 4 °C. Total PBMCs or sorted CD11c^hi B cells (CD19⁺CD11c^hi) or plasma cells (CD19⁺CD27⁺⁺CD38⁺⁺) from SLE donors (n = 4) were added to wells at indicated concentrations in complete media and incubated for 16 h at 37 °C. The plates were then washed and incubated with detection mAb (MT78/145, Mabtech) at a concentration of 1 µg/ml for 2 h at RT, followed by streptavidin-horseradish peroxidase conjugate for 1 h at RT and developed with precipitating TMB substrate (Mabtech) for 15 min. The images were captured using an Immunospot reader (Cellular Technology Limited).

Wang S., Wang J., Kumar V., Karnell J.L., Naiman B., Gross P.S., Rahman S., Zerrouki K., Hanna R., Morehouse C., Holoweckyj N., Liu H., Manna Z., Goldbach-Mansky R., Hasni S., Siegel R., Sanjuan M., Streicher K., Cancro M.P., Kolbeck R, & Ettinger R. (2018). IL-21 drives expansion and plasma cell differentiation of autoreactive CD11chiT-bet+ B cells in SLE. Nature Communications, 9, 1758.

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Quantifying SARS-CoV-2 RBD-Specific Memory B Cells

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SARS-CoV-2 RBD-specific memory B cell numbers were counted using ELISpot. MultiScreenHTS IP Filter Plate, 0·45 µm plates (Merck Millipore) were coated overnight at 4°C with purified anti-human-IgG (MT91/145, Mabtech) or purified anti-human-IgA prepared at 15 μg/mL in PBS. Plates were washed and blocked for 30 min at room temperature with RPMI + 10% FBS. 1 x 10⁶ PBMCs were resuspended in 1 mL RPMI + 10% FBS + 1 μg/mL R848 + 10 ng/mL IL-2, and incubated at 37°C, 5% CO₂ for 5 days to differentiate memory B cells into antibody-secreting cells. After incubation, cells were counted, and 100,000 or 400,000 live cells were taken for ELISpot plating to determine RBD-specific memory B cell numbers. Total IgG secreting cells were determined by plating 1500 or 3000 live cells. Cells were incubated for 18-22 h at 37°C, 5% CO₂ in the ELISpot plate before detection. A combination of RBD-WASP/anti-WASP-ALP or anti-IgG-biotinylated/streptavidin-ALP (Mabtech) were used to detect RBD-specific or total IgG secreting cells respectively.

Goh Y.S., Rouers A., Fong S.W., Zhuo N.Z., Hor P.X., Loh C.Y., Huang Y., Neo V.K., Kam I.K., Wang B., Ngoh E.Z., Salleh S.N., Lee R.T., Pada S., Sun L.J., Ong D.L., Somani J., Lee E.S., Maurer-Stroh S., Wang C.I., Leo Y., Ren E.C., Lye D.C., Young B.E., Ng L.F, & Renia L. (2022). Waning of specific antibodies against Delta and Omicron variants five months after a third dose of BNT162b2 SARS-CoV-2 vaccine in elderly individuals. Frontiers in Immunology, 13, 1031852.

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