In brief, multiscreen 96-well filter plates (Milipore, Burlington, MA) were coated with 5 μg/mL of purified mouse anti-human IgM antibody (Sigma-Aldrich) overnight, washed and blocked with 5% bovine serum albumin (BSA) (Sigma Aldrich) for 2 hours. During this time, B cells were harvested and washed 3x with RPMI 1640, enumerated, and resuspended in supplemented RPMI with 10% bovine calf serum (ThermoFisher) and incubated overnight at 37°. The following day plates were incubated with biotin-conjugated mouse anti-IgM (Sigma Aldrich) and streptavidin-horseradish peroxidase (Sigma Aldrich) for 1 h at 37°C. All incubations were preceded by 3 washes with phosphate buffered saline plus 0.1% Tween-20 (Sigma Aldrich) and 3 washes with nanopure water. IgM positive spots were developed with aminoethyl carbazole staining kit (Sigma Aldrich). Spots were quantified using Immunospot Software (Cellular Technology, Shaker Heights, OH) and normalized to the number of viable cells plated in each well.
Biotin conjugated mouse anti igm
Manufactured by Merck Group
Biotin-conjugated mouse anti-IgM is a laboratory reagent used for the detection and quantification of immunoglobulin M (IgM) in biological samples. It consists of mouse-derived antibodies that specifically bind to IgM molecules, with biotin molecules conjugated to the antibodies. The biotin label allows for easy detection and signal amplification when the reagent is used in various immunoassay techniques.
Automatically generated - may contain errors
Lab products found in correlation
Bovine calf serum (bcs), by Thermo Fisher Scientific (2 mentions)
Aminoethyl carbazole staining kit, by Merck Group (2 mentions)
Streptavidin horseradish peroxidase, by Merck Group (2 mentions)
Mouse anti human igm antibody, by Merck Group (2 mentions)
Multiscreen 96 well filter plate, by Merck Group (2 mentions)
Immunospot software, by Cellular Technology (2 mentions)
Bovine serum albumin (bsa), by Merck Group (2 mentions)
Tween 20, by Merck Group (2 mentions)
Phosphate buffered saline (pbs), by Merck Group (1 mentions)
2 protocols using biotin conjugated mouse anti igm
1
Quantification of IgM-secreting B Cells
2
Quantifying IgM-Secreting B Cells
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In brief, multiscreen 96-well filter plates (Milipore, Burlington, MA) were coated with 5 μg/mL of purified mouse anti-human IgM antibody (Sigma-Aldrich) overnight, washed and blocked with 5% bovine serum albumin (BSA) (Sigma Aldrich) for 2 hours. During this time, B cells were harvested and washed 3x with RPMI 1640, enumerated, and resuspended in supplemented RPMI with 10% bovine calf serum (ThermoFisher) and incubated overnight at 37°C. The following day, filter plates were incubated with biotin-conjugated mouse anti-IgM (Sigma Aldrich) and streptavidin-horseradish peroxidase (Sigma Aldrich) for 1 h at 37°C. All incubations were preceded by 3 washes with phosphate buffered saline plus 0.1% Tween-20 (Sigma Aldrich) and 3 washes with nanopure water. IgM positive spots were developed with aminoethyl carbazole staining kit (Sigma Aldrich). Spots were quantified using Immunospot Software (Cellular Technology, Shaker Heights, OH) and normalized to the number of viable cells plated in each well.
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