The ELISpot assay can be used successfully to quantify the number of immunoglobulin-secreting cells. It was first developed in 1983 as a highly sensitive method for the identification of antibody secretion at the single-cell level.
The frequency of immunoglobulin-secreting B cells can be determined with the ELISpot assay. B cells secreting antigen-specific immunoglobulin, or indeed immunoglobulin of all specificities, can be quantified using this assay. The number of antigen-specific B cells is most often compared to the total number of secreting B cells. B cells that secrete antigen-specific antibodies can be detected in the circulation from 6 to 9 days after antigen exposure (e.g. vaccination). Memory B cells may require polyclonal stimulation before detectable amounts of antibodies are found.
The antigen-specific assay may be performed in two different ways: either (A) the antigen or (B) the specific anti-immunoglobulin antibodies are bound to the plate. Then, antigen-specific antibodies are detected by (A) biotinylated anti-IgG or (B) biotinylated antigen. The latter approach produces spots that are more distinct and it also reduces the amount of antigen required. Simultaneous detection of more than one isotype is possible in the B cell FluoroSpot assay, in which fluorescent rather than enzymatic signals are used. See FluoroSpot for various celltypes.