Intended Use: This 2.5 hour ELISpot kit is developed to detect and visualize of single cells secreting human IL-4. Principle: Add stimulated cells or cells and stimulant to the wells and incubate at 37°C in CO2 incubator for a specified period. Secreted human IL-4 binds to antibody coated microtiter plate. microtiter plate coated with human IL-4 Capture Antibody. Cells and unbounded proteins are washed away. Streptavidin-AP or Streptavidin-HRP is added and binds to the biotinylated detection antibody. Unbounded proteins are washed away. Biotinylated detection antibody is added and binds to the secreted human IL-4. Substrate Solution is added. A colored precipitate forms and appears as spots at the sites of human IL-4 secreting location. Each individual spot representing an individual human IL-4 secreting cell. Background: Interleukin 4 (IL-4) was initially characterized as a B cell stimulatory factor (BSF-1) (1) and B-cell differentiation factor (BCDFγ) (2). It was subsequent