IL-4 analysis in ELISpot

IL-4 is a hallmark cytokine for the Th2-type T-cell responses which are prominent in many types of allergies. However, it is also a cytokine that has been persistently difficult to detect partly due to the often low numbers of specifically responding cells, the relatively low amounts of IL-4 produced per cell and also because of it being consumed by IL-4 receptor bearing cells. Therefore, IL-4 has often escaped detection by ELISA as well as by intracellular staining in combination with flow cytometry. The ELISpot here provides an ideal method with a sensitivity that may be as low as one or a few positive cells in 100,000. Also, as the cytokine is detected at the site of the producing cell, it is virtually unaffected by the consumption by other cells.

In the figure, PBMC from subjects with or without contact allergy to nickel were tested for in vitro reactivity to nickel. The subjects displayed +++ (n=10), ++ (n=11), + (n=10) or no (controls; n=10) patch test reactivity to nickel. PBMC were incubated with or without 50 µM NiCl2 for 40 h and IL-4 responses were assessed in ELISpot and in ELISA using cell culture supernatants. Detectable responses to nickel, above cut off levels, were seen in 23 of the 31 allergic individuals when tested with ELISpot whereas only 4 were defined as positive in the ELISA.