A highly sensitive single-cell assay detects T-helper cell responses missed by conventional interleukin-2-based methods.

One of the most widely used methods for the study of T-helper (T(h)) cell activity is the interleukin 2 (IL-2) assay. Typically, this assay is a two-step process involving (a) the activation of T(h) cells in vitro and (b) the testing of IL-2-dependent indicator cells for growth in the presence of T(h) cell supernatants. The assay has served to quantify and characterize T(h) responses to a variety of unique pathogens and immunization regimens. A one-step, single-cell assay is also available for the testing of mouse T(h) cell hybridomas. In this assay, cells fused with the BWZ.36 parent line are scored for positive responses based on their expression of a beta-galactosidase gene linked to an IL-2 enhancer element. The experiments described in this report were designed to examine the conventional and single-cell assays in a side-by-side comparison. Results revealed a striking difference between the two assays. The single-cell assay proved to be extremely sensitive and identified T(h) responses that were altogether missed by the conventional test. Based on these results, we suggest that (i) the conventional and single-cell assays should not be used interchangeably, and (ii) negative results in the conventional IL-2 assay should be considered preliminary until the more sensitive, single-cell assay can be performed.