Correlation of human CD56+ cell cytotoxicity and IFN-gamma production.

The use of an IFN-gamma ELISPOT assay to evaluate cellular immune responses has gained increasing popularity, especially as a surrogate measure for cytotoxic T lymphocyte (CTL) responses. We have compared the IFN-gamma ELISPOT assay and the traditional(51)Cr release assay for detection of human natural killer (NK) cell activity. The cell populations used for evaluation of these assays included freshly isolated and IL-2-activated peripheral blood mononuclear cells (PBMC). CD56-positive cells were demonstrated to be the primary source of the IFN-gamma signal when PBMC were evaluated with NK-sensitive targets in the IFN-gamma ELISPOT assay. IFN-gamma ELISPOT and(51)Cr release assays showed excellent correlation suggesting that NK activity can be reliably evaluated with methods other than the traditional(51)Cr release assays.

Positive and negative selection of antigen-specific B cells in transgenic mice expressing variant forms of the V(H)1 (T15) heavy chain.

Four variant forms of the V1 (T15-H chain) gene are synthesized in mice. Each V1 variant pairs with a distinct L chain to produce a binding site having specificity for phosphocholine (PC). Transgenic mice expressing variant forms of the V1 gene were analyzed to elucidate the factors driving B cell selection into the peripheral repertoire. In all four lines of H chain transgenic mice analyzed, transgene expression caused complete allelic exclusion of endogenous H chains in the bone marrow (BM), whereas most splenic B cells expressed endogenous H chains. The number of sIgM(+) BM B cells and their sIg receptor number was reduced compared to that of normal transgene-negative controls, suggesting that B cells expressing transgene-encoded H chains were being negatively selected in the BM. Mice expressing autoreactive forms of the V1 transgene with lower affinity for PC (M603H and M167H) exhibit positive selection of PC-specific B cells into the spleen, whereas mice expressing the higher affinity T15H variant exhibited elevated PC-specific B cells in the peritoneal cavity but few V(H)1(+) splenic B cells. These data suggest that the higher affinity T15-id(+) B cells preferentially survive in the peritoneal cavity. When these H chain transgenes were crossed into the mu MT knockout mouse in which surface expression of endogenous H chains is blocked, the percent of splenic V(H)1(+) PC-specific B cells increased up to 5-fold and T15-id(+) B cells were detectable in the spleen of T15H mice. This implies that T15-id(+) PC-specific B cells can be selected into the periphery, but they compete poorly with follicular B cells expressing endogenous Ig.