A COMPARISON OF T-LYMPHOCYTE RESPONSIVENESS OF MONONUCLEAR LEUKOCYTES AND WHOLE BLOOD

Abstract

Cell cultures of whole blood were evaluated as a simplified replacement for mononuclear cells in the lymphocyte proliferation assay and the interleukin-2 bioassay. The cell cultures were stimulated with Phytohemaggluttin, Concanavalin A or anti-CD3 monoclonal antibody (OKT3) for various times of incubation. The results were measured as Tlymphocyte proliferation and interleukin-2 production. In the lymphocyte proliferation assay, both diluted whole blood and mononuclear cell cultures produced satisfactory results. Whole blood diluted 1:8, 1:16 or 1:32 could be used as a replacement for mononuclear cells. The optimal dilution of whole blood was shown to depend on the stimulant used. In the interleukin-2 assay, diluted whole blood cultures produced higher levels of Interleukin-2 (up to 25 Units/mL) than mononuclear cell cultures (less than 5 Units/mL). Low production of Interleukin-2 in the mononuclear cell cultures could possibly be attributed to our modified protocol which used human AS serum in place of the conventional fetal bovine serum. Human AB serum does not contain the same level of foreign antigen as fetal bovine serum, which could account for the low production of Interleukin-2. The recommended protocol for interleukin-2 production is whole blood diluted 1:8 or 1:16 stimulated with Phytahemaggluttin or Concanavalin A. Whole blood diluted 1:32 consistently showed lower interleukin-2 production than whole blood diluted 1:8 or 1:16 and is not a recommended dilution. The low IL-2 production is probably due to a limiting number of T-lymphocytes present at this dilution. OKT3 elicited no detectable levels of Interleukin-2 from any of the cell cultures and is not a recommended stimulant. The use of whole blood as the source of T-lymphocytes is a modification of both the lymphocyte proliferation assay and the Interleukin-2 bioassay. It serves to eliminate the large number of manipulation steps necessary to purify mononuclear cells from peripheral blood. As a result, a reduced volume of blood is required and less time and technical assistance are needed in using these assays to measure immune competence.