Carboxyfluorescein succinimidyl ester‐based proliferative assays for assessment of T cell function in the diagnostic laboratory

作者：DA Fulcher, SWJ Wong · 发表于：Immunology and Cell Biology · 年份：1999 · DOI：10.1046/j.1440-1711.1999.00870.x · 被引用次数：114 · 研究领域：Immunodeficiency and Autoimmune Disorders、Cytomegalovirus and herpesvirus research、T-cell and B-cell Immunology

Immune deficiency diseases are often accompanied by abnormalities in one or both arms of the specific immune system. Impairment can often be detected as a decrease in the number of T or B lymphocytes or their products in the circulation, but questions are often asked as to the functional capabilities of T lymphocytes in patients with recurrent infections. Function of T cells has traditionally been measured by their uptake of [3H]- thymidine following stimulation with antigen or mitogen in vitro. However, the ability of carboxyfluorescein succinimidyl ester (CFSE) to label lymphocytes intracellularly and track their mitotic activity by progressive two-fold reduction in fluorescence intensity prompted an alternative methodology based on flow cytometry, an approach which has the advantage of allowing specific gating on particular T cell subsets and simultaneous assessment of activation markers. This method was therefore evaluated for T cell responses to mitogen and antigen. Phytohaemagglutinin-induced blast transformation of CFSE-labelled T cells was reflected by an increase in forward and orthogonal light scatter and a progressive two-fold decrease in CFSE fluorescence intensity. These changes allowed the derivation of various measures of mitotic activity, which correlated well with [3H]-thymidine uptake. Patients with T cell functional deficiencies showed impairment in their responses by both assays, whereas the CFSE-based assay demonstrated that impaired blastogenesis was not...