Difficulties in precise quantitation of CD4+ T lymphocytes for clinical trials: a review.

Maintenance of CD4+ T helper lymphocyte counts has been used as a surrogate marker of efficacy for drugs in the treatment of AIDS. In a multicenter clinical trial, subtle improvement of CD4+ T cell counts may be masked and misinterpreted if care is not paid to likely sources that can contribute to the variability of measurement of CD4+ T lymphocytes. This review addresses major areas that can contribute to the variability of measurement of CD4+ T lymphocytes, with emphasis on applications to multicenter clinical trials, and proposes areas of improvement that may not be well recognized by the medical community. Whereas there are excellent guidelines for immunophenotyping, equal attention is needed in hematologic enumeration of WBC and absolute lymphocytes. In particular, allowing the margin of error acceptable to blood cell standards for HIV-infected specimens is unsatisfactory. Special attention should also be given to the stability of lymphocytes in the anticoagulant during storage, the lysing method, the quality assurance programs as well as intrasubject fluctuations which may be derived from exercise, medications and diurnal variations. Awareness of these contributing factors by physicians and technical analysts will expedite the discovery of potential therapy in the treatment of AIDS. For a multicenter clinical trial, it is advisable to select a centralized laboratory adopting a uniform protocol with regard to sample preparation and handling, using more stringent quality controls for hematologic analysers, calibration of instruments and immunophenotyping. Pending a true reference standard that can monitor the variation of the entire analytical procedure, we anticipate that future interlaboratory quality assurance programs will include absolute T lymphocyte count, an important parameter for assessing the accuracy and consistency of CD4+ T helper cell counts generated from a laboratory.