A manual bead assay for the determination of absolute CD4+ and CD8+ lymphocyte counts in human immunodeficiency virus-infected individuals

Abstract

CD4+ T lymphocytes are currently the most common surrogate marker indicating disease progression in individuals infected with human immunodeficiency virus (HIV). Since the cost of enumerating lymphocyte phenotypes is quite high, an inexpensive bead assay analyzed by light microscopy (cytosphere assay; Coulter Corporation, Hialeah, Fla.) was developed as an alternative method for counting CD4+ and CD8+ T lymphocytes. To evaluate the reliability of the cytosphere assay, heparinized blood was collected from 117 HIV-infected individuals and tested for both CD4+ and CD8+ lymphocytes by flow cytometry and the cytosphere assay. The Pearson correlation coefficient of the cytosphere assay compared with that of flow cytometry for CD4+ T lymphocytes was 0.93, with mean values +/- standard deviations of 534 +/- 509 by flow cytometry and 499 +/- 477 by the cytosphere assay. The correlation coefficient for CD8+ T lymphocytes was 0.86, with mean values of 831 +/- 543 by flow cytometry and 746 +/- 472 by the cytosphere assay. The sensitivity and specificity of the cytosphere assay in determining absolute CD4+ T-lymphocyte counts of less than 200/microliters were 97.6 and 94.7%, respectively. The positive predictive value was 90.9%, and the negative predictive value was 98.6%. The cytosphere assay was highly correlative to flow cytometry in determining CD4+ and CD8+ T-lymphocyte counts among HIV-infected patients. The ease and limited resources needed to perform this test make it ideal in developing countries and other areas where technology and finances are limited.