It has been reported that acute allograft rejection is associated with heightened expression of the peripheral blood lymphocyte (PBL) early activation marker CD69 and that this may serve as a potential biomarker of rejection. This study sought to determine whether PBL CD69 expression correlates with both acute clinical and subclinical renal allograft rejection as well as clinically inapparent cytomegalovirus (CMV) infection. Flow cytometric determination of PBL CD69 expression was performed at the time of clinical and protocol biopsies (n = 131) in 45 renal transplant recipients. Nineteen patients also underwent weekly monitoring of PBL CD69 expression for the initial 15 wk after transplantation. Simultaneous screening for CMV viremia was performed with a semiquantitative PCR assay. No differences were seen in either CD4+ or CD8+ lymphocyte CD69 expression between the biopsy diagnoses. CMV viremia however, independent of rejection, was associated with greater CD69 expression on CD8+ lymphocytes (17.8 ± 10.4% versus 9.6 ± 4.8%; P < 0.0001) but not CD4+ lymphocytes. No individuals experienced clinical CMV disease. Weekly monitoring of PBL CD69 expression did not change coincident with the diagnosis of rejection; however, CMV viremia coincided with a substantial rise in the proportion of CD8+69+ lymphocytes in a number of individuals. Thus, PBL CD69 expression is neither sensitive nor specific for the noninvasive diagnosis of renal allograft rejection. Furthermore, clinically inapparent CMV viremia is associated with heightened expression of this activation marker on CD8+ lymphocytes. This latter finding suggests that clinically inapparent CMV viremia may be a potential confounder for biomarkers of rejection that examine peripheral blood lymphocytes. E-mail: mkarpinski@hsc.mb.ca