Determination of CD4 lymphocyte percentages using a new simplified and affordable no lyse flow cytometric protocol: a pilot study / Bestimmung der Lymphozyten-abhAOngigen CD4-Prozentwerte mit Hilfe eines neuen, vereinfachten und kostengA14nstigen, durchflusszytometrischen Protokolls ohne Lyse: Eine Testuntersuchung

Although the pathogenesis of HIV infection is independent of age, specific considerations have to be taken into account for the treatment of infants, children and adolescents. In addition to more cautious drug therapy, it is proposed to follow up disease progression by repeated measurement of CD4 lymphocyte percentage (CD4%). Unfortunately, this requires complicated, expensive analytical techniques and complex sample preparation protocols. This is one of the reasons why treatment of the increasing number of children suffering from HIV/AIDS in developing countries remains a challenge. We evaluated a new and simplified no lyse no wash protocol to measure CD4% on a volumetric flow cytometer. Eight blood samples of healthy and HIV-infected individuals were sent to five German hospital laboratories (centres BaF) and were measured by their flow cytometric in-house techniques. The results were compared to those of centre A using the volumetric technique with the new sample preparation protocol. Comparative data with established protocols and flow cytometric techniques showed good correlations for the determination of leukocytes and for lymphocytes. Comparing CD4% results of centre A (new protocol and volumetric measuring technique) with the highest and lowest values of centres BaF by Bland-Altman analysis yielded a bias of a1.02% (SD 3.16) and 4.27% (SD 3.72) over the whole range (4.01%a70.56%), respectively. A comparably simple three-step protocol as introduced for absolute CD4+ cell count can be used to calculate CD4% only by the addition of two antibodies (CD4 and CD45). The cost per test is in the range of an2.50 and therefore far below current prices of an15.00aan40.00.