CD4+ T lymphocytes enumeration by an easy-to-use single platform image cytometer for HIV monitoring in resource-constrained settings.
BACKGROUND
HIV monitoring in resource-constrained settings demands affordable and reliable CD4(+) T lymphocytes enumeration methods. We developed a simple single platform image cytometer (SP ICM), which is a dedicated volumetric CD4(+) T lymphocytes enumeration system that uses immunomagnetic and immunofluorescent technologies. The instrument was designed to be a low-cost, yet reliable and robust one. In this article we test the instrument and the immunochemical procedures used on blood from HIV negative and HIV positive patients.


METHODS
After CD4 immunomagnetic labeling in whole blood, CD4(+) T lymphocytes, CD4(+dim) monocytes and some nonspecifically labeled cells are magnetically attracted to an analysis surface. Combining with CD3-Phycoerythrin (PE) labeling, only CD3(+)CD4(+) T lymphocytes are fluorescently labeled and visible in a fluorescent image of the analysis surface. The number of CD4(+) T lymphocytes is obtained by image analysis. Alternatively, CD3 immunomagnetic selection in combination with CD4 immunofluorescent labeling can also be applied for CD4(+) T lymphocytes enumeration.


RESULTS
The SP ICM system was compared with two single platform flow cytometer (SP FCM) methods: tetraCXP and TruCount methods. The SP ICM system has excellent precision, accuracy and linearity for CD4(+) T lymphocytes enumeration. Good correlations were obtained between the SP ICM and the SP FCM methods for blood specimens of 44 HIV(-) patients, and of 63 HIV(+) patients. Bland-Altman plots showed interchangeability between the SP ICM and the SP FCM methods.


CONCLUSIONS
The immunolabeling methods and the instrumentation are simple and easy-to-handle for less-trained operators. The SP ICM system is a good candidate for CD4(+) T lymphocytes enumeration in point-of-care settings of resource-constrained countries.
Li
Xiao
X
University of Twente, Faculty of Science and Technology, Biophysical Engineering Group, Building Zuidhorst, Dienstweg 1, 7522 ND Enschede, The Netherlands. l.x.lixiao@tnw.utwente.nl
Ymeti
Aurel
A
Lunter
Björn
B
Breukers
Christian
C
Tibbe
Arjan G J
AG
Terstappen
Leon W M M
LW
Greve
Jan
J
D000328:Adult; D015704:CD4 Antigens / Q000032:analysis / Q000276:immunology; D018791:CD4 Lymphocyte Count / Q000191:economics / Q000295:instrumentation* / Q000379:methods; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology* / Q000821:virology; D003362:Cost-Benefit Analysis; D005434:Flow Cytometry / Q000295:instrumentation / Q000379:methods; D005455:Fluorescent Antibody Technique / Q000379:methods; D015658:HIV Infections / Q000097:blood / Q000175:diagnosis* / Q000276:immunology*; D006295:Health Resources / Q000191:economics; D006801:Humans; D019044:Image Cytometry / Q000191:economics / Q000295:instrumentation* / Q000379:methods; D018189:Immunomagnetic Separation / Q000379:methods; D010799:Phycoerythrin; D011237:Predictive Value of Tests; D015203:Reproducibility of Results; D014584:User-Computer Interface
D015704:CD4 Antigens; D010799:Phycoerythrin