We present a simple system for CD4 and CD8 counting for point-of-care HIV staging in low-resource settings. Automatic sample preparation is achieved through a dried reagent coating inside a thin (26 μm) counting chamber, allowing the delayed release of fluorochrome conjugated monoclonal antibodies after the filling of the chamber with whole blood by capillary flow. A custom-built image cytometer is used to capture fluorescence images representing more than 1 μl of blood. The thin layer of blood in combination with the large image area allows the use of whole blood from a finger prick without the need for dilution, lysis or cell enrichment. Automatic cell counting of CD4(+) and CD8(+) T-lymphocytes correlates well with results obtained by flow cytometry.

Background For resource‐poor countries, affordable methods are required for enumeration of CD4+ T lymphocytes of HIV‐positive patients. For infants, additional determination of CD4/CD8 ratio is needed. Methods We determine the CD4+ and CD8+ T lymphocytes as the CD3+CD4+ and CD3+CD8+ population of blood cells. Target cells are CD3‐immunomagnetically separated from the whole blood, and CD4‐Phycoerythrin and CD8‐PerCP immunofluorescently labeled. A point‐of‐care single platform image cytometer was developed to enumerate the target CD3+CD4+ and CD3+CD8+ populations. It has light‐emitting diodes illumination, is fully computer‐controlled, operates from a 12 V battery, and was designed to be cheap and easy‐to‐handle. Target cells are imaged on a CCD camera and enumerated by an image analysis algorithm. The cytometer outputs the absolute number of CD4+ and CD8+ T lymphocytes/μl and CD4/CD8 ratio. Results The quality of the cell images obtained with the cytometer is sufficient for a reliable enumeration of target cells. The image cytometer achieves an accuracy of better than 10% in the range of 50–1700 cells/μl. Analysis of blood samples from HIV patients yields a good agreement with the TruCount method for CD4 and CD8 count and CD4/CD8 ratio. Conclusions The image cytometer is affordable (component costs $3,000), compact (25 × 25 × 20 cm3), and uses disposable test materials, making it a good candidate to monitor progression of immunodeficiency disease in resource‐poor settings. © 2007 International Society for Analytical Cytology.

A DNA‐sensing platform is developed by exploiting the easy surface functionalization of metal–organic framework (MOF) particles and their highly parallelized fluorescence detection by flow cytometry. Two strategies were employed to functionalize the surface of MIL‐88A, using either covalent or non‐covalent interactions, resulting in alkyne‐modified and biotin‐modified MIL‐88A, respectively. Covalent surface coupling of an azide‐dye and the alkyne–MIL‐88A was achieved by means of a click reaction. Non‐covalent streptavidin–biotin interactions were employed to link biotin–PNA to biotin–MIL‐88A particles mediated by streptavidin. Characterization by confocal imaging and flow cytometry demonstrated that DNA can be bound selectively to the MOF surface. Flow cytometry provided quantitative data of the interaction with DNA. Making use of the large numbers of particles that can be simultaneously processed by flow cytometry, this MOF platform was able to discriminate between fully complementary, single‐base mismatched, and randomized DNA targets.

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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