Flow cytometric analysis of cell lineage and immune activation markers using minimal amounts of human whole blood—Field method for remote settings

Abstract

Remote laboratory settings – such as those where studies on neglected tropical diseases are performed – often lack specialized equipment required for flow cytometric analysis of immune cell subsets, which complicates evaluations on a single cell level using peripheral blood. Our aim was to establish a method to use whole blood for phenotypic characterization of T-cells for specific markers including CD3, CD4, HLA-DR, CD38, CCR5, CD27, CD45RA, CD25, and FoxP3. This method uses 100 μL whole blood which is stained for extracellular markers, lysed, and cryopreserved at −20 °C at a field laboratory before transferring to liquid nitrogen for long-term storage and transportation. Cells can then be transported to a central laboratory for flow cytometry analysis. The method was initially established using samples from healthy donors; expression levels after cryopreservation were comparable to fresh whole blood samples from the same individuals. Moreover, data sets were also comparable to those which were stored in liquid nitrogen for up to one year. The method was then transferred to field studies in a remote area of Ghana which was used to observe its practicality and robustness in limited resource settings. Collectively, the low amount of whole blood (such as that taken from a finger prick), lack of any specialized equipment, and ease of use make this method suitable for utilization in remote field locations.