The sialome or display of sialic acids on the surface of human immune cells can vary according to immune response and activation state. Here, human peripheral blood mononuclear cells (PBMCs) were isolated and activated with anti-CD3 antibody and the cell surface sialome was quantified using a combination of click chemistry, confocal microscopy and flow cytometry techniques. Carbohydrate click chemistry was used to detect and measure the incorporation of an azido-m65odified sialic acid precursor molecule, N-acetylmannosamine (ManNaz) sugar into the PBMC surface sialome. Incorporation of sialic acid into the PBMC glycocalyx was visualized using copper-catalyzed click conjugation of Alexa 488 alkyne and confocal microscopy and further quantified using flow cytometry. The use of these methods indicate that regulating the sialome content on the surface of activated immune cells may be monitored during immunomodulatory responses and anti-inflammatory therapies.
Keywords: Click chemistry; Glycocalyx; Glycosylation; Immune cells; Inflammation; PBMCs; Sialome.
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