Antibodies (Abs) that mediate antibody-dependent cellular cytotoxicity (ADCC) activity againstHIV-1 are of major interest. A widely used method to measure ADCC Abs is the rapid andfluorometric antibody-dependent cellular cytotoxicity (RFADCC) assay. Antibody-dependentkilling of a labelled target cell line by PBMC is assessed by loss of intracellular CFSE but retention ofmembrane dye PKH26 (CFSE-PKH26+). Cells of this phenotype are assumed to be derived fromCFSE+PKH26+ target cells killed by NK cells.We assessed the effector cells that mediate ADCC in this assay. Backgating analysis andphenotyping of CFSE-PKH26+ revealed that the RFADCC assay's readout mainly representsCD3-CD14+ monocytes taking up the PKH26 dye. This was confirmed for 53 HIV+plasmapurifiedIgG samples when co-cultured with PBMC from three separate healthy donors.Emergence of the CFSE-PKH26+monocyte population was observed upon co-culture of targetswith purified monocytes but not with purified NK cells. Image flow cytometry and microscopyshowed a monocyte-specific interaction with target cells without typical morphologicalchanges associated with phagocytosis, suggesting a monocyte-mediated ADCC process.We conclude that the RFADCC assay primarily reflects Ab-mediated monocyte function.Further studies on the immunological importance of HIV-specific monocyte-mediated ADCCare warranted. © 2012 Elsevier B.V. All rights reserved.
Kramski, M., Schorcht, A., Johnston, A. P. R., Lichtfuss, G. F., Jegaskanda, S., De Rose, R., ... Kent, S. J. (2012). Role of monocytes in mediating HIV-specific antibody-dependent cellular cytotoxicity. Journal of Immunological Methods, 384(1/2), 51-61. https://doi.org/10.1016/j.jim.2012.07.006