Detection of xenoantibodies using a simple flow cytometric assay

Higher primates, including humans, have high levels of preexisting naturall y circulating antibodies that predominantly recognize the epitope Gal (1,3- Gal), which is highly expressed on the surface of xenogenic cells. Depositi on of these antibodies on the endothelial cell surface of vascularized xeno grafts leads to an activation of the classical pathway of the complement sy stem, resulting in tissue ischemia and necrosis with rapid demise of the xe nograft. This hyperacute rejection (HAR) is always a major barrier in xenog raft transplantation and should be minimized by accurately monitoring the n aturally occurring antibodies. In the present study, we utilized a simple a nd rapid flow cytometric (FCM) assay to monitor the presence of these natur ally occurring antibodies. We found that the FCM assay is very effective in measuring human antibodies bound to the xenogenic cells, which cause cytot oxicity. This assay could be useful in the pre- and post-xenotransplantatio n monitoring of xenoantibodies, thus, helping in the development of strateg ies to block the binding of preformed human antibodies to the xenograft in order to overcome the problem of HAR.