Background: Xenotransplantation using genetically engineered pig organs is considered a valid potential for the human donor shortage. However, in addition to acute and chronic rejection of pig-to-human xenotransplantation by immunology barriers, the blood type incompatibility between pig and human is also one of the reasons for the failure of xenografts. Different from people, pigs only have blood types A and O. Currently, the reliable method to blood type pigs is by immunofluorescence and immunohistochemical staining of tissues (such as kidneys), but the application of this method is limited by the relative difficulty of obtaining material and the trauma associated with tissue biopsy. Although the immunofluorescence method of buccal mucosa smear is relatively simple and feasible, there is a risk of contamination with food debris and obtaining too few oral mucosal cells. In the present study, we have found a simple and reliable method for blood typing of pigs.
Methods: The expression of blood group antigen on erythrocyte surface was directly detected by flow cytometry using commercial monoclonal antibodies against blood group antigen in the donor pigs (7 wild-type and 4 gene-edited pigs) undergoing allogeneic or xenogeneic kidney transplantation in our center. Clinically common methods such as slide agglutination and reverse cassette gel were used as controls, and the accuracy was verified by immunofluorescence using kidney tissue and oral buccal mucosa smears.
Results: Kidney tissue and oral mucosa smear were detected in 8 cases with blood group A and 3 cases with blood group O. However, the blood type of pigs detected by slide agglutination method was O-type, and the blood type of pigs detected by reverse cassette gel method was difficult to determine. The results of blood group antigen detection on the surface of porcine red blood cells using our flow cytometry method were consistent with the immunofluorescence results of kidney tissue and oral mucosa In addition, our results also suggest that there are differences in the expression of A antigen on the surface of pig RBC and kidney, and flow cytometry can detect weak expression of A antigen on the surface of RBC. Conclusions: Although the expression of A antigen on the surface of pig RBC is weak and difficult to be detected by routine clinical blood group detection methods, flow cytometry using commercial anti-A monoclonal antibody can accurately detect the expression of A antigen on porcine red blood cells, so it is a simple and reliable method.