Objective To detect the reliability of α-1，3-galactosyl（α-Gal）antigen knockout and the expression of human protein CD46（hCD46）on the surface of genetically modified pigs at the protein level, to provide reliable donors for xenotransplantation. Methods A total of 500 μl blood from wild type（WT），α-1，3-galactosyl transferase knockout（GTKO），hCD46 and GTKO/hCD46 Bama miniature swine were collected for peripheral blood mononuclear cell（PBMC）isolation. Expression of α-Gal antigen was detected by flow cytometry after coincubation with fluorescein isothiocyanate-Griffonia simplicifolia isolectin B4（FITC-GSIB4）and PBMC. Expression of hCD46 protein was detected by flow cytometry after co-incubation of PBMC with fluorescein isothiocyanate-CD46 membrane protein antibody（anti-CD46-FITC）. Results The α-Gal expression on the surface of PBMC were negative in GTKO and GTKO/hCD46 Bama pigs and positive in WT and hCD46 Bama pigs，consistent with the sequencing results. The hCD46 protein was detected positively on the surface of PBMC of hCD46 and GTKO/hCD46 Bama pigs and was negative in WT and GTKO Bama pigs，which was consistent with the results of genotype identification after birth. Conclusion A method for visually detecting the expression of α-Gal antigen and hCD46 protein in donor porcine using microscale blood was established.