Background: Recent studies highlighted the presence of anti-\alpha-Gal antibodies in patients implanted with commercial bioprosthetic heart valves (BHVs). BHVs expose residual \alpha-Gal xenoantigen and their recognition by the circulating anti-Gal antibodies leads to opsonization of the device’s tissue component with the consequent triggering of a deterioration pathway that culminates with calcification. Small animal models such as mice and rats have been broadly involved in the in vivo testing of biomaterials by subcutaneous implantation, especially for the effectiveness of BHVs anti-calcific treatments. However, since models employed for this purpose express \alpha-Gal antigen, the implantation of BHVs’ leaflets does not elicit a proper immunological response, so the calcification propensity may be dramatically underestimated. Methods: An \alpha-Gal knockout (KO) mouse model has been created, using the CRISP/Cas9 approach, and adopted to assess the calcification potential of commercial BHVs leaflets through the surgical implantation in the back subcutis area. Calcium quantification was performed by inductively coupled plasma analysis; immune response against the BHVs leaflets and \alpha-Gal silencing was evaluated through immunological assays. Results: Two months after the implantation of commercial BHV leaflets, the anti-Gal antibody titers in KO mice doubled when compared with those found in wild-type (WT) ones. Leaflets explanted from KO mice, after one month, showed a four-time increased calcium deposition concerning the ones explanted from WT. The degree of silencing of \alpha-Gal varied, depending on the specific organ that was assessed. In any case, the animal model was suitable for evaluating implanted tissue responses. Conclusions: Such mouse model proved to be an accurate tool for the study of the calcific propensity of commercial BHVs leaflets than those hitherto used. Given its reliability, it could also be successfully used to study even other diseases in which the possible involvement of \alpha-Gal has been observed.