Introduction: Human neutrophil antigens 3 (HNA-3a and HNA-3b) are located on choline transporter protein 2 (CTL2) and expressed on neutrophils, lungs, kidneys and other human tissues. Alloantibodies directed against these antigens are formed from allogeneic exposure and are mainly associated with transfusion-related acute lung injury (TRALI) and immune neutropenias, however due to the presence of this antigen in renal tissue and the doubt that HNA-3 alloimmunization may also be involved in cases of kidney transplant rejection, the study of the frequency of anti-HNA-3 antibodies in this context becomes a focus of great clinical relevance.

Objective: Investigate the presence of anti-HNA-3 antibodies in the serum of patients who had kidney transplant rejection.

Materials and Methods: A total of 604 patients with kidney transplant rejection were included in the study. The Granulocyte Agglutination Test (GAT) was performed as a screening test in all individuals included in the present study with a panel of granulocytes from at least three individuals previously genotyped for all HNA systems. Positive samples in GAT were tested using the microsphere-based technique (LABSCreen Multi kit, One Lambda); we analyzed the normalized background values ≥ 10 and immunofluorescence ≥ 1000 as a positive result for HNA-3 antibodies. HNA-3 genotyping by PCR-RFLP was performed only in individuals who had a positive result in serological techniques.

Results: We detected 85/604 (14.1%) positive samples in GAT and 11/85 positive samples in both GAT and LSM multi for anti-HNA antibodies. Regarding the specificity of these 11 samples, 6 (54.5%) individuals presented an anti-HNA-3b antibody confirmed by genotyping (HNA- 3a/HNA-3a). In addition to the anti-HNA-3b antibodies, other specificities of anti-HNA antibodies were also identified: 1/11 (9.1%) anti-HNA-1a, 2/11 (18.2%) anti-HNA-1b, 1/11 (9.1%) anti-HNA-FCγRIIIb and 1/11 (9.1%) anti-HNA-2. Furthermore, 14/85 (16.5%) individuals had anti-HLA antibodies detected by LSM multi: 8/14 (57.1%) class I, 3/14 (21.4%) class II and 3/14 (21.4) class I and II. No patient presented simultaneously anti-HNA and HLA antibodies.

Conclusion: Our data show that HNA-3b alloimmunization has a greater tendency in patients who rejected kidney transplantation compared to healthy blood donors (1.0% vs. 0.2% respectively, p=0.05). These findings indicate that the detection of anti-HNA-3b antibodies may allow a better understanding of patients' immune response against renal allograft when no HLA antibody is identified supporting evidence of immunological risk.