Details

Autor(en) / Beteiligte

• Otsu, Makoto
• Tanabe, Yuji
• Iwakiri, Ayako
• Arima, Kazuna
• Uchiyama, Anna
• Yamamoto, Marina
• Ohtani, Shinichi
• Endo, Hiroshi
• Komoto, Mina
• Miyazaki, Koji

Titel

A report on a modified protocol for flow cytometry‐based assessment of blood group erythrocyte antigens potentially suitable for analysis of weak ABO subgroups

Ist Teil von

• Transfusion (Philadelphia, Pa.), 2023-03, Vol.63 (3), p.463-469

Ort / Verlag

Hoboken, USA: John Wiley & Sons, Inc

Erscheinungsjahr

2023

Quelle

MEDLINE

Beschreibungen/Notizen

• Background Flow cytometry (FC) has proven its utility in scrutinizing AB antigen expression in red blood cells (RBCs), cooperating with serological tests for accurate blood group typing. However, technical difficulties may impair the characterization of weak ABO subtypes when background noises appear at non‐negligible levels. Study design and methods We sought to establish an FC method that could prevent antibody‐induced hemagglutination and an increase in cellular autofluorescence, two major issues inherent to RBC‐FC analysis of AB expression. We optimized fixatives, multicolor‐staining protocols, and sequential gating strategies. Blood samples from weak ABO subtype cases, Bm and Ael, were analyzed with the established protocol. Results The optimized mixture of glutaraldehyde and formaldehyde successfully generated fixed RBCs resistant to agglutination while maintaining low autofluorescence. These features allowed co‐staining of leukocyte‐ and erythrocyte‐markers, which enabled sequential gating strategies facilitating the precise AB antigen analysis in purely single RBCs with minimum background noises. By the established FC analysis, we could detect in the Bm sample a small RBC population exhibiting weak B antigen expression. The assay also proved it feasible to identify a small population (0.04%) of RBCs weakly expressing the A antigen in the Ael sample confirmed as harboring a rare c.816dupG ABO variant allele. Conclusion The RBC‐FC analysis described here allows the detection of AB antigens weakly expressed in RBCs while achieving minimum background noise levels in negative control samples. Overall, the modified protocol provides a quick and reliable assay valuable in transfusion medicine and is potentially applicable to the characterization of rare weak ABO variants.