Background and objectives: Serologic tests for granulocyte antibodies, i.e., the granulocyte agglutination test and the granulocyte immunofluorescence test, require panels of typed granulocytes that cannot be preserved for more than a few hours. We have developed a new method in which granulocyte antigens, extracted into saline containing 3% sucrose, are coated onto U-type Terasaki plates. With this new method, we evaluated the micro-mixed passive hemagglutination test (EG-MPHA) for screening for granulocyte antibodies.
Materials and methods: We tested the ability of the EG-MPHA to detect granulocyte antigens using 5 human antibodies specific for NA1, NA2, NB1, 5b, and Sar(a), and 8 different monoclonal antibodies for NA1, CD11a, CD11b, CD13, CD16, CD18 and HLA class I. Sera from 94 alloimmunized patients were screened by the chloroquine-treated EG-MPHA method.
Results: NA1, NA2, NB1, 5b, Sar(a), CD11a, CD11b, CD13, CD16, CD18 and HLA class I antigens were present in the extracted granulocyte antigen preparation. CD11b and HLA class I antigens were removed when the extracted granulocyte antigens were treated with chloroquine. Granulocyte antibody screening of sera from alloimmunized patients showed that approximately 30% of the anti-HLA-positive and 10% of the anti-HLA-negative sera were positive for granulocyte antibody by the chloroquine-treated EG-MPHA. The extracted granulocyte antigen panels could be stored frozen for at least 1 year at -80 degrees C.
Conclusion: This new method is preferable for screening for granulocyte antibodies. In addition, it has the advantage of requiring only 5 microl of serum for each test.