Detection of human monocyte-reactive alloantibodies by flow cytometry after selective downmodulation of the Fc receptor I.
Monocyte-reactive human alloantibodies may be of importance in situations such as transfusion reactions and bone marrow and kidney transplantation. So far, only complement-binding monocyte-reactive antibodies can be detected with a cytotoxicity assay. No antiglobulin assays are yet available that also detect noncomplement-fixing monocyte-reactive antibodies. The binding of monomeric IgG with high affinity to the Fc receptor I (FcRI) on monocytes has severely hampered the development of such an assay until now. We report on the selective removal of the FcRI from monocytes to test human sera in a flow cytofluorometry assay for the presence of monocyte-reactive IgG alloantibodies. Selective downmodulation of FcRI was accomplished by incubating the cells with murine monoclonal antibodies against FcRI followed by a second incubation with goat-antimouse IgG polyclonal antibodies. With such modified cells, human complement-binding and noncomplement-binding IgG and IgM alloantibodies against polymorphic determinants of the HLA class I and II glycoproteins, the human monocyte antigen system and polymorphic antigenic determinants of the LFA complex, can be detected in a sensitive and reproducible manner.
Kuijpers
R W
RW
Department of Immunologic Haematology, Central Laboratory of The Netherlands Red Cross Blood Transfusion Service, Amsterdam.
Dooren
M C
MC
von dem Borne
A E
AE
Ouwehand
W H
WH
D000017:ABO Blood-Group System; D000941:Antigens / Q000276:immunology; D003165:Complement System Proteins / Q000378:metabolism; D015536:Down-Regulation; D005260:Female; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D007518:Isoantibodies / Q000032:analysis*; D016169:Lymphocyte Function-Associated Antigen-1 / Q000235:genetics; D009000:Monocytes / Q000276:immunology*; D009504:Neutrophils / Q000276:immunology; D011110:Polymorphism, Genetic; D011961:Receptors, Fc / Q000502:physiology*
D000017:ABO Blood-Group System; D000941:Antigens; D007518:Isoantibodies; D016169:Lymphocyte Function-Associated Antigen-1; D011961:Receptors, Fc; D003165:Complement System Proteins