title: Application of flow cytometry in detection of red-cell-bound IgG in Coombs-negative AIHA.
abstract: Coombs negative autoimmune hemolytic anemia (AIHA) is characterized by laboratory evidence of in vivo hemolysis along with a negative direct antiglobulin test (DAT) performed by conventional tube technique (CTT) in clinically suspected AIHA patients. The sensitive gel test (GT) and flow cytometry (FC) can effectively diagnose such patients where CTT does not detect low level of red cell autoantibodies. We investigated the use of FC in the serological evaluation of CTT DAT negative AIHA and its comparison with GT DAT. Of the 50 patients with suspected AIHA, CTT DAT was negative in 5 patients (Coombs negative AIHA). GT DAT could detect red cell autoantibodies in 4 of these 5 patients. Monospecific GT DAT showed IgG and/or C3d as the responsible autoantibody. FC was considered as reactive when MFI was >3.6 (mean of 20 healthy negative volunteers +2SD). FC was reactive in all five Coombs negative AIHA patients. The mean MFI in five known CTT DAT positive samples taken for comparison was significantly higher compared to 5 DAT negative AIHA (18.3 +/- 7.78 vs. 7.88 +/- 1.35, p < 0.05). There was poor correlation between strength of GT DAT and MFI by FC. We conclude that FC is more sensitive test than the CTT and helps in the serological diagnosis of Coombs negative AIHA. However, in resource poor settings, GT DAT can be a good alternative to FC.
authors.affiliation: Department of Transfusion Medicine, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow, India. rkcchaud@sgpgi.ac.in
authors.forename: Rajendra
authors.initials: R
authors.lastname: Chaudhary
authors.forename: Sudipta S
authors.initials: SS
authors.lastname: Das
authors.forename: Ritu
authors.initials: R
authors.lastname: Gupta
authors.forename: Dheeraj
authors.initials: D
authors.lastname: Khetan
mesh_terms: D000293:Adolescent; D000328:Adult; D000744:Anemia, Hemolytic, Autoimmune / Q000097:blood / Q000175:diagnosis* / Q000453:epidemiology / Q000209:etiology; D000888:Antibodies, Anti-Idiotypic / Q000097:blood*; D001172:Arthritis, Rheumatoid / Q000097:blood / Q000150:complications; D015933:Complement C3d / Q000276:immunology; D003298:Coombs Test; D003906:Developing Countries; D004912:Erythrocytes / Q000276:immunology*; D005188:False Negative Reactions; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D006462:Hemolytic Plaque Technique; D019693:Hepatitis, Autoimmune / Q000097:blood / Q000150:complications; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood*; D015994:Incidence; D007194:India / Q000453:epidemiology; D008297:Male; D008875:Middle Aged; D011446:Prospective Studies; D012680:Sensitivity and Specificity
chemical_list: D000888:Antibodies, Anti-Idiotypic; D007074:Immunoglobulin G; D015933:Complement C3d