Immunohematological characterization of direct antiglobulin test-negative autoimmune hemolytic anemia through simple, sensitive methods: Experience from a tertiary care hospital in India

Background and Objectives: A negative DAT does not rule out AIHA. The incidence of this clinical entity known as DAT negative AIHA has been reported to be 2-4%. Sensitive techniques like enzyme linked antiglobulin test (ELAT), flow cytometry (FC), complement-fixation antibody consumption test etc. have been described for diagnosis of DAT negative AIHA. Majority of blood bank laboratories lack these advanced methods. Here we share our experience of investigating DAT negative AIHA using simple, sensitive methods which are otherwise less practised. Methods: The prospective study included 377 anemic patients clinically suspected of AIHA. Blood samples received in blood bank were subjected to polyspecific DAT using both conventional tube test (CTT) and column agglutination technique (CAT). Polyspecific DAT negative results were further evaluated using sensitive but simple methods. Hematological and biochemical parameters of all patients were obtained from hospital information system. In vivo hemolysis was categorized as per the criteria established by previous workers. SPSS statistical software (version 13, USA) was used for all statistical evaluation. Results: Of the final 353 clinical AIHA patients evidence of autoimmunization by CTT was observed in 335. Where DAT negative AIHA was observed in 18 (5.1%) patients, 14 showed evidences of autoimmunization by extended sensitive methods. Four patients responded well to AIHA therapy despite DAT negativity by available methods. The mean Hb, Retic, S. Bil and LDH were found to be 6.9 g/dL, 3.25%, 2.8 mg/dL and 512 IU/mL respectively. Conclusion: We conclude that DAT negative patients with clinical suspicion of AIHA and positive laboratory evidences should be evaluated for the presence of autoantibody by alternate sensitive methods which are otherwise less practiced. Blood banks may establish these useful simple techniques and stick to the defined protocols to diagnose DAT negative AIHA.