Verification of column agglutination technology with conventional tube technology for naturally occurring antibody titration

Abstract

Background: Pre-transfusion compatibility testing is performed in order to prevent transfusion of incompatible donor red cells that might result in an immune mediated hemolytic transfusion reaction, thereby forming a critical element in the process of transfusion to enhance blood safety. Titers are generally determined with a semi-quantitative assay by serial double-fold dilution. Several different techniques have been employed for titration, of which the conventional tube test (CTT) and column gel card test –Column Agglutination Technique (CAT) are the most frequently used. Historically, CTT has been used as the standard technique for immunohaematological studies, such as direct antihuman globulin test for the diagnosis of autoimmune haemolytic anaemia and in screening for specific antibodies in transfusion medicine. Study Design and Methods: For this study, 51 plasma samples from donors were subjected to antibody titration using the two techniques CAT and CTT. For the purpose of standardization and reproducibility of results, all 51 samples were tested by two technicians and the results obtained were a total of 70 antibody titers. Dilutions were continued to a titer of 1:1024 with the aim of allowing for at least two negative tubes beyond the titer end-point. The statistical analysis of the data has been done in MS-Excel. Results: Out of the 70 titer samples, 57 samples showed identical titer values in the methods performed by both the technicians. 8 samples showed titer values higher in Technician 2 by one dilution as compared to Technician 1, whereas 5 samples showed titer values lower in Technician 2 when compared to Technician 1 (one dilution low in 4 samples and two dilutions low in 1 sample). Conlcusion: CAT titers demonstrated up to a twofold difference as compared to CTT titres, as demonstrated in the figures presented. This disparity is not unusual and can be attributed to the lack of standardization which can be reduced by Column Agglutination Technology. CAT uses specific volumes of red blood cells and the reagents are dispensed by pipettes. Also, the procedure is technically simpler due to the elimination of washing steps and results in well-defined end points. Column Agglutination Technique offers not only sensitivity, but also the best titer turnaround time, eliminating almost 45 minutes of incubation period alone. The results of testing by CAT are comparable to CTT but have the advantages of ease of performance, better standardization of cells, most importantly stable results and reproducibility. Implementation of antibody titers by the gel method would clinically benefit the management of ABO incompatible solid-organ transplant patients.

Keywords

• Antibodies
• antibody titration
• column agglutination technique
• conventional tube technology
• standardization
• validation