New method for overcoming the interference produced by anti-CD38 monoclonal antibodies in compatibility testing

Abstract

Background - Plasma of patients taking anti-CD38 monoclonal antibodies (MoAbs) leads to panagglutination in the indirect antiglobulin test (IAT), that can mask clinically significant alloantibodies. Dithiothreitol (DTT) treatment of test RBCs is the more widespread method for avoiding this interference. Current DTT 0.2 mol/L method is time consuming and damages several red blood groups antigens. This study aims to evaluate low concentration DTT treatment of RBCs adapted for gel testing.
Materials and methods - Four DTT concentrations (0.01, 0.02, 0.03, and 0.04 mol/L), and three gel test brands were evaluated on six DARA patient’s samples. Briefly, the method consists of pipetting 50 μL of 0.8% RBCs on AHG micro columns, followed by 25 μL of DTT, thoroughly mixing and 15 min incubation at 37 °C. Then, 25 μL of serum/plasma is added to proceed to IAT. In order to asses the effect of DTT 0.04 mol/L on different blood group antigens, serial dilutions of sera containing anti-K, -k, -Kp^b, -Lu^b, -Yt^a and anti-JMH antibodies were tested against DTT-RBCs. One sample of a DARA patient with known alloantibodies as well as samples of two patients inoculated with anti-K and anti-Fy^a were evaluated.
Results - RBCs treatment with DTT 0.04 mol/L for 15 min completely eliminated anti CD38 panagglutination in all samples studied and worked with different reactivity intensities in IAT and gel brands. The new method allowed the detection of underlying anti-D, anti-E, anti-K and anti-Fy^a alloantibodies. Titration assays  demonstrated no denaturation of Kell, Lutheran, Cartwright and JMH antigens.
Discussion - The new DTT method adapted for gel testing is efficacious, simple and only adds 15 min over regular IAT. Pheno/genotyping before DARA treatment or transfusion of K negative RBCs may be unnecessary.