DTT Test: A New Method to Differentiate IgM and IgG Erythrocyte Antibodies¹

Abstract

A new test to determine the immunoglobulin class of erythrocyte antibodies is presented. Dithiothreitol (DTT), at a concentration of 0.01 M, has been used in a rapid 15 min, 37°C incubation test system. Dialysis is not required. The accuracy of this new DTT test was compared to a standard 2-ME procedure. Human sera containing saline-acting and incomplete varieties of erythrocyte antibodies were tested. Identical results were obtained by both methods with 292 of 302 (96.7%) test sera. Anion-exchange chromatography was employed to produce erythrocyte antibody solutions of known immunoglobulin class. Incubation of these solutions with DTT decreased the immunologic reactivity of IgM erythrocyte antibodies, without affecting the activity of IgG varieties.

A Simple Technique for the Inactivation of IgM Antibodies Using Dithiothreitol

“…All of these studies were carried out before we learned of Pirofsky and Rosner's [6] work, and involved a 30-minute incubation of DTT and test serum. Because Pirofsky and Rosner [6] used a 15-min in cubation, seven of the sera reported above were retested using both a 15-and a 30-min incubation with DTT.…”

“…Because Pirofsky and Rosner [6] used a 15-min in cubation, seven of the sera reported above were retested using both a 15-and a 30-min incubation with DTT. There was no difference in the extent of inactivation of the antibody activity, whether a 15-or a 30-min incubation was used (table V).…”

“…This odorless compound was prepared to maintain monothiols such as coenzyme A in the reduced state, but also has the property of reducing disulfides quantitatively [1], Because of these properties of DTT, we have investigated its value as a simple method for the inactivation of IgM red cell antibodies [4]. Pirofsky [5] had previously mentioned the possible use of DTT for this purpose and since initiation of our studies, Pirofsky and Rosner [6] have re ported detailed results of their work.…”

IgM Anti-D as Auto-Antibody in a Case of ‘Cold’ Auto-Immune Haemolytic Anaemia

“…Table I , and anti-C4 (Behring-Hoechst, Hounslow, UK) were used in direct antiglobulin tests (DAT). Reduction of IgM antibodies was carried out using dithiothreitol (DTT) [9], and elutions were per formed using diethyl ether [10]. …”

Complement-mediated Killing of Ureaplasma Urealyticum by Antibody Directed Against Components of the Growth Medium

“…To inactivate IgM (Pirofsky and Rosner, 1974;Olson et al, 1976), sera were treated with 4 0 m~ dithiothreitol (DTT; Calbiochem) at room temperature for 1 h and then dialysed against PBS overnight at 4°C. Sera treated only with PBS were included as controls.…”