CHARACTERIZATION OF SPECIFIC BINDING OF A HUMAN IMMUNOGLOBULIN-M MONOCLONAL-ANTIBODY TO LIPOPOLYSACCHARIDE AND ITS LIPID-A DOMAIN

The human immunoglobulin M monoclonal antibody HA-1A was first described as an antibody which bound specifically to the lipid A region of lipopolysaccharide (LPS) (N. N. H. Teng, H. S. Kaplan, J. M. Herbert, C. Moore, H. Douglas, A. Wunderlich, and A. Braude, Proc. Natl. Acad. Sci. USA 82:1790-1794, 1985) and provided significant protection when administered to patients with gram-negative bacteremia and shock (E. J. Ziegler, C. J. Fisher, Jr., C. L. Sprung, R. C. Straube, J. C. Sadoff, G. E. Foulke, C. H. Wortel, M. P. Fink, R. P. Dellinger, N. N. H. Teng, 1. E. Allen, H. J. Berger, G. L. Knatterud, A. F. LoBuglio, C. R. Smith, and the HA-1A Sepsis Study Group, New Engl. J. Med. 324:429-436, 1992). Since that original report, questions have arisen in the scientific literature concerning the specificity of this antibody in LPS and/or lipid A binding. Experiments have, therefore, been carried out with a variety of assay formats to determine the capacity of this HA-IA antibody to bind to lipid A and LPS. Direct binding experiments with a sensitive enzyme-linked immunosorbent assay (ELISA) system have established that HA-IA will bind to purified lipid A from both Escherichia coli and Salmonella spp. These results have been confirmed by using a fluid-phase antigen-antibody competitive inhibition assay with purified lipid A and an antibody-antibody competitive inhibition assay with a monoclonal antibody with known specificity for lipid A. The HA-1A monoclonal antibody has also been shown to bind to a panel of R-chemotype LPS by ELISA and, unlike many other previously reported anti-lipid A antibodies, binding of RA-1A to R-chemotype LPS and lipid A is comparable. Although binding of HA-1A to S-LPS (smooth, wild-type LPS) could not be detected by direct ELISA, competitive inhibition experiments with some preparations of S-LPS have been able to show specific HA-IA binding. Collectively, these data confirm the binding specificity of RA-1A for the lipid A component of LPS and provide evidence that this monoclonal antibody manifests a relatively uncommon profile in its capacity to bind lipid A and R-chemotype LPS as well as some preparations of S-LPS.