Monoclonal antibodies to enterobacterial common antigen and to Escherichia coli lipopolysaccharide outer core: demonstration of an antigenic determinant shared by enterobacterial common antigen and E. coli K5 capsular polysaccharide

Abstract

We established hybridoma cell lines producing monoclonal antibodies against enterobacterial common antigen (ECA) and a substructure of the outer core of different Escherichia coli lipopolysaccharides (LPSs). Anti-ECA antibodies 865 and 898 reacted with ECA in extracts of heated E. coli and with ECA-bound Rl and R4 core-containing LPS preparations, as well as with a purified sample of ECA from Salmonella montevideo. Antibody 865, but not antibody 898, cross-reacted with K5 capsular polysaccharide, suggesting that 4-linked ce-N-acetylglucosamine is part of an antigenic determinant shared by both K5 polysaccharide and ECA. Anti-LPS antibody 786 recognized an outer core structure common to E. coli K-12, B, R2, and R4 core type LPS, but not to Rl and R3 core type LPS. Its most probable target is the trisaccharide sequence Hexp(1->2)-a-D -Glcp(1-*3)a-D-Glcp->(Hepp) (where Hex is hexose, p is phosphate, Glc is glucose, and Hep is heptose), the first glucose being the immunodominant moiety. These monoclonal antibodies may be used not only for the detection of ECA, K5, and LPS core structures but also for analysis of the molecular forms resolved on polyacrylamide gels (banding patterns) of both ECA and LPS, independently of one another.on July 15, 2020 by guest http://iai.asm.org/ Downloaded from

ECA, the enterobacterial common antigen

“…1), indicating a size heterogeneity of chain lengths as demonstrated for smooth lipopolysaccharides [22]. Such patterns were also observed with monoclonal ECA antibodies for crude bacterial extracts [23,24]. From the number of bands ( Fig.…”

“…ECALp s, like ECApo, reacts with monoclonal ECA antibodies in EIA ( Fig. 7; [23,36]). That this is really due to the core-bound ECALPs, and not to a cross-reaction with the R-cores, is indicated by the lack of reactivity of R-LPS from the respective isogenic ECA-negative mutants (1469 and 1327 in Fig.…”

“…This classic way of ECA detection via serological cross-reaction is not needed any longer: antisera against strains with core-bound 'immunogenic' ECA (e.g., E. coli O14, ECA ÷) can be absorbed with isogenic ECA-negative mutants (e.g., E. coli O14, ECA-) to yield monospecific anti-ECA sera. In 1985, two monoclonal anti-ECA antibodies (one IgG, one IgM type) were described [23]. As with polyclonal sera, both monoclonal antibodies showed no more reactivity with ECA upon esterification of the mannosaminuronic acid; the reactivity could be re-established by saponification [11].…”

Monoclonal antibodies and their use in microbiology

“…Thus, at first sight, MCA specific for LPS might appear to have most promise in the recognition of salmonellas. The Enterobacterial Common Antigen to which Peters et al (1985) prepared MCA is probably too widespread amongst Enterobacteriaceae to be suitable for identifying one genus. A variety of other moieties, including outer membrane proteins, occur relatively widely but so far no MCA with specificity for any of them has been found to recognize the genus Salmonella.…”

“…Haernophilus injuenzae type b comes into this category as it causes disease in infants but fails to stimulate protective antibody production until the child is about 18 months old. It frequently causes disease in immunocompromized adults (Petola et al 1977). Antibiotic-resistant strains are being reported relatively frequently (Ward et al 1978).…”