Crossed Immunoelectrophoresis and Enzyme-linked Immunosorbent Assay of the Cell-surface Antigens of Bacteroides fragilis

Cousland, Gary; Poxton, Ian R.

doi:10.1099/00221287-130-3-645

Cited by 7 publications

(5 citation statements)

References 9 publications

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“…Kasper et al, 1983) assume that the surface carbohydrates of this Gram-negative bacterium consist simply of lipopolysaccharide (LPS) and 'capsular polysaccharide'. Hofstad (1 98 1) and Cousland & Poxton (1984), however, have demonstrated a complex set of surface carbohydrates. Much of the apparent confusion stems from the difficulty in preparing pure surface components.…”

Section: Introductionmentioning

confidence: 99%

Immunochemistry of the Surface Carbohydrate Antigens of Bacteroides fragilis and Definition of a Common Antigen

Poxton

Brown

1986

Microbiology

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The components extracted by aqueous phenol from whole cells of Bacteroides fragilis were analysed by SDS-PAGE and immunoblotting and shown to consist of a series of strain-specific, cross-reactive and common antigens, Regularly-spaced ladder patterns on silver-stained gels indicated that in most strains the LPS was present as a predominantly smooth type, but with chain lengths of varying molecular mass, ranging within each particular strain from essentially rough forms to long chain-length smooth forms. The rough form of the LPS at the gel front possessed an antigen common to most of the strains investigated. Another antigen, which migrated behind the rough LPS on SDS gels, was common to all strains of the species. The smooth LPS forms and the other high molecular mass components were strain-specific antigens. Previously published methods are not capable of producing pure LPS or capsular polysaccharide for this organism. claimed to have developed methods for the preparation of pure capsular polysaccharide free from LPS, and of LPS free from capsular polysaccharide, protein and nucleic acid. Starting with an aqueous phenol extract of whole cells, capsular polysaccharide was prepared by gel filtration in the presence of a detergent, whereas LPS was prepared by further extraction with a phenol/chloroform/petroleum spirit mixture. The authors suggested that the LPS of B.fragi1i.s is a rough-type molecule and is both chemically and antigenically similar in all the strains investigated.The development of a method for demonstrating the heterogeneity of LPS on polyacrylamide gels (Tsai & Frasch, 1982) has been applied to many species of Gram-negative bacteria. In a preliminary report (Cousland & Poxton, 1983), we showed that B. fragilis possesses a complex mixture of aqueous phenol-extractable surface carbohydrate antigens. A series of closely spaced bands in a ladder pattern on the silver-stained polyacrylamide gel was reminiscent of the pattern produced by the smooth form of LPS from other bacteria. This led us to suggest that B. fragilis possesses smooth type LPS. We also showed that a common antigen ran at the gel front and was associated in some unknown way with the rough form of the molecule.

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Section: Introductionmentioning

confidence: 99%

Immunochemistry of the Surface Carbohydrate Antigens of Bacteroides fragilis and Definition of a Common Antigen

Poxton

Brown

1986

Microbiology

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“…Cross reactivity between lipopolysaccharides isolated from strains of the species Bacteroides fragilis has frequently been observed (Cousland & Poxton 1983;Hofstad 1977;Weintraub et al 1985;Cousland & Poxton 1984). Incidentally also cross reactions between phenol/water extracted antigens of several serotype specific strains of the Bacteroides fragilis group were reported (Hofstad 1977;Meisel-Miko/ajczyk & Ga/kowska 1976;Babb & Communs 1981;Linko-Kettunen et al 1984).…”

Section: Discussionmentioning

confidence: 98%

“…During our biological and immunochemical investigations of cell wall antigens of the Bacteroides fragilis group we were frequently confronted with the poor solubility of the phenol/water extracted antigenic fractions. In the B. fragilis group several polysaccharide surface antigens have been identified: capsular polysaccharide (Kasper 1976a), lipopolysaccharide (Kasper 1976b;Hofstad 1976;Poxton & Brown 1986) and other carbohydrate antigens (Hofstad 1981;Cousland & Poxton 1984;Reid et al 1987). These are all present in the water-soluble phase after phenol/water extraction.…”

Section: Introductionmentioning

confidence: 99%

The effects of deoxycholate and sodium dodecyl sulphate on the serological reactivity of antigens isolated from six Bacteroides reference strains

Beckmann

Meisel-Mikołajczyk²,

Wallenburg

1990

Antonie van Leeuwenhoek

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The detergents sodium dodecyl sulphate (SDS) and sodium deoxycholate (NaD) are frequently used as solvents for macromolecular polysaccharide complexes in immunochemical and serological techniques. The influence of the disaggregating surfactants on the serological reactivity of endotoxins isolated from six serotype specific reference strains of the Bacteroides fragilis group was investigated by comparing haemagglutinating and precipitating reactivities of antigen solutions in phosphate buffered saline (PBS), NaD and SDS. All antigens were phenol/water extracted endotoxins. Solutions of antigens isolated from serotypes A, B, C and D in PBS exhibited mainly serotype specificity and a few well known low-titer cross reactions; solutions in NaD showed additional cross reactivity, which was enhanced by solubilization of the antigens in SDS. In immunoelectrophoresis endotoxins isolated from serotypes A and C and dissolved in NaD or SDS showed additional precipitation lines compared to solutions of the same antigens in PBS. These changes in the serological reactivity are of relevance for investigations where the serological specificity of antigens is in question.

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“…Two of them (2 and 6) reacted with periodic acid-Schiffs reagent but not with Calcofluor, and the same two showed some periodate effect. Cousland & Poxton (1984) used periodate sensitivity to help identify carbohydrate antigens. We assume antigens 2 and 6 to be glycoproteins of the cell wall.…”

Section: Discussionmentioning

confidence: 99%

Immunoelectrophoretic Analysis of Streptococcus agalactiae Serotype Ia Antigens

Somerset

Davis

1986

Microbiology

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Rabbit antibodies to heat-killed whole cells of Streptococcus agalactiae serotype Ia were used to establish an antigen map using Triton X-100 sonicates of homologous cells and crossed immunoelectrophoresis. A total of 11 antigens were identified but the density of immunoprecipitates was varied and only seven could be reliably detected, one of which dominated the immunoprecipitate pattern by its intensity. The antigens were partially characterized by immunological, chemical and cell-location methods. Five of the antigens contained carbohydrate and two of those were sensitive to trypsin and probably represent cellwall compounds. Of the three most prominent antigens, one was surface located and represented the type and shared type antigens (Iabc), one was a cell-wall carbohydrate and very sensitive to periodate, and one was a proteinlcarbohydrate complex which was heat-labile and trypsin sensitive. Group B epitopes were detected in three immunoprecipitates. Cross-reactions between type Ia and other serotypes and streptococci were recorded.

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Crossed Immunoelectrophoresis and Enzyme-linked Immunosorbent Assay of the Cell-surface Antigens of Bacteroides fragilis

Cited by 7 publications

References 9 publications

Immunochemistry of the Surface Carbohydrate Antigens of Bacteroides fragilis and Definition of a Common Antigen

Immunochemistry of the Surface Carbohydrate Antigens of Bacteroides fragilis and Definition of a Common Antigen

The effects of deoxycholate and sodium dodecyl sulphate on the serological reactivity of antigens isolated from six Bacteroides reference strains

Immunoelectrophoretic Analysis of Streptococcus agalactiae Serotype Ia Antigens

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