Immunochemical Analysis of Lipopolysaccharides with 2-D Gel Electrophoresis and Monoclonal Antibodies
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Phenol extracted, alkali treated lipopolysaccharide from vaccine strain (S19) Brucella abortus was demonstrated by twodimensional gel electrophoresis to have at least ten silvers taining analogues. When tested on nitrocellulose immune blots all ten were antigenically reactive with bovine anti-B. abortus polyclonal sera, but only six reacted with murine monoclonal anti-O-antigen antibody. Analogues of LPS focusing at distinct pIs were polydisperse and in several cases polyionic, suggesting the incorporation of groups in the 0-antigen side chain which modified the pl. Analogues focusing at different pIs were concluded to arise from differences in either core or O-antigen side chain structure. While no qualitative differences were observed, LPS from a pathogenic B. abortus strain (S2308) had lesser amounts of analogues 1,2,5,6, and 8 t ha n S19 when examined by 2-D Gel.
The 2-D Gel method has proven to be valuable in the study of LPS from E . coli and B. abortus and should be useful for study of LPS from a wide range of sources. Given molecules with discrete isoelectric points and affinities for sodium do decylsulfate or other ionic detergents, the possibility exists that other large biopolymers may be studied with the 2D method as well. Used in conjunction with electroblotting and monoclonal antibody techniques, complex mixtures or fragments of purified materials may be readily separated and studied. It should also be possible to gene rate monoclonal or polyclonal antibodies to selected LPS subfractions separated by 2-D gel by the use of methods such as those reported by Boulard, Lecroisey, and Tracey, et al. and discussed by Pearson and Anderson using LPS fractions as well as protein immunogens contained i n exci sed acrylamide gel patterns 815.
KeywordsVaccine Strain Protein Immunogen Immunochemical Analysis Brucella Abortus Ionic Detergent
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