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Production and characterization of monoclonal antibodies against tubulin from intestinal and tissue nematodes (Ascaris suum & Brugia pahangi)

Monoclonal antibodies (MAbs) have been raised against $ beta$-tubulin of B. pahangi and A. suum. Anti-B. pahangi MAbs were used to investigate the heterogeneity of tubulins from nematodes and mammals. One-dimensional SDS-PAGE showed that MAbs P3D and 1B6 react with $ beta$-tubulin from a number of filarial and intestinal nematodes, but not with tubulin from protozoan and mammalian cells. Two-dimensional SDS-PAGE demonstrated that MAb P3D recognizes two isoforms of $ beta$-tubulin and 1B6 recognizes one. Limited proteolysis showed that MAb 1B6 reacted with the amino-terminal fragments and MAb P3D with the carboxyl-terminal fragments of $ beta$-tubulin. The effect of anti-B. pahangi MAbs on the viability of adult B. pahangi was assessed using MTT assay. It was found that MAbs P3D and 1B6 caused an 80% and 40% reduction respectively, in worm viability, whereas anti-chick MAb 357 or mebendazole drug had no effect. Immunogold labelling of B. pahangi demonstrated the presence of tubulin in the median and basal layers of the cuticle, hypodermal layer and somatic muscle blocks, as well as the uterus of B. pahangi. The reduction in the viability of worms may, therefore, be due to the disruption of microtubules in the body wall muscle of B. pahangi. The total MBZ binding was highest in the intestine followed by the body wall muscle and in the reproductive tract extracts of A. suum. Electron microscopy of A. suum tissues demonstrated that the tubulin content decreased from the intestine through the body wall muscle to the reproductive tract. One dimensional SDS-PAGE revealed the presence of $ alpha,$ $ beta sb1$ and $ beta sb2$ tubulin subunits in all tissues of A. suum. This data confirmed the reduction of tubulin from the intestine through the body wall muscle to the reproductive tract. Two dimensional SDS-PAGE followed by Western blotting demonstrated that $ alpha$ and $ beta$ tubulin isoform patterns are dissimilar in different tissues of A. suum. Body wall muscle, inte

Identiferoai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.70284
Date January 1992
CreatorsBughio, Nasreen Inayat
PublisherMcGill University
Source SetsLibrary and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada
LanguageEnglish
Detected LanguageEnglish
TypeElectronic Thesis or Dissertation
Formatapplication/pdf
CoverageDoctor of Philosophy (Institute of Parasitology.)
RightsAll items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated.
Relationalephsysno: 001287122, proquestno: AAINN74615, Theses scanned by UMI/ProQuest.

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