PMID: 7540809Apr 10, 1995Paper

Preparation of anti-mucin polypeptide antisera to study mucin biosynthesis

Analytical Biochemistry
K M TytgatJ Dekker

Abstract

Mucins are very heavily O-glycosylated glycoproteins. For in depth studies on the cell biological aspects of mucins, anti-polypeptide antibodies are essential. We therefore developed a method for the preparation and screening of polyclonal antisera against mucin peptide epitopes. Mucins from five different tissues were isolated using CsCl/guanidinium.HCl density gradient centrifugation, and polyclonal antisera were prepared. Specificity for mucin peptide epitopes was determined by Western blotting, immunohistochemistry, and immunoprecipitation. The versatility of each anti-mucin antiserum for the study of mucin biosynthesis was tested in metabolic labeling experiments on tissue explants. All polyclonal antisera were directed primarily against peptide epitopes of mucin precursors as well as of fully glycosylated mucins. Each of the polyclonal antisera enabled us to study the mucin biosynthesis in the organ where the mucin was isolated from originally. Our mucin isolation method yields very pure mucins with sufficiently intact polypeptides to reproducibly elicit polyclonal anti-polypeptide antisera. As the sera recognized the polypeptides, primarily independent of the state of O-glycosylation, the intermediate steps in the biosyn...Continue Reading

Citations

Oct 29, 2000·Diseases of the Colon and Rectum·H P MeijerA W Einerhand
Oct 11, 2003·Helicobacter·Jeroen H B Van de BovenkampJan Dekker
Nov 6, 2002·The Journal of Histochemistry and Cytochemistry : Official Journal of the Histochemistry Society·Melissa VerburgJan Dekker
May 13, 2010·European Journal of Gastroenterology & Hepatology·Henrike M HamerRobert-Jan M Brummer
Mar 5, 2011·Pediatric Research·Patrycja J PuimanIngrid B Renes
Jun 18, 2002·American Journal of Physiology. Gastrointestinal and Liver Physiology·Ingrid B RenesAlexandra W C Einerhand
Oct 29, 2000·American Journal of Physiology. Gastrointestinal and Liver Physiology·M VerburgJ Dekker
Jan 24, 2002·American Journal of Physiology. Gastrointestinal and Liver Physiology·Ingrid B RenesAlexandra W C Einerhand
Jan 14, 1999·The American Journal of Physiology·B J van KlinkenJ Dekker
Jun 5, 1998·The American Journal of Physiology·B J van KlinkenA W Einerhand
Dec 29, 1998·Analytical Biochemistry·B J Van KlinkenJ Dekker

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