title: Characterization of Antibodies to Products of Proinsulin Processing Using Immunofluorescence Staining of Pancreas in Multiple Species.
abstract: The efficient processing of proinsulin into mature insulin and C-peptide is often compromised under conditions of beta cell stress, including diabetes. Impaired proinsulin processing has been challenging to examine by immunofluorescence staining in pancreas tissue because the characterization of antibodies specific for proinsulin, proinsulin intermediates, processed insulin and C-peptide has been limited. This study aimed to identify and characterize antibodies that can be used to detect products of proinsulin processing by immunofluorescence staining in pancreata from different species (mice, rats, dog, pig and human). We took advantage of several knockout mouse lines that lack either an enzyme involved in proinsulin processing or an insulin gene. Briefly, we report antibodies that are specific for several proinsulin processing products, including: a) insulin or proinsulin that has been appropriately processed at the B-C junction; b) proinsulin with a non-processed B-C junction; c) proinsulin with a non-processed A-C junction; d) rodent-specific C-peptide 1; e) rodent-specific C-peptide 2; and f) human-specific C-peptide or proinsulin. In addition, we also describe two 'pan-insulin' antibodies that react with all forms of insulin and proinsulin intermediates, regardless of the species. These antibodies are valuable tools for studying proinsulin processing by immunofluorescence staining and distinguishing between proinsulin products in different species.
authors.affiliation: Laboratory of Molecular and Cellular Medicine, Department of Cellular & Physiological Sciences, Life Sciences Institute, University of British Columbia, Vancouver, BC, Canada (AA, JEB, TJK)
authors.forename: Ali
authors.initials: A
authors.lastname: Asadi
authors.affiliation: Laboratory of Molecular and Cellular Medicine, Department of Cellular & Physiological Sciences, Life Sciences Institute, University of British Columbia, Vancouver, BC, Canada (AA, JEB, TJK)
authors.forename: Jennifer E
authors.initials: JE
authors.lastname: Bruin
authors.affiliation: Laboratory of Molecular and Cellular Medicine, Department of Cellular & Physiological Sciences, Life Sciences Institute, University of British Columbia, Vancouver, BC, Canada (AA, JEB, TJK)|Department of Surgery, University of British Columbia, Vancouver, BC, Canada (TJK)
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Kieffer
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology*; D004285:Dogs; D005455:Fluorescent Antibody Technique; D006801:Humans; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D010179:Pancreas / Q000276:immunology* / Q000378:metabolism; D011384:Proinsulin / Q000172:deficiency / Q000276:immunology* / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D013045:Species Specificity; D013194:Staining and Labeling; D013552:Swine
chemical_list: D000906:Antibodies; D011384:Proinsulin
keywords: beta cells; diabetes; immunofluorescence staining; islets; proinsulin processing