Demonstration of insulin and glucagon mRNA in routinely fixed and processed pancreatic tissue by in-situ hybridization.
Human insulin and glucagon mRNA were identified in routinely processed pancreatic tissue by non-radioactive in-situ hybridization using digoxigenin-labelled oligonucleotide probes. Cocktails of synthetic oligonucleotides complementary to human insulin and glucagon mRNA were labelled with digoxigenin using terminal deoxynucleotidyl transferase (Tdt). Specific hybrids were detected with alkaline phosphatase-labelled anti-digoxigenin antibody and visualized by BCIP-nitroblue tetrazolium indicator substrate. The results showed highly sensitive and specific staining of islet cells on a range of routinely formalin-fixed and paraffin-embedded tissues. Post-mortem pancreatic tissue from adults and stillborn neonates yielded acceptable signals as long as tissue morphology was well preserved. Preliminary investigations using pancreatic endocrine cell tumours gave clear easily interpretable signals which were comparable to conventional immunostaining. The application of this technique promises to be of value in the investigation of pancreatic disease.
Department of Pathology, University of Leicester, Leicester Royal Infirmary, U.K.
K
K
Shorrock
P
P
Roberts
J H
JH
Pringle
I
I
Lauder
D000328:Adult; D004076:Digoxigenin; D005934:Glucagon / Q000235:genetics*; D006801:Humans; D007231:Infant, Newborn; D007328:Insulin / Q000235:genetics*; D007515:Islets of Langerhans / Q000737:chemistry; D009377:Multiple Endocrine Neoplasia / Q000737:chemistry; D009693:Nucleic Acid Hybridization; D010179:Pancreas / Q000737:chemistry*; D010190:Pancreatic Neoplasms / Q000737:chemistry; D012333:RNA, Messenger / Q000032:analysis*
D007328:Insulin; D012333:RNA, Messenger; D005934:Glucagon; D004076:Digoxigenin