Differential detection of rat islet and brain glutamic acid decarboxylase (GAD) isoforms with sequence-specific peptide antibodies
| Autor: | Linsong Li, Denis G. Baskin, W. A. Hagopian, M. Skelly, Åke Lernmark, A E Karlsen, Jianji Jiang |
|---|---|
| Rok vydání: | 1995 |
| Předmět: |
Gene isoform
endocrine system Histology endocrine system diseases Immunoblotting Glutamate decarboxylase Immunocytochemistry Antibodies Islets of Langerhans Animals Rats Inbred BB Antiserum geography geography.geographical_feature_category biology Glutamate Decarboxylase Brain nutritional and metabolic diseases Islet Immunohistochemistry Rats Isoenzymes Biochemistry Polyclonal antibodies biology.protein Anatomy Peptides Sequence Analysis Immunostaining |
| Zdroj: | Journal of Histochemistry & Cytochemistry. 43:53-59 |
| ISSN: | 1551-5044 0022-1554 |
| DOI: | 10.1177/43.1.7822765 |
| Popis: | We studied the distribution of the M(r) 65,000 and M(r) 67,000 isoforms of glutamic acid decarboxylase, GAD65 and GAD67, in rat islets and brain by immunocytochemistry. Synthetic peptides representing selected GAD65 or GAD67 sequences were used to produce sequence-specific antibodies, allowing differential immunocytochemical detection of the two isoforms. GAD-specific reactivity of each peptide antiserum was confirmed by ELISA, immunoblotting, and immunoprecipitation. Immunostaining specificity was verified by displacement with either immunizing or irrelevant peptide. Dual immunostaining with GAD isoform-specific antibodies and polyclonal antibodies to glucagon showed that GAD65 was primarily detected in rat pancreatic islet beta-cells, whereas alpha-cells had weak GAD65 staining. In contrast, GAD67 was detected primarily in alpha-cells. In rat brain, GAD65 and GAD67 were present in neuron cell bodies and processes. These data demonstrate that antibodies raised against the N-terminus of GAD allow differential immunocytochemical identification of GAD67 and GAD65. Differential expression of GAD isoforms within islet alpha- and beta-cells supports the role of GAD65 in autoimmune diabetes and stiff-man syndrome. |
| Databáze: | OpenAIRE |
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