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| An Improved Method to Measure Bioactivity of the Fusion Protein GGH Based on the Intracellular cAMP Level |
| GENG Yan, REN Yi-lin, XU Zheng-hong, DOU Wen-fang |
| School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China |
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Abstract Objective: To improve a method to determine bioactivity of the fusion protein GGH based on the intracellular cAMP level. Methods: According to GGH is the analog of GLP-1 which can promote islet β cells to produce second messenger cAMP, detecting intracellular cAMP levels by enzyme-linked immunosorbent assay (ELISA) can be used to characterize GGH biological activity. Results: Exenatide or GGH was used to stimulate rat insulinoma cell line RIN m5f to produce cAMP. The optimal cAMP protective agent and drug diluent were 100nmol/L IBMX and DPBS. We found that freezing and thawing cells in liquid nitrogen 2 times led to maximum and stable production of cAMP in the supernatant measured by ELISA. Conclusion: An in vitro high-throughput screening method for detecting the bioactivities of the fusion protein GGH was successfully modified. Using this method, the biological activity of GGH and GLP-1 analogs can be rapidly identified.
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Received: 20 August 2013
Published: 25 November 2013
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