Does anyone have a complete protocol for an insulin secretion assay in a MIN-6 cell line? I'd like to test the effect of our drug on insulin secretion and am still learning about cell culture. Also, is it possible to measure insulin secretion and to be able to use the very same cells to perform western blot or RT-PCR (to try to probe for the mechanism influencing the secretion)?
Thank you so muc
I have done insulin secretion assays in isolated islets with HEPES-balanced KRB buffer. If you want I can give the protocol for that. Generally we use only 10-20 size-matched islets. At the end of the low and high glucose treatment, we use the islets for 1) measuring the insulin content or 2) protein content. Once I demolish the islets by ultra-sonicator for insulin or protein content I also measure the amount of DNA. Generally I express my data as ng insulin/ug DNA/hour. I thing with small modifications it will be easy to use the same protocol for MIN6 cell line. Good Luck!
Try Henningsson R, Salehi A and Lundquist I. Role of nitric oxide synthase isoforms in glucose-stimulated insulin release. Am J Physiol Cell Physiol. 2002; 283: C296-C304.
Thank you so much, I'll try that paper.
Ercument , it seems like the protocol would be helpful too. Do you mind sending it?
Thanks again
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