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Open Forum on Methodology

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Western blot detection of GPCR:GFP fusion proteins

16 January 2001

Jay Slack

We have been expressing several GPCR:GFP fusion constructs in HEK293 cells and have been unable to confirm expression of the appropriate fusion protein. Cellular distributions differ for each of the constructs, suggesting that the correct protein is being made. However, we need to confirm that the correct fusion protein is made before proceeding with functional analyses. Any suggestions/input regarding preparation of lysates, cell numbers needed/evidence of lack of reactivity of anti-GFP antibodies to GPCR:GFP fusions would be greatly appreciated.

Thanks,

Jay Slack

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