Calcium measurements - optimising loading conditions when using Calcium Dyes (Dec/12/2006 )

Hi

I was wondering if anyone can help with a couple of questions.

I am making Ca2+ measurements in CHO cells using Fluo-4 AM dye.
The protocol that I am using involves adding Pluronic acid F127 to improve the loading conditions.

My questions is: How does Pluronic acid F127 enhance the loading?

Also some other colleagues of mine have used a mixture of Pluronic acid and BSA, how would the BSA affect the loading conditions?

Looking forward to hearing from people!

Thanks,



Madu

Advantages of the use of pluronic acid F-127 (a non ionic surfactant polyol) are in the speed of the Fluo-4 AM (AcetoxyMethyl Ester) entry, Intracellular trapping of Fluo-4, eliminating any possible hydrolysis of Fluo-4 AM by external esterases, and more efficient loading while maintaining cell integrity.