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Can Oligofectamine be used in this way? - (Oct/10/2009 )

I use Oligofectamine to transfect siRNA in 24 wells HeLa cells. In the application, I did some calculations and make a scale-up proportion. For example, it is suggested to dilute 3uL oligofectamine in 12uL Opti-MEM, while I mix 60uL oligofectamine in 240uL Opti-MEM and incubate for 5 minutes.

Once I performed this kind of mixture, I observed a big turbid white clump formed in the mixture (it looked alike fungus floating in media, you know. Please imagine :P ). Well, after 5 minutes incubation time, i re-suspend the mixture and mix with different siRNA. The transfection was not successful. No silencing was observed. This was repeated twice with same results. Alternatively, I have performed single mixture (3uL oligofectamine + 12uL Opti-MEM) for each and every reaction and they worked fine.

Does anybody have experience with this? I'm new to RNAi in my institute and nobody can be consulted :(


Well...after two and half months, this problem remained. I have purchased new Oligofectamine, try to dilute in DMEM (w/o serum) instead of Opti-MEM but the precipitate persisted. The only things I did not change are the pipette tips and eppendorf tube for mixing :rolleyes:

Just want to get reponse from my dear forumer, does anyone observe such condition when using Oligofectamine? Or nobody encountered this problem before?

If I would like to change my transfection reagent, is there any recommendation? I'm transfecting siRNA into HeLa cells.

Thank you!