I am trying to culture and differentiate PC12 cells in an open-faced PDMS microchannel. I have had success differentiating PC12 cells with NGF in culture dishes by coating with poly-L and collagen type IV. However, when I am having problems doing this in the PDMS microchannels. I seed the undifferentiated cells in the channels for 2-3 days prior to adding 100 ng/mL of NGF to the media. The channels are submerged in a dish with DMEM supplemented with 1 % pen/strep, 10 % horse serum and 5 % FBS (culture media) and/or DMEM with 1 % horse serum and 1 % pen/strep (differentiating media). The undifferentiated cells survive in the channels/divide, but there is no growth of dendrites with the NGF is added, even after 7 days. I add 100 ng/mL of NGF and change the media every other day. If anyone has tried something similar and could offer some advice, I would appreciate it!
What are the dimensions of your microchannel? Have you functionalized your PDMS device with ECM? If so, which one(s)?
Thanks for the responses. The width of my channel is 500 microns wide and ~ 50 microns deep. I functionalized my channel with PEG-silane (after activation in a low RF plasma cleaner) to render it hydrophilic prior to incubation with poly-L.
Maybe the absence of ECM is precluding neurite outgrowth. Can you try to functionalize your PDMS with collagen IV or a combination of ECM + PLL instead?
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