I've been using glass coverslips to minimize the volume of solutions that I need for washes, etc of my PDMS chips with combed DNA. Since I began this modification, I've noticed that the DNA "disappears" from my PDMS during the protocol. How likely is it that the DNA is being pulled off the PDMS and onto the surface of the glass coverslip?
If the DNA is immobilized covalently to the PDMS, I think its unlikely. If the DNA only adheres to the PDMS by physisorption I think it's possible. But it should also happen if you use normal microscope slides.
Did you use in your former protocol glass microscope slides?
What is your immobilization strategy for the PDMS?
I do not think the DNA is covalently bound to the PDMS. How does one go about covalently binding DNA to PDMS? For my previous protocol involved soaking the PDMS chip in a significant volume of reaction solutions. the idea behind the glass coverslips were to reduce that volume, but if I'm losing all the DNA it is not going to work.
It sounds like phyisorption is the method for attaching DNA to the PDMS. If that is the case, the rinsing step could provide enough shear stress to remove it. We found this to be the case with blocking non-specific binding in microchannels in some of our work. Do you have some sort of tag for the DNA? The easiest way to test would be to tag the glass slide following the rinse, or maybe do PCR on the flushed fluid from the rinse to get DNA replication.
Unfortunately, I have been using different coverslips during each incubation step to avoid contamination. So I don't have a single source to go back and probe. I've been considering (for a number of reasons) intentionally transferring the DNA to a glass slide. Will it stick more robustly to glass?
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