PVDF for science geeks

Factual Questions
1

I know this is a somewhat technical question, but what the hay!

Why is PVDF used in “low protein binding” syringe filters; but also used extensively in western blotting to bind lots of protein? Are the membranes used in filters somehow different than those used in western blotting? If so, what is the difference chemically?

Well, there you go…Now I’ll sit back and watch this thread sink into oblivion.

2

I can’t tell you why they are used in low protein binding filters, but I can tell you that the to major determinants of protein binding in PDVF filters are hydrophobicity and water-accessibility/wettability. I know that seems like a contradiction, but the two are not incompatible. If water can easily permeate the filter and has good access to every region of the (hydrophobic) filter surface, it will greatly increase the strength of the hydrophobic reactions through exclusionary repulsion. PDVF can be made to score well on both.

I’m sorry, I don’t know how low-protein binding PDVF filters are made, which is probably the heart of your question. I suspect it is application specific. In many applications, the filter could be saturated in a ‘harmless’ protein that will not affect the work being done. With proper buffers (pH, ionic content, etc.) the protein-PDVF bond is remarkably strong and resistant to chaotrophic agents. The ‘factory installed’ proteins won’t dislodge to any appreciable degree, leaving little opportunity for ‘research’ proteins to bind. There were a lot of studies on this in the late 80s/early 90s when I was active in molecular bio, but I wasn’t using these materials so I can’t comment further.

e.g. Tovey ER, Baldo BA. “Protein binding to nitrocellulose, nylon and PVDF membranes in immunoassays and electroblotting.” J Biochem Biophys Methods. 1989 Aug-Sep;19(2-3):169-83.

3

I figured the “non-protein” binding variety would be the “normal” PVDF since it is similar to teflon and touted as a non stick material. I don’t know what is done (if anything) to make it amenable to protein binding.

4

According to Millipore, the low protein binding is accomplished by chemically modifying the surface – usually with polyacrylate, making the surface more hydrophilic. The native hydrophobicity of PVDF facilitates protein binding.

5

It’s been a while since I used PVDF, but

  1. In SDS-PAGE and electroelution, proteins look very different from how they are in nature. They have been denatured and coated with SDS to make their charge/length ratio constant.
  2. PVDF does require a methanol pre-soak and a special buffer for binding.
  3. According to the Millipore product lit, Durapore and the PES filters have been “surface-modified to render them both hydrophilic and low protein binding.”
  4. There can be vastly different properties of such polymeric substances based on the length of the polymer. In filters, it is often about pore size which is usually closely correlated with the polymer lengths. I would assume the pore size on Western blot PVDF is much, much smaller.