Particle Separations with Cellulose Membranes
I tried particle separations with the 30kD cellulose membranes from Millipore using the same setup as previous experiments. Basically the results are that neither 5 nor 20nm particles pass through the membranes whether in water or 10mM KCl.
Absorbance Data:
Sizing Data:
No peaks were found in the nm range for any of the filtrates.
We may not be seeing separations because 30kD is too low of a cutoff for a 5nm particle. For reference albumin is about 6nm and 67kD. Using the density formula (D = M/V) and assuming proteins have the same density, we can roughly find that a 30kD protein would have a diameter around 4.6. This means a membrane with a 30kD cutoff would probably have pores smaller than 4.6nm to retain a 30kD protein. 5nm particles then are probably too big, but it is tricky to convert polymer membranes to an absolute pore size because of their characteristics. We are unable to detect our 2nm particles, so the direction to go with this is to pick a higher cutoff membrane.
Update for 100 kD Millipore cellulose membranes:
Ok same deal as before but using Millipore Microcons – a cellulose centrifuge tube insert. Here are the results of the separations:
Again it looks like nothing made it through. It’s interesting, but it seems like the smaller species that are giving off that UV absorbance also had trouble making it through in these membranes.
According to LaTulippe’s seminar:
100kD 6nm pore
300kD 9nm pore
1000kD 12nm pore
Perhaps we want to try 300 or 1000?
Red w/ black spots (closely aggregated gold). Might have dried to some degree on the surface. This doesn’t rinse off.
Back side was perfectly white.
Were the cellulose membranes obviously colored gold after this work? Was it for PES that you took pictures of gold stained membranes?
Yes (I just posted the images). The picture from before was PES – that one was darker and more obvious.