Dye Diffuses through UV-Ozone Treated and Untreated SC267 Samples

In my last post I presented the results of an experiment in which it appeared that BSA did not diffuse through a SC267 sample treated in the UV-Ozone system for 20min at 150C. As a control I also ran BSA dissusion through an untreated SC267 sample and showed that BSA did move across the membrane. Taking a step back, I wanted to ensure that the membrane’s pores were still open by running a dye diffusion experiment over the weekend.

Using Jess’ diffusion cell I ran 4 SC267 samples: 1) untreated+rhodamine, 2) treated+rhodamine, 3) untreated+fluoroscein, and 4) treated+fluoroscein. The hope was that in both the treated and untreated cases we would observe the positively charged rhodamine diffusing across the membrane. At the same time I hoped to show that the negatively charged fluoroscein was able to diffuse across the untreated membrane, but after UV-Ozone treatement the fluoroscein diffusion would be blocked due to the increase in negative charge on the membrane. Unfortunately after analyzing the diffusion results both rhodamine and fluoroscein were able to diffuse across all SC267 samples; treated or untreated.

In the image above the assay area labeled “W267” shows the untreated rhodamine, treated rhodamine, untreated fluoroscein, and treated fluoroscein from left to right.

From this experiment it is shown that the pores in the membrane are still open after UV-Ozone treatment, allowing for the diffusion of both rhodamine and fluoroscein. As for the reason that the treated sample allowed for the fluoroscein to diffusion I am not sure and would appreciate any input. Could it be that the membrane pores are too large to prevent diffusion of a small molecule like fluoroscein with an increase in membrane charge? My last post showed UV-Ozone tratement preventing BSA diffusion, could this be a result that BSA is simply a larger molecule?