Adhesion Testing: HUVEC and Fibroblasts
Adhesions tests are being performed to see if cellular attachment varies between our material, glass, and plastic. These experiments are motivated by lower initial cell densities observed on pnc-Si in growth tests; we are investigating if the lower initial densities are due to the cells rolling of the edge of the raised pnc-Si substrate or decreased affinity of the cells to our material.
Adhesion tests comparing the number of cells in four conditions:
-Glass with complete media +10% FBS
-pnc-Si with complete media +10% FBS
-Tissue Culture Plastic with complete media +10% FBS
-Tissue Culture Plastic Glass with complete media and no additional FBS
Cells are passaged into cloning rings attached to the various materials with silicone vacuum grease. After most cells had attached the cloning rings were removed and the cells imaged using the Cell Tracker dye. Materials are run in triplicate during each trial. The results for the first four tests are shown below.
Thus far there is no confidently distinguishable difference between adhesion to pnc-Si and other materials, though in three of four cases there is a lower cell density on the pnc-Si. I am unsure of how we can consolidate the different results to tighten up the error bars, as we are seeding at different dilutions. Even if we ran three experiments in a row using the same dilution, I feel like that still wouldn’t work as the cells being used will likely vary in number. I suppose we can scale up and run like 6-9 samples for each experiment. Does anyone have any suggestions?
You can normalize to the seeding density.
Remember to ask Jess about ‘teflon’ coverslips. For these studies, teflon will be a great negative control, polylysine coated coverslips would be a great positive control.
Yes we would normalize it with the initial seeding density. We haven’t been doing that till now. We plan to use hemocytometer and check the cell count in the suspension.