A couple months ago, I tried to use tetramethylrhodamine (TAMRA) as a fluid-phase marker to label vacuoles.  The idea is that the only way this stain could be internalized is if it is pinocytosed, eventually ending up in vacuoles.  If…

It’s been a while since I’ve done long-term BBB co-culture experiments on transwells but I recently completed a study.  I was particularly interested to determine whether I could reproduce this spike in TEER that I measured in 1 previous experiment.…

Last time I cultured fibroblasts on pnc-Si transwells, the cells got super-confluent over the membrane and made the analysis difficult.  Basically, I wanted to see if fibroblasts formed vacuoles/2D structures over free-standing membranes.  I thought the multi-layer cells could have…

Along with differently sized square membranes, I also designed a wafer with different sizes of slits.  On each chip, there is a 4000, 2000, 1000 and 500um long window.  The thicknesses of these windows are 200, 100, 50 and 25um. …

In my last post on this subject, I showed that cutting off permeability of pnc-Si chips inhibits vacuole formation in endothelial cells.  To do this, I seeded cells on the membrane side, allowed them to attach, inverted them onto a…

One of the hidden secrets of endothelial angiogenesis assays on Matrigel is that non-endothelial cell types also form 2D capillary-like structures (tubes) on those gels.  Motivated by this observation, I wanted to look at fibroblast behavior on pnc-Si transwells, specifically…

There’s been lots of discussion about “non-porous” membranes and the possibility that they are actually porous.  By “non-porous”, I mean that a TEM of that membrane doesn’t have obvious pores by pore processing/our eyes.  This could mean a couple things:…

The other (see below) just-to-be-sure experiment I performed this week was to compare staining between 2 different live cell dyes.  I typically use calcein AM to stain live cells green.  This is a membrane-permeant dye which, once inside cells, is…

I recently started up another BBB co-cultures and I had a couple of leftover samples.  I decided to use one for a ‘just-to-be-sure’ experiment since I realized that I never did a calcein-Hoechst 33342 co-stain.  That is, I never used…

I designed new masks with different sized squares and slits in order to study how the aspect ratio of free-standing membranes would affect endothelial cells.  In this 1st experiment, I seeded bEnd at 50000 cells/cm2 and let them grow for…