The next day, when checking my cells (which we grow in spinner flasks with a loose cap) I noticed 2 things: 1. when coloring the cells with trypan blue some of the preps had web-like structures clumping cells and there was a lot of junk around. I emphasise "some" because other times when doing a trypan blue coloration on the same line, the same day, these wouldn't appear. 2. even without coloration I started noticing long filaments. Not many (5-6 for both chambers of the haemocytometer).
I've attached pictures for both cases.
We then decided to bin everything just in case (cells, media, trypan) and restarted a new cell line. However I see the same thing now as before (occasional, isolated filaments). A professor in our lab suggested that they might be cellulose filaments resulting from wiping the haemocytometer with white tissue paper, but I am skeptical. Now I don't know if it's paranoia and if these things were always there (you know how when routine sets in, you tend to blank certain recurring things) or if it's indeed contamination, so I need your suggestions.
Other, maybe relevant, observations:
> The presence of these things doesn't seem to influence cell viability, just cell growth. Cell growth has slowed down considerably and they hover around 1x106 for long periods (days).
> When using trypan blue the filaments stain. I've read about fungi and it seems that for most species the outer wall is permeable to trypan blue.
> The number of filaments doesn't seem to increase at a high rate, if at all. Nothing else indicates contamination (culture smells as usual, nothing visible floating around, even after 5-6 days).
> I'm using pen-strep, but not fungizone.
> When looking at trypan blue alone, I could see a lot of junk and debris, probably because it's not filtered, so it is possible that the web-like thing around the cells could be due to filthy trypan (we're in the process of recieving new dye soon, which we will filter). Also when looking at just the surface of the haemocytometer after ethanol-wiping it, I can sometimes see 1-2 filaments so the cellulose from tissue theory might be correct.
Are you guys seeing any of this? And if not, does it look like fungus? Thank you for your help.
Edited by Jull, 28 February 2011 - 04:32 AM.