Hello

I have a severe problem with mdck cells on matrigel.

I incubate them for 48h at 37°C in 5% CO2 and DMEM 5% FCS on the substratum. Afterwards I fix with 4% PFA in order to analyze. Thus far, so good. But after fixing, several rinsing steps are due (and I do it very gently, belief me): I lose my cells while rinsing. I think, they don't attach well in the first place. Yet I figure, that 48h and fixing should be sufficient??? When I look into the microscope after PFA treatment, you can literally see the cells loosing contact and rounding off.

What do I do wrong? I feel like a beginner again, but nobody in my lab seems to come up with an idea:mad:

Thanks :)

I'm trying to organise and locate a source of decent quality and relatively cheap FBS in particular one that is good for the growth of 293s and Hela but in particular FLPin Trex 293s because they seem to be the only ones that didn't grow well when doing a batch comparison between my old batch of FBS and PAAs FBS Gold. Can anyone recommend a particular company?

Dear all,

I got a big problem for my cell culture recently, the S1 (HMT-3552) refuses to behave themselves in 3D culture (in Matrigel). Normally, they stop proliferating (exit from cell cycle) after day7 when I remove the EGF, however, they start to continue growing after day 7 recently. And some lab members doubt that we may get mycoplasma contamination, however, I did not see any changes for the medium (it is not turbid but as clear and transparent as normal).

I wonder whether there is any perceptible signs for mycoplasma contamination that I can use to judge my case?

Many thanks.


Lei

Hi all

Lately, our research group demonstrated the expression of D3 dopamine receptors in a rat Malignant Peripheral Nerve Sheat Tumor (MPNST) cell line. Simply put, schwannoma cells (straightforward, though old nomenclature).

Thus, we would like to measure some parameters following a dopamine treatment. We already have some dopamine hydrochloride (a well-known dopamine salt used in medicine to strenghten heart beat).

Are there any protocol and/or constraints in treating an non-primary, non-neuronal cell line with dopamine hydrochloride? I.e., should we perform some pre-treatment? Should we set some particular medium conditions? Has anybody here ever used dopamine-related molecules in similar situations?

Apparently there is little on Pubmed about in vitro dopamine hydrochloride-exposure.

Thank you in advance

"Bradley K. Sherman" <bks@panix.com> wrote in message
news:hcvggu$s8i$1@reader1.panix.com...
Thanks Bradley,
That's neat, looks so much like an insect walking on that bubble.
Just so I'm clear, that is a single cell and the cilia are part of that
cell.
Many more on the site that I'll check out too.
Thanks,
The Mav