Are there antibodies to differentiate epithelial from fibroblasts cells? - (Jul/04/2007 )

Best fellow Lab Rodents,

I depicture my issue:
We have been given a valuable immortalized cell population from a befreinded lab. This should be a pure strain of BME-UV1 cells.
But We did not recieve a certificate of quality control with the cell line, so we don't know the passage number & we don't know if it's free of contaminations.

We have done 5 passages with this immortalized cell line.
We have the feeling that 2 different cell morphologies are to be viewed under the microscope.
We found some pictures from a former employee who did work with a qc'd vial of those cell line (not from befreinded lab).
The pictures from the former employee shows a monolayer of hexagonal cells.
We see a non uniform monolayer containing 'hexagonal' cells and longitudinal cells with buds (like fibroblasts).
So our hypothesis is that this cell line could very well be contaminated with fibroblast.

So now comes the tricky part: How would you guys differentiate Epithelial (Bovine Mammary) cells from Fibroblast cells (undefined species).

Or putted in different words: what antibodies would only work on fibroblasts not on epithelial cells?

Thanks for all future suggestions,

IronLunatic (who is staying anonymous for the moment)

Ps: In attachment i add a picture of the cells

-IronLunatic-

QUOTE (IronLunatic @ Jul 4 2007, 06:23 AM)

It looks to me that the "fibroblastic " looking cells are epithelial cells at the margins of the cell islands. This is fairly typical of epithelial cells in growth. The marginal cells are "looking for neighbours". When they contact cells in a monolayer the morphology will be revert back to a typical epithelial morphology. Cell lines do however have a finite life span and can change over time...what is the passage number of your cells?
Have your cells ever been mycoplasma tested?

-Rhombus-

Dear Rhombus,

Thanks for your fast reply.
Here are some further answers (and questions) regarding your points:

QUOTE (Rhombus @ Jul 4 2007, 04:42 PM)

It looks to me that the "fibroblastic " looking cells are epithelial cells at the margins of the cell islands.

> A: Hmmm, might indeed be epithelial cells. I attach 2 other pictures from 80-90% confluency.
I got the feeling they are 'cluttered'.
I have the feeling that the 2 morfologies are fighting for the same space.

I don't know if this is related to my problem but they also don't get fully 100% confluent:
There are always some 'cell empty' spots.

QUOTE (Rhombus @ Jul 4 2007, 04:42 PM)

...what is the passage number of your cells?

A: I don't have that information. What's for sure is that this -with SV40 immortalized- cell line have had more than 20 passages.

QUOTE (Rhombus @ Jul 4 2007, 04:42 PM)

Have your cells ever been mycoplasma tested?

A: Actually, no. Could this be related to the strange morphology?

Best regards,

Ironlunatic

-IronLunatic-

Just to say that i 'chewed' on the 'Horrible mycoplasma contamination' topic on this forum.
This topic gives a straight answer to Rhombus third point.
So no need to reply on the third point.

-IronLunatic-

I cannot read out the image;

try as an fibroblast-specific antibody FSP-1

take a look at:

http://antibodybeyond.com/reviews/antigen-f/fsp1.htm

-The Bearer-

I would second Rhombus point of view, as "fibroblastic" cells are in margin of cell islands. Confluent cells seems to adopt the hexagonal phenotype, as the margin's one tend to be more elongated. Picture is hard to read though.

Have you asked for any pictures of the cells from the lab who sent the cells to you?

-Madrius-