Vacuoles/Empty spaces in Endothelial cells

Dave Bates dobates at
Mon Mar 27 15:21:06 EST 1995

In article <3kqav6$of9 at>
djack at (Dowdy Jackson) writes:

> In article <Pine.OSF.3.91.950319022615.20687A-100000 at>,
> Jason at UofSask <jir130 at> wrote:
> >I'm currently culturing bovine brain endothelial cells.  I've noticed 
> >that in some of the cells there appears to be what looks like vacuoles or 
> >empty spaces (ranging from small ones to some that are about 1/4 the size 
> >of the cell) in the cytoplasm of the cells, usually near the nucleus.  
> >The cells don't seem to be dying as they are able to reach confluence.  
> >Does anyone have any ideas as to what may be causing the formation of 
> >these vacuoles/spaces and how can it be eliminated from the culture?  
> >Could the cause be the serum (calf serum) or the basic fibroblast 
> >growth factor (human, recombinant) being used?
> >
> >TIA for any help/advice or comments given.
> >
> >Jason Ram
> >
> We have seen the same things in our bovine endothelial cells but this usuall
> happens because of :
> a) Cells are close to the max passage number.
> b) The serum concentration or mitogen concentrations are too low.
> c) The CO2 in the incubators is incorrect.
> We use Endothelial Cell Growth Mitogen from Biomedical Technologies as our
> mitogen becasue it works well and is cheaper than using fibroblast growth
> factor.
> Dowdy Jackson
> Department of Biochemistry, Molecular Biology and Cell Biology

I see this from time to time too. I would agree with Dowdy, especially
on the CO2 and the passage number. Also, be careful about your pH. If
the media is too alkaline then they get vacuolar (also caused by low
CO2 or by the tops of the flasks being airtight). Also make sure you
put fresh L-Glu in your media at least every couple of days. I use
fetal bovine serum not Newborn Calf. They like this better and I use it
at 10% (although others say 7% is fine). Keep the passage number below
20 if you can. For important experiments use 12 or less. A lot of stuff
changes in these cells as you passage them - from 2 or 3 upwards. There
was a poster on von Wilebrand Factor expression at ASCB last year which
said that it is produced at a lower rate after passage 6.

Dave Bates PhD                                     Tel: 916 752-7081
Postdoctoral Researcher                            Fax: 916 758-2554 
Dept of Human Physiology                         email:
dobates at
University of California at Davis                drink: anything
Davis, California 95616, USA            No I don't have a sense of
                                                now buy me a beer      

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