how-to-count-cells

kresten no-sp at m.org
Wed Aug 8 03:10:25 EST 2001


For some cell types you can also measure acidic phosphatase activity. This
is a very easy assay:

Add pNPP in tritonx100 and acetate buffer to the cells, incubate e.g. 30 min
(eg on a shaker), stop reaction (and convert the nitrophenol to the
phenolate) by NaOH addition, spin to remove any bubles formed by triton,
measure the yellow colour. Very easy. Works well. Reproducible.

There is a paper describing this:

Yang TT, Sinai P, Kain SR
An acid phosphatase assay for quantifying the growth of adherent and
nonadherent cells.
Anal Biochem 1996 Oct 1;241(1):103-8
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_ui
ds=8921171&dopt=Abstract

In my experience you may observe "edge-effects", that is the outer row in
the microtitre (96-well) plate will grow differently and show higher
variance than inner-wells. Therefore it is a good idea to make an outer row
of water or medium. The later can of course be used to measure background -
this is generally very low. Remember to give your cells a bit of new medium
once in a while :-)

Good luck,
Kresten

"J. Arturo García-Horsman" <arturo.garcia-horsman at uku.fi> wrote in message
news:3B700261.EDBA0549 at uku.fi...
> Hello all,
>
> I was wondering if anybody knows a method to determine the number cells
> growing in a 96 well plate. Something like measuring the turbidity
> (absorbance) or refringence, other than actually counting them. I don't
> have access to a cell counter, and I don't want to spend hours counting
> them below the microscope well by well.
>
> Any help would be appreciated.
>
> Cheers,
>
> Arturo.
> _______________________________________________________________
>
> Arturo Garcia-Horsman, Ph.D.
> Research Specialist
> Department of Pharmacology and Toxicology
> University of Kuopio
> Canthia, Harjulantie 1 A, 1. krs.
> 70210 Kuopio, Finland
> Tel: +358-017-162422
> Fax: +358-017-162424
> E-mail: Arturo.Garcia-Horsman at uku.fi
>
>
>





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