[Cytometry] PI intensity moves during analysis

Ana Leda Longhini analeda.consultoria at gmail.com
Tue May 10 10:00:20 EDT 2011


Lizzy, I would like to take advantage of your mail and add a question. Why
there is a shift between samples? That happens all the time with my
customers, even doing lineage cell cycle.

Thanks in advance,

Ana Leda F. Longhini
Flow Cytometry Lab
Unicamp - SP- Brazil



On Tue, May 10, 2011 at 10:35 AM, Elisabeth Freyer <
Elisabeth.Freyer at hgu.mrc.ac.uk> wrote:

> Hello everybody,
>
> I am observing some strange shifts of the PI intensity when i do cell cycle
> analysis. I'm not talking about the shifts between samples, my whole
> histogram of the same sample actually moves up in intensity within the first
> minute of running it on the machine. After a minute it stays about the same.
>
> I use an Aria2, measure PI in 488-685/35BP
> It ONLY happens when I use protocols that require treating the cells with
> Pepsin or Trypsin (e.g. Vindelov) prior to  PI staining. no movement w/ EtOH
> fixation and staining in PI/RNAse solution only.
> The samples are sitting on ice until they get loaded onto the machine. I
> thought it might be temperature related, but also with a cooled sample
> chamber it still moves.
>
> To record I now usually wait for a minute for the PI intenisty to stabilize
> and record then. It gets a bit difficult when I have samples with very small
> cell numbers :(
>
> I'd greatly appreciate if somebody could enlighten me where that comes from
> and if there is anything I can do to stop this.
>
> Cheers
> Lizzie
> :)
>
> --
> Elisabeth Freyer
> Flow Cytometry Facility
> MRC Human Genetics Unit
> Western General Hospital
> Crewe Road
> Edinburgh
> EH4 2XU
>
> Tel: 0131 332 2471
> email: Elisabeth.Freyer at hgu.mrc.ac.uk
>
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>



-- 
Ana Leda F. Longhini
Consultora Cientifica
Fones: (19) 8169-6696
           (19) 3342-3114
e-mail: analeda.consultoria at gmail.com


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