Réf. : platelet
MailingList/general/Biocytex at biocytex.fr
Thu Jun 3 13:39:45 EST 2004
We often "play" with human platelets . Please find a few quick suggestions
1/ Try to gate on dual scatter as usual. Check the purity of the platelet
cloud using back-gating i.e. gate dual scatter plot from CD41b positive
events. Also check that you have > 90% of CD41b + in your dual scatter
gate. If you can not achieve that and you need to analyse low level
markers, then you have to turn to dual color fluorescence.
2/ You may be interested to compare this analysis with many others which
will follow, and potentially to other values already reported in the
litterature. In that case, the best would be to report the mean number of
specific Antibody molecules Bound per Cell (sABC) i.e. ABC corrected from
apparent ABC of the isotype- and concentration-matched irrelevant control
MAb. ABC can be derived from MFI (GeoMean) using appropriate calibrators .
If you are working on human platelets, PLATELET Calibrator is the tool of
choice (see www.biocytex.fr).
3/ Best is to avoid as far as possible aggregated platelets : no Ca2+ in
buffer, no wash step, gentle staining procedure, no temperature shock e.g.
maintain blood sample and staining tubes at room temperature, avoid the
cold, avoid activators red cell. hemolysis, dilute blood or PRP sample
before staining (e.g. initially 1:4 dilution for whole blood, 1:8 for PRP;
you may then stain 20 µL diluted blood (or PRP) with 20 µL MAb and then 20
µL anti-mouse IgG-FITC reagent and finally dilute to 2 mL before FCM
Hope that helps.
In case you need more detailed infos, do not hesitate to communicate
directly with us.
Philippe Poncelet, PhD
Director, Research and Technology
140, Chemin de l'Armée d'Afrique
13010 Marseille France
Tel: +33 (0) 4 96 12 20 40
Fax: +33 (0) 4 91 47 24 71
P.S. Take the opportunity to thank all participants of ISAC for this really
exciting meeting. Hope you all have appreciated the sunny south of France.
"Elham Ashouri" <ashourie at pearl.sums.ac.ir> le 02/06/2004 08:12:36
Veuillez répondre à ashourie at sums.ac.ir
Pour : Cytometry Mailing List <cytometry at flowcyt.cyto.purdue.edu>
Objet : platelet
I try to work with platelet . I am going to run platelet sample.
Some questions about it:
1.if I have low platelet number then can I analysis it as usual?
how can I find correct gate?
2.how can I report the intensity of CD41b(percent, MFI)?
3.if the sample continuing jaint or aggregate platelet how can I
Looking forward to hearing from you soon
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