[Cytometry] Apoptotic Progenitor Cells

Donna Cross donna_cross at health.qld.gov.au
Thu Jul 9 02:50:50 EDT 2009


Hi fellow flow'ers
 
We aim is to stain CD34+ cyropreserved progenitor cells in order to quantitate apoptotic cells using Aldeflour (StemCell Technologies) which fluoresces in the green region in combination with TruCount beads.
 
Our current protocol for CD34 quantitation utilises TruCount beads in combination with CD34PE, CD45FITC and 7AAD (single stage assay, ISHAGE gating). We were hoping to eliminate the need for CD45, CD34 and 7AAD.
 
Using the Aldeflour without the TruCount beads works beautifully.
 
The minute the two are used together the assay doesn't work. TruCount beads are hydrophobic. 
 
To eliminate the possibility of quenching, the cells plus Aldeflour were incubated together, then added to the beads. Once again the assay failed to work.
 
>From here:
 
I'd be interested in hearing what experiences people have had with quantitating apoptotic cells.
 
Has anyone managed to get Aldeflour and TruCount beads working in the same tube?
 
Are there any other suggestions for using the Aldeflour to obtain a quantitative value?
 
What other dyes are available for apoptotic staining that could be used in combination with hydrophobic beads?
 
Are there any single colour quantitative beads available on the market that are not hydrophobic?
 
All responses would be greatly appreciated
 
Donna
 
 

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Donna Cross
 
Health Practitioner
Pathology Queensland
Haematology Laboratory
Princess Alexandra Hospital, Ipswich Road, Woolloongabba, Brisbane, 4102
ph:-(07) 3240 5083
fax:-(07) 3240 7042

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