M.G.Kelly@queens-belfast.ac.uk M.G.Kelly at queens-belfast.ac.uk
Fri Jul 6 10:07:41 EST 2001

Hi Greg,
we have been able to perform flow cytometry on induced and spontaneous sputum
from normal subjects, COPD patients and patients with bronchiectasis. It is
feasible, tho' may require a bit of working with. what we do is use a protocol
whereby we incubate the sputum with 0.1% DTT for 15min followed by adding some
PBS. We then centrifuge and resuspend the cells. We label with annexin V and
propidium iodide to look at apoptosis. It works well, and if you would like
some further information, please get in touch with me. Other groups have also
performed flow cytometry on sputum e.g. Hargreave's group at Mc Master
University, Hamilton, Ontario. A Pubmed search should reveal this. they are
also the pioneers of sputum induction and processing.
One thing to remember is that processing with 0.1% DTT may have some effect on
some cell surface receptors and a German group have written on this. Look for a
paper in the Eur Respir J by Loppow et al.
Best wishes.

Dr. Martin Kelly
Department of Respiratory Medicine, Belfast city Hospital, Northern Ireland &
Department of Clinical Biochemistry, Queens University Belfast.
Email: m.g.kelly at qub.ac.uk
Quoting Greg Jasper <gjasper at box-g.nih.gov>:

> greetings flowers!
> Our lab has received a sputum specimen for flow cytometric studies.
> Does anyone have any experience, or suggestions, with processing such
> specimens for flow, especially separating the cells from the sputum.
> Or is this type of sample not recommended for flow?
> Any comments would be helpful.
> regards,
> Greg Jasper, MT
> Flow Cytometry Unit
> NIH, NCI Lab of Pathology

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