Microparticles detection

Nebe-Von-Caron, G g.nebe-von-caron at unipath.com
Fri Mar 30 05:28:01 EDT 2007


Forward scatter is more than optimistic for such an attempt. In many
machines it is not even corellate with the ranking of the particle size
(http://www.phnxflow.com/pdfs/PFS.ISAC.XXI.Poster.pdf
<http://www.phnxflow.com/pdfs/PFS.ISAC.XXI.Poster.pdf> ). 
Sidescatter on cuvette based instruments should go down to 200nm latex
ond 100nm gold ( the latter scattering more). That has nothing to do
with a 100nm microparticle. 1000nm bugs tend to scatter less than 400nm
beads (unless well fed) and in most instruments the particle load from
the sheath will already drown your signals at that level.
I suggest to get some 200nm particles to use as reference and trigger on
side scatter "or" fluorescence to see as many as you can. Remember
degassing solutions and clean sheath tubing.
Regards
Gerhard
P.S. If becomes very important to filter "everything" down to 100nm. and
avoid immunoprecipitation

________________________________

From: Ray Hester [mailto:rhester at jaguar1.usouthal.edu]
Sent: Mon 26/03/2007 21:42
To: cyto-inbox
Subject: Microparticles detection



Hi,

An investigator wants to detect microparticles from endothelial cells
(EMP) that will be labeled with antibodies specific for a variety of
markers including E-selectin and Pecam.  As I understand it, these EMP
are generally from 0.1 to 1 micron in diameter.

We have only a FACSVantage DiVa at our disposal to do these analyses and
don't have much experience detecting particles equal to or smaller than
1.0 micron.  If it's not possible to detect them triggering on FSC, I
know we can try triggering on fluorescence, but there are several papers
published in which FSC is used.  I haven't looked at all of them but the
most recent (SHOCK 26:464, 2006) used an LSRII to detect EMP by light
scatter.  Since the LSR is a cuvette-type rather than stream- in-air
cytometer, I wondered if it would be easier to detect EMP with that type
of flow cytometer, i.e., Aria, Canto, FACSCan, etc.

If someone is using a FCASVantage DiVa to detect particles around 0.5
micron in diameter, will you let us know ?

Thanks.

Ray Hester
Univ of South Alabama






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