Sorting Cryptosporidium Oocysts.

From: PETER BERCZ (BERCZ.PETER@EPAMAIL.EPA.GOV)
Date: Thu Jan 30 1997 - 11:05:33 EST


Dear Flow Cytometrists:

Problem Statement:  In  wanting to
determine the minimally infective dose
(oral route in a neonatal mouse model as
well as in a human colon tumor cell line) of
cryptosporidium oocysts, we must obtain
highly accurately sorted and calibrated
pure cyst suspensions from crude density
gradient preps.  Main contaminants in
crude are yeasts.

We have a highly specififc FITC-MAB,
fluor tagging of cysts has been well
worked out.

We have a Coulter Epics Elite  equipped
with the SPE fast sorter and autoclone,
we are experienced sorters. ( We also
have a Meridian ULTIMA confocal scope
as an adjunct tool)

My inquiry is about:

1. Does anyone have experience with flow
sorting and/or confocal mapping of
protozoan oocysts, about potential 
problems and technical tricks, etc?

2. Would FITC-MAB tagging and flow
sorting of oocysts  have an adverse
impact on cyst viability, infectivity
and virulence?

3. Do I need to worry about aerosol-borne
 infectious control.

Any advice or dialogue  will be greatly
appreciated.

J. Peter Bercz
USEPA/NERL/EERD   Cincinnati



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