kschell at facstaff.wisc.edu
Mon Aug 10 15:00:00 EST 1998
We have an investigator who is looking at fibronectin in asthmatic
patients. Currently she has been able to demonstrate mRNA in lymphocytes,
monocytes, eosinophils, and macrophages from alveolar lavages. In addition
she also finds high levels of fibronectin in the alveolar fluid. She wants
to demonstrate intracellular fibronectin, like intracellular cytokines.
She stains the cells, unfixed, and is able to demonstrate low levels of
fibronectin on the surface of most of the subpopulations in asthmatics (but
not in normals) she looks at. When she fixes and permeabilizes (saponin
and 4% paraformaldehyde) she sees a huge increase in fluorescence in these
populations. Her critics suggest that the only reason she sees this
increase is due to the exposure, by detergent and paraformaldehyde, of
hidden epitopes and that she cannot really demonstate intracellular
fibronectin. Does anyone know if this is true of this technique? Has it
been documented? Any suggestions to identify intracellular fibronectin
versus surface fibronectin with newly exposed epitopes?
Thanks in advance.
Supervisor, UWCCC Flow Cytometry Facility
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