(185b) Isoelectric Focusing without Carrier Ampholytes

Analytical
applications of isoelectric focusing use carrier ampholytes to maintain a
linear pH gradient between the anolyte and catholyte.  There are applications
where use of carrier ampholytes is troublesome, including subsequent sample analysis
by mass spectrometry. 

We
report here on capillary isoelectric focusing when carrier ampholytes are removed. 
This is motivated partially by presentations at last year's AES meeting, where
results were presented and interpreted that differ considerably from ours.  We
report on the nature of the pH gradient measured by the change of speciation,
ultraviolet absorbance and fluorescence of molecular pH probes.  These results
are compared with theory derived from first principles.  Finally we show how single
proteins and simple protein mixtures focus in such a carrier-ampholyte-free
system using imaging capillary isoelectric focusing.

This
text is added to conform with the stated requirement on the minimum word count
of an abstract that will be accepted by the American Institute of Chemical
Engineers and the American Electrophoresis Society.