High performance capillary electrophoresis - T.E.A.M.

Instrumentation/Operation
Calculation of mobility or use of an internal standard can
greatly improve relative migration time reproducibility.
Since mobility is calculated relative to the EOF, it is in
effect an internal standard. Variations in temperature due to
changes in ambient or from Joule heating, interaction of the
running buffer with the surface (that is, equilibration), or by
slight changes in buffer composition or concentration can
alter EOF but generally do not affect selectivity. In these
cases an internal standard can improve reproducibility, as
exemplified in table 18 for a peptide mapping separation.
The internal standard can either be added to the sample
mixture or can be a component in the original sample. The
EOF, often present as a negative-going peak due to displacement
of buffer, can be a useful reference for calculating
mobility or for use as an internal standard.
Migration time [min]
Relative migration
Run# Peak 1 Peak 2 Peak 3 Peak 3 / Peak 1
Table 18
Migration time reproducibility
— absolute and internal standard
1 15,193 17.965 18.421 1.212
3 14,971 17.672 18.12 1.210
5 15,107 17.914 18.391 1.217
7 15,303 18.119 18.597 1.215
9 15,237 18.04 18.512 1.215
11 14,713 17.337 17.79 1.209
13 14,575 17.145 17.58 1.206
15 14,401 16.95 17.402 1.208
17 14,522 17.137 17.61 1.213
19 14,869 17.584 18.073 1.215
(n = 20)
Average 14.942 17.654 18.119 1.213
s 0.342 0.45 0.462 0.004
% RSD 2.29 2.55 2.55 0.32
4.3 Detection
Detection in CE is a significant challenge as a result of the
small dimensions of the capillary. Although CE requires only
nanoliter volumes of sample, it is not a “trace” analysis
technique since relatively concentrated analyte solutions or
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