Laser ablation construction of on-column reagent addition devices for capillary electrophoresis.
A simple and reproducible technique for constructing perfectly aligned gaps in fused-silica capillaries has been developed for postcolumn reagent addition with capillary electrophoresis. This technique uses laser ablation with the second harmonic of a Nd:YAG laser (532 nm) at 13.5 mJ/pulse and a repetition rate of 15 Hz to create these gaps. A capillary is glued to a microscope slide and positioned at the focal point of a cylindrical lens using the focused beam from a laser pointer as a reference. Gaps of 14.0 +/- 2.2 microm (n = 33) at the bore of the capillary are produced with a success rate of 94% by ablation with 400 pulses. This simple method of gap construction requires no micromanipulation under a microscope, hydrofluoric acid etching, or use of column fittings. These structures have been used for reagent addition for postcolumn derivatization with laser-induced fluorescence detection and have been tested for the separation of proteins and amino acids. Detection limits of 6 x 10(-7) and 1 x 10(-8) M have been obtained for glycine and tranferrin, respectively. Separation efficiencies obtained using these gap reactors range from 38,000 to 213,000 theoretical plates.
Rezenom
Yohannes H
YH
Department of Chemistry, University of Tennessee, Knoxville 37996-1600, USA.
Lancaster
Joseph M
JM
Pittman
Jason L
JL
Gilman
S Douglass
SD
D019075:Electrophoresis, Capillary / Q000295:instrumentation* / Q000379:methods; D005998:Glycine / Q000032:analysis; D007202:Indicators and Reagents; D007834:Lasers; D012680:Sensitivity and Specificity; D014168:Transferrin / Q000032:analysis
D007202:Indicators and Reagents; D014168:Transferrin; D005998:Glycine