| Capillary electrophoresis has shown great potential for the separation of biological species. The small dimensions (less than 50 mum) of the capillaries facilitate analysis of minute quantities of analyte. Our instruments use ultra-sensitive laser induced fluorescence detectors to monitor fluorescently labeled molecules as they exit the capillary in a sheath flow cuvette. Primary amines of proteins and biogenic amines are labeled with the fluorogenic reagent 3-(2-furoyl)-quinoline-2-carboxaldehyde (FQ) to produce a highly fluorescent product. The instrumental limit of detection was less than 1 zeptomole (10-21 mol) for FQ labeled protein.; Separations of complex cellular homogenates are performed using capillary zone electrophoresis, micellar electrokinetic capillary chromatography, and capillary sieving electrophoresis as separations modes. One-dimensional separations are optimized to reduced analysis time to less than 2 minutes, while maintaining separation efficiency and peak capacity.; To increase the peak capacity of the system, two separation modes are preformed during two-dimensional capillary electrophoresis (2D-CE). Analysis times of one and two dimensional separations were dramatically reduced through the use of dynamic coatings and narrower inner diameter capillaries. Day to day and within day reproducibility was shown to be better than 2% in both dimensions for 2D-CE separations of BE cellular homogenate.; We apply this sensitive and reproducible technique to analyze tissue biopsies from patients with Barrett's Esophagus. Barrett's Esophagus is a premalignant condition which afflicts those who suffer from gastro-intestinal reflux disease. Barrett's, squamous, and fundus tissues are excised from three patients and analyzed by 2D-CE. Separated components are classified as proteins and biogenic amines, and identified by co-migration with standards. Statistical analysis of 2D-CE spot volumes showed that several amino acids are present at different levels across the different tissue types, and in those patients with the highest risk of progression to cancer. |
