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Analysis of rabbit tear fluid using capillary electrophoresis with UV or laser-induced fluorescence detection.
J Capillary Electrophor. 1997 Jan-Feb; 4(1):1-6.JC

Abstract

In preliminary studies of the development of tear analysis methodology that may eventually be useful in the clinical setting, the authors evaluated various protocols for analyzing rabbit tears by capillary zone electrophoresis (CZE). Conditions included the use of a 50-mM monosodium phosphate buffer, pH 2.5, or a 400-mM sodium borate buffer, pH 8.9, both with ultraviolet (UV) detection, as well as a 50-mM borate buffer, pH 8.5, with laser-induced fluorescence (LIF) detection of ATTO-TAG CBQ (Molecular Probes, Inc., Eugene, OR, U.S.A.) derivatized tears. All CZE analyses were performed with a P/ACE System 2100 instrument equipped with System Gold software (Beckman Instruments, Fullerton, CA, U.S.A.), using a 50 microns x 57 cm (50 cm to the window) fused-silica capillary, at 25 degrees C, with constant voltage of 20 kV for UV detection and 11 kV for LIF detection. Tear samples were collected from normal rabbit eyes by means of 10-microL glass micropipets. The volume of each sample was approximately 2 microL. Analysis using the phosphate buffer with UV detection produced as many as 35 peaks in each sample, of which 11 peaks were readily discerned. This compared favorably with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis, which produced 32 bands with silver staining and 11 quantifiable bands with Coomassie brilliant blue staining. Many of the tear protein components have yet to be identified. CZE analysis with the high-ionic-strength borate buffer with UV detection produced only four peaks, and the low-ionic-strength borate buffer with LIF detection produced only six peaks. CZE analysis was completed in less than 1 hr, compared with 7-8 hr for SDS-PAGE. In summary, CZE analysis of tear fluid is comparable to CZE analysis of other bodily fluids and shows great potential for use in clinical diagnosis as well as for enhancing our understanding of the cellular actions of tears on the front of the eye.

Authors+Show Affiliations

Department of Ophthalmology, Louisiana State University Medical Center School of Medicine, New Orleans, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Comparative Study
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

9384713

Citation

Varnell, R J., et al. "Analysis of Rabbit Tear Fluid Using Capillary Electrophoresis With UV or Laser-induced Fluorescence Detection." Journal of Capillary Electrophoresis, vol. 4, no. 1, 1997, pp. 1-6.
Varnell RJ, Maitchouk DY, Beuerman RW, et al. Analysis of rabbit tear fluid using capillary electrophoresis with UV or laser-induced fluorescence detection. J Capillary Electrophor. 1997;4(1):1-6.
Varnell, R. J., Maitchouk, D. Y., Beuerman, R. W., Salvatore, M. F., Carlton, J. E., & Haag, A. M. (1997). Analysis of rabbit tear fluid using capillary electrophoresis with UV or laser-induced fluorescence detection. Journal of Capillary Electrophoresis, 4(1), 1-6.
Varnell RJ, et al. Analysis of Rabbit Tear Fluid Using Capillary Electrophoresis With UV or Laser-induced Fluorescence Detection. J Capillary Electrophor. 1997 Jan-Feb;4(1):1-6. PubMed PMID: 9384713.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Analysis of rabbit tear fluid using capillary electrophoresis with UV or laser-induced fluorescence detection. AU - Varnell,R J, AU - Maitchouk,D Y, AU - Beuerman,R W, AU - Salvatore,M F, AU - Carlton,J E, AU - Haag,A M, PY - 1997/1/1/pubmed PY - 1997/12/31/medline PY - 1997/1/1/entrez SP - 1 EP - 6 JF - Journal of capillary electrophoresis JO - J Capillary Electrophor VL - 4 IS - 1 N2 - In preliminary studies of the development of tear analysis methodology that may eventually be useful in the clinical setting, the authors evaluated various protocols for analyzing rabbit tears by capillary zone electrophoresis (CZE). Conditions included the use of a 50-mM monosodium phosphate buffer, pH 2.5, or a 400-mM sodium borate buffer, pH 8.9, both with ultraviolet (UV) detection, as well as a 50-mM borate buffer, pH 8.5, with laser-induced fluorescence (LIF) detection of ATTO-TAG CBQ (Molecular Probes, Inc., Eugene, OR, U.S.A.) derivatized tears. All CZE analyses were performed with a P/ACE System 2100 instrument equipped with System Gold software (Beckman Instruments, Fullerton, CA, U.S.A.), using a 50 microns x 57 cm (50 cm to the window) fused-silica capillary, at 25 degrees C, with constant voltage of 20 kV for UV detection and 11 kV for LIF detection. Tear samples were collected from normal rabbit eyes by means of 10-microL glass micropipets. The volume of each sample was approximately 2 microL. Analysis using the phosphate buffer with UV detection produced as many as 35 peaks in each sample, of which 11 peaks were readily discerned. This compared favorably with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis, which produced 32 bands with silver staining and 11 quantifiable bands with Coomassie brilliant blue staining. Many of the tear protein components have yet to be identified. CZE analysis with the high-ionic-strength borate buffer with UV detection produced only four peaks, and the low-ionic-strength borate buffer with LIF detection produced only six peaks. CZE analysis was completed in less than 1 hr, compared with 7-8 hr for SDS-PAGE. In summary, CZE analysis of tear fluid is comparable to CZE analysis of other bodily fluids and shows great potential for use in clinical diagnosis as well as for enhancing our understanding of the cellular actions of tears on the front of the eye. SN - 1079-5383 UR - https://www.unboundmedicine.com/medline/citation/9384713/Analysis_of_rabbit_tear_fluid_using_capillary_electrophoresis_with_UV_or_laser_induced_fluorescence_detection_ DB - PRIME DP - Unbound Medicine ER -