Analysis of rabbit tear fluid using capillary electrophoresis with UV or laser-induced fluorescence detection.

R. J. Varnell, D. Y. Maitchouk, R. W. Beuerman, Michael Francis Salvatore, J. E. Carlton, A. M. Haag

Research output: Contribution to journalArticleResearchpeer-review

3 Citations (Scopus)

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.

Original languageEnglish
Pages (from-to)1-6
Number of pages6
JournalJournal of capillary electrophoresis
Volume4
Issue number1
StatePublished - 1 Jan 1997

Fingerprint

Capillary electrophoresis
Capillary Electrophoresis
Electrophoresis
Tears
Buffers
Lasers
Fluorescence
Rabbits
Borates
Fluids
Sodium Dodecyl Sulfate
Osmolar Concentration
Polyacrylamide Gel Electrophoresis
Sodium dodecyl sulfate
Polyacrylates
Ionic strength
Phosphates
Molecular Probes
Silver Staining
Gels

Cite this

Varnell, R. J. ; Maitchouk, D. Y. ; Beuerman, R. W. ; Salvatore, Michael Francis ; Carlton, J. E. ; Haag, A. M. / Analysis of rabbit tear fluid using capillary electrophoresis with UV or laser-induced fluorescence detection. In: Journal of capillary electrophoresis. 1997 ; Vol. 4, No. 1. pp. 1-6.
@article{9bdba6ca990743dbb167a580155c46d8,
title = "Analysis of rabbit tear fluid using capillary electrophoresis with UV or laser-induced fluorescence detection.",
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.",
author = "Varnell, {R. J.} and Maitchouk, {D. Y.} and Beuerman, {R. W.} and Salvatore, {Michael Francis} and Carlton, {J. E.} and Haag, {A. M.}",
year = "1997",
month = "1",
day = "1",
language = "English",
volume = "4",
pages = "1--6",
journal = "Journal of capillary electrophoresis",
issn = "1079-5383",
publisher = "International Scientific Communications Inc.",
number = "1",

}

Analysis of rabbit tear fluid using capillary electrophoresis with UV or laser-induced fluorescence detection. / Varnell, R. J.; Maitchouk, D. Y.; Beuerman, R. W.; Salvatore, Michael Francis; Carlton, J. E.; Haag, A. M.

In: Journal of capillary electrophoresis, Vol. 4, No. 1, 01.01.1997, p. 1-6.

Research output: Contribution to journalArticleResearchpeer-review

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, Michael Francis

AU - Carlton, J. E.

AU - Haag, A. M.

PY - 1997/1/1

Y1 - 1997/1/1

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.

AB - 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.

UR - http://www.scopus.com/inward/record.url?scp=0030631514&partnerID=8YFLogxK

M3 - Article

VL - 4

SP - 1

EP - 6

JO - Journal of capillary electrophoresis

JF - Journal of capillary electrophoresis

SN - 1079-5383

IS - 1

ER -