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Title: ELECTROLYTE, COLUMN AND DETECTOR SELECTION FOR THE SEPARATION AND ANALYSIS OF METALLOTHIONEIN BY CAPILLARY ELECTROPHORESIS

Author
item BEATTIE JOHN H - THE ROWETT RESEARCH INST
item GRAEDLER FLORIAN - THE ROWETT RESEARCH INST
item POMBO SUSANA - THE ROWETT RESEARCH INST
item Richards, Mark

Submitted to: Capillary Electrophoresis Conference Proceedings
Publication Type: Proceedings
Publication Acceptance Date: 8/26/1994
Publication Date: N/A
Citation: N/A

Interpretive Summary:

Technical Abstract: Several recent studies have demonstrated the use of capillary zone electrophoresis and micellar electrokinetic capillary chromatography for the separation of closely related isoforms of metallothionein (MT), a low Mr metal-inducible and metal-binding protein. Separation of purified MT by these high resolution techniques has in many cases revealed a degree of heterogeneity only previously suspected from the identification of multiple functioning MT genes. Work is therefore in progress to develop a quantitative assay for these different proteins. As part of this work, we are investigating methods to improve the specificity and sensitivity for analysis of MT isoforms in tissue extracts. Untreated silica capillaries plugged with C18 material have been used to concentrate small Mr compounds prior to their elution and separation by capillary electrophoresis techniques. We therefore decided to investigate the potential of C18 and other affinity media for the preconcentration of MT. Cartridges 5 mm in length were made from fine-bore polythene capillary tubing and were fitted to the sampling end of the silica capillary which was installed on a Beckman P/ACE 2050 instrument. Using C18 packing material for example, samples were loaded by pressure injection, eluted with 50% acetonitrile and separated at 30 kV using phosphate buffer as the electrolyte. Some selectivity over contaminants and considerable enhancement in sensitivity was achieved while demonstrating a linear relationship between loading time and peak area for the different isoforms. The method has considerable potential for MT analysis and also for wider applications.