Analytical and Bioanalytical Chemistry

, Volume 382, Issue 3, pp 783–788 | Cite as

The transitional isoelectric focusing process

Original Paper
First Online:
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Abstract

The transitional isoelectric focusing (IEF) process (the course of pH gradient formation by carrier ampholytes (CAs) and the correlation of the focusing time with CA concentration) were investigated using a whole-column detection capillary isoelectric focusing (CIEF) system. The transitional double-peak phenomenon in IEF was explained as a result of migration of protons from the anodic end and hydroxyl ions from the cathodic end into the separation channel and the higher electric field at both acidic and basic sides of the separation channel. It was observed that focusing times increase logarithmically with CA concentration under a constant applied voltage. The correlation of focusing time with CA concentration was explained by the dependence of the charge-transfer rate on the amount of charged CAs within the separation channel during focusing.

Keywords

Isoelectric focusing Carrier ampholytes Transitional IEF process Focusing time 

Abbreviations

IEF

Isoelectric focusing

CIEF

Capillary isoelectric focusing

CAs

Carrier ampholytes

pI

Isoelectric point

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Notes

Acknowledgements

The authors gratefully acknowledge financial support from the Natural Sciences and Engineering Research Council of Canada (NSERC).

References

  1. 1.
    Righetti PG (1983) Isoelectric focusing: theory, methodology and applications. Elsevier Biomedical, New YorkGoogle Scholar
  2. 2.
    Catsimpoolas N (1976) Isoelectric focusing. Academic, New YorkGoogle Scholar
  3. 3.
    Svensson H (1961) Acta Chem Scan 15:325–341Google Scholar
  4. 4.
    Rodrigez-Diaz R, Zhu M, Wehr T (1997) J Chromatogr A 772:145–160Google Scholar
  5. 5.
    Rodrigez-Diaz R, Wehr T, Zhu M (1997) Electrophoresis 18:2134–2144PubMedGoogle Scholar
  6. 6.
    Catsimpoolas N, Wang J (1971) Anal Biochem 39:141–155PubMedGoogle Scholar
  7. 7.
    Chrambach A, Yotis WW, Griffith AL, Rodbard D (1975) Arch Biochem Biophys 163:113–121Google Scholar
  8. 8.
    Dishon M, Weiss GH (1977) Anal Biochem 81:1–8Google Scholar
  9. 9.
    Mosher RA, Thormann W, Bier M (1986) J Chromatogr 351:31–38Google Scholar
  10. 10.
    Mosher RA, Thormann W (1990) Electrophoresis 11:717–723PubMedGoogle Scholar
  11. 11.
    Mao Q, Pawliszyn J, Thormann W (2000) Anal Chem 72:5493–5502PubMedGoogle Scholar
  12. 12.
    Thormann W, Huang T, Pawliszyn J, Mosher R (2004) Electrophoresis 25:324–337PubMedGoogle Scholar
  13. 13.
    Pospichal J, Deml M, Bocek P (1993) J Chromatogr 638:179–186Google Scholar
  14. 14.
    Pospichal J, Glovinova E (2001) J Chromatogr A 918:195–203PubMedGoogle Scholar
  15. 15.
    Huang T, Wu XZ, Pawliszyn J (2000) Anal Chem 72:4758–4761PubMedGoogle Scholar
  16. 16.
    Hamann CH, Hamnet A, Vielstich W (1998) Electrochemistry. Wiley-VCH, TorontoGoogle Scholar
  17. 17.
    Stoyanov AV, Righetti PG (1998) J Chromatogr A 799:275–282Google Scholar
  18. 18.
    Stoyanov AV, Righetti PG (1998) Electrophoresis 19:2269–2272PubMedGoogle Scholar
  19. 19.
    Crow DR (1988) Principles and applications of electrochemistry. Chapman and Hall, LondonGoogle Scholar
  20. 20.
    Kuhn R, Hoffsterrer-Kuhn S (1993) Capillary electrophoresis: principles and practice. Springer, Berlin Heidelberg New YorkGoogle Scholar
  21. 21.
    Weiberger R (2000) Practical capillary electrophoresis. Academic, New YorkGoogle Scholar
  22. 22.
    Huang T (2003) New carrier-ampholytes-free isoelectric focusing approach to separate proteins. PhD Thesis, University of WaterlooGoogle Scholar

Copyright information

© Springer-Verlag 2004

Authors and Affiliations

  1. 1.The Guelph–Waterloo Centre for Graduate Work in Chemistry, Department of Chemistry (GWC)University of WaterlooWaterlooCanada

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