Skip to main content
Protein Science : A Publication of the Protein Society logoLink to Protein Science : A Publication of the Protein Society
. 1992 Jul;1(7):935–944. doi: 10.1002/pro.5560010712

Analysis of peptide mixtures by capillary high performance liquid chromatography: a practical guide to small-scale separations.

M T Davis 1, T D Lee 1
PMCID: PMC2142154  PMID: 1338976

Abstract

Capillary HPLC is a very effective means of separating small amounts of peptides and proteins. Capillary columns ranging from 0.01 mm to 0.5 mm in diameter can be constructed using recycled supports and inexpensive fused silica capillary tubing. Commercial pumping systems and UV detectors can be readily converted for operation in the flow rate range of 0.5-50 microL/min. Detailed procedures are given for the construction of columns and UV detector flow cells. A mixture of peptides derived from the endo Lys C digest of horse heart cytochrome c was used to illustrate various aspects of capillary chromatography of peptides and compares the performance of various-sized capillary columns and UV detector flow cell types.

Full Text

The Full Text of this article is available as a PDF (2.1 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Cobb K. A., Novotny M. High-sensitivity peptide mapping by capillary zone electrophoresis and microcolumn liquid chromatography, using immobilized trypsin for protein digestion. Anal Chem. 1989 Oct 15;61(20):2226–2231. doi: 10.1021/ac00195a003. [DOI] [PubMed] [Google Scholar]
  2. Coutant J. E., Chen T. M., Ackermann B. L. Interfacing microbore and capillary liquid chromatography to continuous-flow fast atom bombardment mass spectrometry for the analysis of glycopeptides. J Chromatogr. 1990 Aug 3;529(2):265–275. doi: 10.1016/s0378-4347(00)83833-5. [DOI] [PubMed] [Google Scholar]
  3. Hemling M. E., Roberts G. D., Johnson W., Carr S. A., Covey T. R. Analysis of proteins and glycoproteins at the picomole level by on-line coupling of microbore high-performance liquid chromatography with flow fast atom bombardment and electrospray mass spectrometry: a comparative evaluation. Biomed Environ Mass Spectrom. 1990 Nov;19(11):677–691. doi: 10.1002/bms.1200191107. [DOI] [PubMed] [Google Scholar]
  4. Kassel D. B., Williams K. P., Musselman B. D., Smith J. A. Optimization of the fragmentation in a frit-fast atom bombardment ion source for the sequencing of peptides at the picomole level. Anal Chem. 1991 Sep 15;63(18):1978–1983. doi: 10.1021/ac00018a015. [DOI] [PubMed] [Google Scholar]
  5. Kirkland J. J., Yau W. W., Stoklosa H. J., Dilks C. H., Jr Sampling and extra-column effects in high-performance liquid chromatography; influence of peak skew on plate count calculations. J Chromatogr Sci. 1977 Aug;15(8):303–316. doi: 10.1093/chromsci/15.8.303. [DOI] [PubMed] [Google Scholar]
  6. Lee T. D., Shively J. E. Enzymatic and chemical digestion of proteins for mass spectrometry. Methods Enzymol. 1990;193:361–374. doi: 10.1016/0076-6879(90)93427-m. [DOI] [PubMed] [Google Scholar]
  7. Novotny M. Recent advances in microcolumn liquid chromatography. Anal Chem. 1988 Apr 15;60(8):500A–510A. doi: 10.1021/ac00159a001. [DOI] [PubMed] [Google Scholar]

Articles from Protein Science : A Publication of the Protein Society are provided here courtesy of The Protein Society

RESOURCES