Protein staining

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A protein staining includes biochemical methods for staining proteins . It is a method for protein characterization .

properties

Protein staining occurs through binding of a chromophore . The bond can take place via ionic , polar and van der Waals interactions , hydrogen bonds , disulfide bridges , hydrophobic effects or covalently .

By photometry , the protein concentration can e.g. B. by Bradford test , by Lowry test , by biuret reaction , by BCA test or, in the case of higher protein concentrations, also by the absorption of the peptide bond , each method having its own interfering substances which, if present, exclude the use of this method. In the case of one- or two-dimensionally separated proteins, such as in SDS-PAGE or in Western Blot , a photograph or a densitogram is usually made.

Protein noncovalent dyes

Various dyes preferentially bind proteins in a non-covalent manner, e.g. B. Coomassie brilliant blue , silver staining , Fast Green FCF , amido or Ponceau S . Acid proteins can be stained with stains-all .

As a fluorescent dye , for. B. Nile red , brilliant sulfaflavin , 8-anilinonaphthalene-1-sulfonic acid (8-ANS), scopoletin , iridium complexes, trichloroethanol (reacts covalently with tryptophans after UV radiation), SYPRO Orange , SYPRO Red , SYPRO Ruby , SYPRO Tangerine, Flamingo , Krypton , Coomassie Fluor Orange, Lucy 506, Lucy 565, Lucy 569 or Epicocconon ( Lightning Fast , Deep Purple Stain ) are used.

Immunostaining

A protein can be indirectly stained by binding an antibody .

Protein covalent dyes

Typical modifiable groups in proteins are e.g. B. amines (in lysine and arginine ), sulfhydryls ( cysteine ) or phenols ( tyrosine ). Amines are labeled with dabsyl chloride , FITC , TRITC or DABITC , among others .

Applications

Protein dyes are used in biochemistry for protein characterization . In the textile industry, covalently binding protein dyes are mostly used to dye protein-based textiles such as wool or silk , as they are washed out more slowly in washing machines and during dry cleaning .

literature

Individual evidence

  1. ^ A b c C. M. Wilson: Studies and critique of Amido Black 10B, Coomassie Blue R, and Fast Green FCF as stains for proteins after polyacrylamide gel electrophoresis. In: Analytical biochemistry. Volume 96, Number 2, July 1979, pp. 263-278, PMID 89822 .
  2. ↑ CM Wilson: Staining of proteins on gels: comparisons of dyes and procedures. In: Methods in enzymology. Volume 91, 1983, pp. 236-247, PMID 6190068 .
  3. ↑ A. Málnási-Csizmadia, G. Hegyi, F. Tölgyesi, AG Szent-Györgyi, L. Nyitray: Fluorescence measurements detect changes in scallop myosin regulatory domain. In: FEBS Journal . Volume 261, Number 2, April 1999, pp. 452-458, PMID 10215856 .
  4. ^ Y. Chen, J. Yang, Z. Wang, X. Wu, F. Wang: Scopoletine as fluorescence probe for determination of protein. In: Spectrochimica acta. Part A, Molecular and biomolecular spectroscopy. Volume 66, Number 3, March 2007, ISSN  1386-1425 , pp. 686-690, doi: 10.1016 / j.saa.2006.04.012 , PMID 16859971 .
  5. ↑ Junli Jia, Hao Fei, Ming Zhou: Luminescent iridium (III) complexes as novel protein staining agents. In: Electrophoresis . Volume 33, Issue 9-10, 2012, pp. 1397-1401, doi: 10.1002 / elps.201100693 .
  6. ↑ C. Li, M. Yu, Y. Sun, Y. Wu, C. Huang, F. Li: A nonemissive iridium (III) complex that specifically lights-up the nuclei of living cells. In: Journal of the American Chemical Society. Volume 133, Number 29, July 2011, ISSN  1520-5126 , pp. 11231-11239, doi: 10.1021 / ja202344c , PMID 21682270 .
  7. ↑ Carol L. Ladner, Jing Yang, Raymond J. Turner, Robert A. Edwards: Visible fluorescent detection of proteins in polyacrylamide gels without staining. In: Analytical Biochemistry Vol. 326, 2004, pp. 13-20, PMID 14769330 .
  8. ↑ Jennifer E. Gilda, Aldrin V. Gomes: Stain-Free total protein staining is a superior loading control to β-actin for Western blots. In: Analytical Biochemistry Vol. 440, 2013, pp. 186-188, PMID 23747530 .
  9. ↑ a b c d Aleksandr Petrovich Demchenko: Advanced Fluorescence Reporters in Chemistry and Biology III: Applications in Sensing and Imaging Volume 3 of Advanced Fluorescence Reporters in Chemistry and Biology . Springer 2011. ISBN 9783642180354 . P. 179ff.
  10. ↑ Christian P. Moritz, Sabrina X. Marz, Ralph Reiss, Thomas Schulenborg, Eckhard Friauf: Epicocconone staining: a powerful loading control for Western blots. In: Proteomics . PMID 24339236 .