Fast Protein Staining in Sodium Dodecyl Sulfate Polyacrylamide Gel using Counter ion-Dyes, Coomassie Brilliant Blue R-250 and Neutral Red

  • Choi, Jung-Kap (College of Pharmacy and Research Institute of Drug Development, Chonnam National University) ;
  • Yoo, Gyurng-Soo (College of Pharmacy and Research Institute of Drug Development, Chonnam National University)
  • Published : 2002.10.01

Abstract

A fast and sensitive protein staining method in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) using both an acidic dye, Coomassie Brilliant Blue R-250 (CBBR) and a basic dye, Neutral Red (NR) is described. It is based on a counter ion-dye staining technique that employs oppositely charged two dyes to form an ion-pair complex. The selective binding of the free dye molecules to proteins in an acidic solution enhances the staining effect of CBBR on protein bands, and also reduces gel background. It is a rapid staining procedure, involving fixing and staining steps with short destaining that are completed in about 1 h. As the result, it showed two to fourfold increase in sensitivity comparing with CBBR staining. The stained protein bands can be visualized at the same time of staining.

Keywords

References

  1. Bradford, M.M., A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem., 72, 248-254 (1976) https://doi.org/10.1016/0003-2697(76)90527-3
  2. Jung D. W, Yoo, G. S., and Choi, J. K., Mixed-dye staining method for protein detection in polyacrylamide gel electrophoresis using calconcarboxylic acid and rhodamineB. Electrophoresis, 19, 2412-2415 (1998) https://doi.org/10.1002/elps.1150191409
  3. Jung, D. W, Tak, G. H., Lim, J. W, Bae, C. J., Kim, G. Y., Yoo, G. S., and Choi, J. K., Detection of proteins in polyacrylamide gels using eriochrome black T and rhodamineB. Anal. Biochem., 263, 118-120 (1998) https://doi.org/10.1006/abio.1998.2813
  4. Laemmli, U. K., Structural proCleavage of steins during the assembly of the head of bacteriophage T4. Nature, 227, 680-685 (1970) https://doi.org/10.1038/227680a0
  5. Merril C. R., Gel-staining techniques. Methods Enzymol., 182, 477-488 (1990) https://doi.org/10.1016/0076-6879(90)82038-4
  6. Neuhoff, V., Stamm, R., and Eibl, H., Clear background and highly sensitive protein staining with Coomassie blue dyes in polyacrylamide gels: A systematic Analysis. Electrophoresis, 6, 427-448 (1985) https://doi.org/10.1002/elps.1150060905
  7. Neuhoff V, Stamm R, Pardowitz I, Arold N, Ehrhardt W, Taube D., Essential problems in quantification of proteins following colloidal staining with coomassie brilliant blue dyes in polyacrylamide gels, and their solution. Electrophoresis, 11, 101-117 (1990) https://doi.org/10.1002/elps.1150110202
  8. Nivinskas H., and Cole K. D., Environmentally benign staining procedure for electrophoresis gels using Coomassie Brilliant Blue. Biotechniques, 20, 380-385 (1996)
  9. Patton W. F., Detection technologies in proteome analysis. J. Chromatogr B Analyt. Technol. Biomed. Life Sci., 771, 3-31(2002) https://doi.org/10.1016/S1570-0232(02)00043-0
  10. Sreeramulu, G., Singh, N. K, Destaining of Coomassie Brilliant Blue R-250-stained polyacrylamide gels with sodium chloride solutions. Electrophoresis, 16, 362-365 (1995) https://doi.org/10.1002/elps.1150160162
  11. Tal M., Silberstein A, and Nusser E., Why does Coomassie Brilliant Blue R interact differently with different proteins? A partial answer. J Biol Chem., 260(18), 9976-9980 (1985)
  12. Tsubouchi M., Spectrophotometric determination of anions by solvents extraction with neutral red. Anal. Chim. Acta, 54(1) 143-148 (1971) https://doi.org/10.1016/S0003-2670(01)81864-X
  13. Wirth P. J., and Romano A, Staining methods in gel electrophoresis, including the use of multiple detection methods. J. Chromatography, 698, 123-143 (1995) https://doi.org/10.1016/0021-9673(94)00879-E