Protein Protocols Protein Protocols

Zn 2+ -Reverse Staining 257
agarose matrix. The gel is then rinsed in water for 5–8 min to remove any excess of the
staining reagents from the gel surface. The reverse stained electropherogram is developed
by incubating the gel for 5–8 min in 1% Na 2CO 3 (Fig. 3). Of note, due to poorly understood
factors, agarose gels are not as amenable to reverse staining as polyacrylamide gels.
Patterns of positive and negative bands are often seen in agarose gels.
Acknowledgment
The author is indebted to Drs. L. Castellanos-Serra and E. Hardy (Centre for Genetic
Engineering and Biotechnology (CIGB), Havana, Cuba) for their contributions to the
development of the reverse staining concept, to the CIGB for supporting this work and
to Dr. D. Stuart (Dept. of Biochemistry, University of Alberta, Canada) for useful
discussions. The author is a Fellow of the Canadian Institutes of Health Research and
the Alberta Heritage Foundation for Medical Research, for support and to Dr. Stuart
(Department of Biochemistry, University of Alberta, Canada) for useful comments on
the manuscript.
References
1. Lee, C., Levin, A., and Branton, D. (1987) Copper staining: a five-minute protein stain for
sodium dodecyl sulfate-polyacrylamide gels. Analyt. Biochem. 166, 308–312.
2. Dzandu, J. K., Johnson, J. F., and Wise, G. E. (1988) Sodium dodecyl sulfate-gel electrophoresis:
staining of polypeptides using heavy metal salts. Analyt. Biochem. 174, 157–167.
3. Adams, L. D. and Weaver, K. M. (1990) Detection and recovery of proteins from gels
following zinc chloride staining. Appl. Theor. Electrophor. 1, 279–282.
4. Fernandez-Patron, C. and Castellanos-Serra, L. (1990) Abstract booklet from in Eighth
International Conference on Methods in Protein Sequence Analysis, Kiruna, Sweden,
July 1–6.
5. Fernandez-Patron, C., Castellanos-Serra, L., and Rodriguez P. (1992) Reverse staining of
sodium dodecyl sulfate polyacrylamide gels by imidazole-zinc salts: sensitive detection of
unmodified proteins. Biotechniques 12, 564–573.
6. Fernandez-Patron, C., Calero, M., Collazo, P. R., Garcia, J. R., Madrazo, J., Musacchio,
A., et al. (1995) Protein reverse staining: high-efficiency microanalysis of unmodified proteins
detected on electrophoresis gels. Analyt. Biochem. 224, 203–211.
7. Castellanos-Serra, L. R., Fernandez-Patron, C., Hardy, E., and Huerta, V. (1996) A procedure
for protein elution from reverse-stained polyacrylamide gels applicable at the low
picomole level: an alternative route to the preparation of low abundance proteins for
microanalysis. Electrophoresis 17, 1564–1572.
8. Castellanos-Serra, L. R., Fernandez-Patron, C., Hardy, E., Santana, H., and Huerta, V.
(1997) High yield elution of proteins from sodium dodecyl sulfate-polyacrylamide gels at
the low-picomole level: application to N-terminal sequencing of a scarce protein and to
in-solution biological activity analysis of on-gel renatured proteins. J. Protein Chem. 16,
415–419.
9. Fernandez-Patron, C., Castellanos-Serra, L., Hardy, E., Guerra, M., Estevez, E., Mehl, E.,
and Frank, R. W. (1998) Understanding the mechanism of the zinc-ion stains of
biomacromolecules in electrophoresis gels: generalization of the reverse-staining
technique. Electrophoresis 19, 2398–2406.
10. Bioinorganic Chemistry: Inorganic Elements in the Chemistry of Life (1994) (Kaim W.
and Schwederki, B., eds.), John Wiley & Sons, Chichester New York Brisbane Toronto
Singapore.