ANTIBODY LABELING WITH REMAZOL BRILLIANT VIOLET 5R, A VINYLSULPHONIC REACTIVE DYE

Abstract

Colloidal gold is the first choice for labeling antibodies to be used in Point Of Care Testing. However, there are some recent reports on a family of textile dyes—named “reactive dyes”—being suitable for protein labeling. In the present article, protein labeling conditions were optimized for Remazol Brilliant Violet 5R, and the sensitivity of the labeled antibodies was assessed and compared with that of colloidal-gold labeled antibodies. Also, the accelerated stability was explored. Optimal conditions were pH 10.95, dye:Ab molar ratio of 264 and an incubation time of 132 min. Labeled antibodies were stable, and could be successfully used in a slot blot assay, detecting as low as 400 ng/mL. Therefore, the present work demonstrates that vinylsulphonic reactive dyes can be successfully used to label antibodies, and are excellent candidates for the construction of a new generation of Point of Care Testing kits.

Keywords:

• antibody
• blot
• dyes
• labeling
• vinylsulphonic

ACKNOWLEDGMENTS

The authors would like to thank Luis Eduardo Santos from Surfactan S.A. for kindly providing the reactive dyes. This work was financially supported by ANPCyT (National Agency for the Promotion of Science and Technology, PICT 2010-1258).

Notes

^§Response is expressed in arbitrary units of OD.

^¥The extent of labeling was calculated as the molar ratio of bound dye and antibody.

*Extent of labeling could not be determined because the absorbance of the solution at 542 nm was negligible.