Introduction

The Borenpharm PFP-(PEG)n-Biotin is a biotin labeling reagent that reacts with primary and secondary amines. The pentafluorophenyl (PFP) ester-activated biotin is less subject to hydrolysis than NHS esters, resulting in more efficient reactions. PFP-(PEG)n-Biotin must be first dissolved in a minimal amount of an organic solvent, such as dimethylsulfoxide (DMSO) or dimethylformamide (DMF) and then added to the buffer containing the protein or other molecule. The reagent forms an emulsion that allows the reaction to proceed.

Important Product Information

PFP-(PEG)n-Biotin is moisture-sensitive. Store the vial of biotin reagent at 4°C with desiccant. To avoid moisture condensation onto the product, equilibrate vial to room temperature before opening.

As directed in the procedure, dissolve the biotin reagent immediately before use. The PFP moiety readily hydrolyzes and becomes non-reactive; therefore, weigh and dissolve only a small amount of the reagent at a time, and do not prepare stock solutions for storage. Discard any unused reconstituted reagent.

Avoid buffers containing primary amines (e.g., Tris or glycine) as these will compete with the reaction. If necessary, dialyze or desalt to exchange the protein sample into an amine-free buffer such as phosphate-buffered saline (PBS).

When biotinylating proteins in solution, excess nonreacted biotin is easily removed by size exclusion using either desalting columns or dialysis (see Additional Information and Related Thermo Scientific Products). A 10mL desalting column is best suited for processing biotinylation reactions involving 1-10mg of protein in approximately 0.5-2mL. For smaller amounts of protein and/or smaller reaction volumes, both the biotinylation reaction and subsequent buffer exchange may be performed in a single Thermo Scientific Slide-A-Lyzer MINI Dialysis Unit. For larger reaction volumes than can be processed with a desalting column, either split the sample between two columns or use an appropriate Slide-A-Lyzer® Dialysis Cassette.