Affinity biotinylation: Nonradioactive method for specific selection and labeling of cellular proteins

Antonio Cosma*

*Corresponding author for this work

Research output: Contribution to journalArticleResearchpeer-review

9 Citations (Scopus)


Biotinylation has become a popular alternative to radioiodination for labeling cell surface proteins, whereas labeling of the total cellular protein pool is usually achieved metabolically with [35S]methionine and [35S]cysteine. In this paper we describe a new technique in which total cellular lysate proteins that have been affinity bound to a solid phase are labeled efficiently with biotin. This labeling technique is preferable to direct biotinylation of cell lysate since the unreacted biotin can be readily removed from the sample by washing. The affinity step permits preselection of the molecules to be labeled, thereby decreasing the potential for nonspecific binding during subsequent immunoprecipitation. We applied this affinity biotinylation method to a human cellular lysate in order to preselect the total glycoprotein pool for subsequent immunoprecipitation of HLA class I. Following immunoprecipitation, SDS-PAGE, and Western blot, the biotinylated protein could be readily revealed by enhanced chemiluminescence. The results were comparable to those obtained by radiometabolic labeling and Western blot using a monoclonal antibody probe. Overall, the affinity biotinylation method is faster and more practical than conventional radiolabeling, without any loss in sensitivity.

Original languageEnglish
Pages (from-to)10-14
Number of pages5
JournalAnalytical Biochemistry
Issue number1
Publication statusPublished - 1 Oct 1997
Externally publishedYes


Dive into the research topics of 'Affinity biotinylation: Nonradioactive method for specific selection and labeling of cellular proteins'. Together they form a unique fingerprint.

Cite this