Entrapment of Proteins Within Columns for High-Performance Affinity Chromatography

Saumen Poddar; Sadia Sharmeen; David S Hage

doi:10.1007/978-1-0716-2176-9_14

Entrapment of Proteins Within Columns for High-Performance Affinity Chromatography

Methods Mol Biol. 2022:2466:205-227. doi: 10.1007/978-1-0716-2176-9_14.

Authors

Saumen Poddar¹, Sadia Sharmeen¹, David S Hage²

Affiliations

¹ Department of Chemistry, University of Nebraska-Lincoln, Lincoln, NE, USA.
² Department of Chemistry, University of Nebraska-Lincoln, Lincoln, NE, USA. dhage1@unl.edu.

PMID: 35585320
DOI: 10.1007/978-1-0716-2176-9_14

Abstract

Entrapment is a noncovalent immobilization method that enables a large biological binding agent, such as a protein, to be put within a support without modifying the structure of the binding agent. This chapter describes an on-column entrapment method that can be used with proteins and HPLC-grade silica to prepare columns for high-performance liquid chromatography. In this method, a protein is trapped within a dihydrazide-activated silica support by using oxidized glycogen as a capping agent. This method allows the protein to be placed within the support in a soluble form and with little or no loss of activity. The approach and reagents needed for this method are described in this chapter, along with some applications reported for columns that have been made using on-column protein entrapment.

Keywords: Entrapment; High performance affinity chromatography; Noncovalent immobilization; Protein immobilization; Silica.

Publication types

Research Support, U.S. Gov't, Non-P.H.S.
Research Support, Non-U.S. Gov't
Research Support, N.I.H., Extramural

MeSH terms

Chromatography, Affinity / methods
Chromatography, High Pressure Liquid / methods
Glycogen / chemistry
Proteins*
Silicon Dioxide* / chemistry

Substances

Proteins
Silicon Dioxide
Glycogen

Grants and funding

R01 DK069629/DK/NIDDK NIH HHS/United States