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Related Concept Videos

Overview Of Cell Separation And Isolation01:20

Overview Of Cell Separation And Isolation

Cell separation was first achieved in 1964 by S. H. Seal, who separated large tumor cells from the smaller blood cells using filtration. Two years later, Pohl and Hawk performed experiments on how cells respond differently to a nonuniform electric field based on the cell type. Such observations were the inception of cell separation methods, which allow isolating a single cell type from a heterogeneous sample.
Affinity Chromatography01:03

Affinity Chromatography

Affinity chromatography is a powerful technique extensively utilized for separating and purifying specific biomolecules from complex mixtures. It capitalizes on the highly selective binding between an analyte and its counterpart, such as antibody-antigen interactions. The counterpart is immobilized on the stationary phase, forming an affinity column. The stationary phase typically consists of solid support, such as agarose or porous glass beads, immobilizing the affinity ligand. The mobile...
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Capillary Electrophoresis: Applications

Capillary electrophoretic separations offer various modes, each with unique applications. These modes include capillary zone electrophoresis, capillary gel electrophoresis, capillary array electrophoresis, capillary isoelectric focusing, capillary isotachophoresis, micellar electrokinetic chromatography, and capillary electrochromatography.
Capillary zone electrophoresis (CZE) separates ionic components based on their electrophoretic mobility. It has been used to separate proteins, amino acids,...
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Types Of Column Chromatography

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DNA Agarose Gel Electrophoresis

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Related Experiment Video

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Protein Complex Affinity Capture from Cryomilled Mammalian Cells
10:37

Protein Complex Affinity Capture from Cryomilled Mammalian Cells

Published on: December 9, 2016

Cell separation using cryogel-based affinity chromatography.

Ashok Kumar1, Akshay Srivastava

  • 1Department of Biological Sciences and Bioengineering, Indian Institute of Technology Kanpur, Kanpur, India. ashokkum@iitk.ac.in

Nature Protocols
|October 30, 2010
PubMed
Summary
This summary is machine-generated.

This study presents a novel cryogel affinity matrix for rapid, type-specific cell separation. The method efficiently captures and recovers viable mammalian cells using protein A-functionalized matrices in under 30 minutes.

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Area of Science:

  • Biotechnology
  • Chromatography
  • Materials Science

Background:

  • Cell separation relies on specific interactions between cell surface receptors and immobilized ligands.
  • Existing methods can be time-consuming and may impact cell viability.

Purpose of the Study:

  • To develop a generic and efficient cell separation approach using cryogel affinity matrices.
  • To functionalize cryogel matrices with protein A for antibody-mediated cell capture.

Main Methods:

  • Preparation of supermacroporous polyacrylamide and polydimethylacrylamide cryogel monoliths.
  • Functionalization of cryogels with protein A via epoxy derivatization, ethylenediamine, and glutaraldehyde.
  • Antibody-labeled cell capture and recovery through mechanical squeezing of the cryogel matrix.

Main Results:

  • Cryogel matrices exhibit highly interconnected pores (up to 100 μm) facilitating mammalian cell migration.
  • Protein A-immobilized cryogels effectively captured antibody-labeled target cells.
  • High yields of viable cells were recovered by mechanically squeezing the elastic cryogel.

Conclusions:

  • The developed cryogel affinity matrix offers a rapid (<30 min) and efficient method for type-specific cell separation.
  • This generic approach preserves cell viability, making it suitable for various downstream applications.
  • The supermacroporous nature and mechanical properties of the cryogel are key to its performance.