Separation of high-density lipoproteins into apolipoprotein E-poor and apolipoprotein E-rich subfractions by fast protein liquid chromatography using a heparin affinity column

The available methods for the separation of membrane proteins and lipoproteins are sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS/PAGE), followed by immunoblotting, isoelectric focusing (IEF), capillary electrophoresis (CE) and high‐performance liquid chromatography (HPLC). In this article it is shown that HPLC techniques, given their wide versatility, relative ease of use, and high resolution, may be considered the most valuable tool for the characterization of virtually any hydrophobic protein. Moreover, HPLC is not a destructive technique and therefore proteins, once separated, are available for further analytical investigations. Application examples are described and comparisons with other methods are discussed.

Chromatography of Membrane Proteins and Lipoproteins

Zolla¹

2000

The available methods for the separation of membrane proteins and lipoproteins are sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS/PAGE), followed by immunoblotting, isoelectric focusing (IEF), capillary electrophoresis (CE) and high‐performance liquid chromatography (HPLC). In this article it is shown that HPLC techniques, given their wide versatility, relative ease of use, and high resolution, may be considered the most valuable tool for the characterization of virtually any hydrophobic protein. Moreover, HPLC is not a destructive technique and therefore proteins, once separated, are available for further analytical investigations. Application examples are described and comparisons with other methods are discussed.

Chromatography of Membrane Proteins and Lipoproteins

Zolla

D’Alessandro

2011

The available methods for the separation of membrane proteins and lipoproteins are sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS/PAGE), followed by immunoblotting, isoelectric focusing (IEF), and capillary electrophoresis (CE), along with the recently introduced gel‐based native techniques (blue native (BN) and clear native (CN)), and high‐performance liquid chromatography (HPLC). In this article, it is shown that HPLC techniques, given their wide versatility, relative ease of use, and high resolution, may be considered the most valuable tool for the characterization of virtually any hydrophobic protein. Application examples are described, and comparisons with other methods are discussed. Moreover, HPLC is not a destructive technique, and therefore, proteins, once separated, are available for further analytical investigations. Among these techniques, quantitative and qualitative analyses of the separated fractions can be obtained through other biophysical approaches, such as crystallography or structural spectroscopy. Most of these approaches require preliminary protein purification (90% or higher), which could be rapidly obtained through preliminary HPLC.

Chromatography of Membrane Proteins and Lipoproteins

Zolla

D’Alessandro

Lana

2016

The available methods for the separation of membrane proteins and lipoproteins are sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS/PAGE), followed by immunoblotting, isoelectric focusing (IEF), and capillary electrophoresis (CE), along with the recently introduced gel‐based native techniques (blue native (BN) and clear native (CN)), and high‐performance liquid chromatography (HPLC). In this article, it is shown that HPLC techniques, given their wide versatility, relative ease of use, and high resolution, may be considered the most valuable tool for the characterization of virtually any hydrophobic protein. Application examples are described, and comparisons with other methods are discussed. Moreover, HPLC is not a destructive technique, and therefore, proteins, once separated, are available for further analytical investigations. Among these techniques, quantitative and qualitative analyses of the separated fractions can be obtained through other biophysical approaches, such as crystallography or structural spectroscopy. Most of these approaches require preliminary protein purification (90% or higher), which could be rapidly obtained through preliminary HPLC.