This is an application note that describes a high recovery HPLC method for the identification of human brain lipid raft membrane proteins based on a macroporous reversed-phase column.
Membrane proteins are critical for cellular processes but are difficult to study due to their hydrophobic nature and low abundance. This document presents a robust workflow developed by Agilent Technologies to overcome these challenges, specifically focusing on the analysis of lipid raft domains. The method utilizes a novel macroporous reversed-phase column to achieve high protein recovery and excellent separation.
| Column Dimensions | 4.6-mm x 50-mm |
|---|---|
| Part Number | 5188-5231 |
| Flow Rate | 0.75 mL/min |
| Column Temperature | 80 °C |
| Detection Wavelength | 280 nm |
| Pressure Limit | 250 bar |
| Proteins Identified | 158 proteins (73 membrane) |
The part number for the 4.6-mm x 50-mm mRP-C18 column is 5188-5231.
The recommended column temperature is 80 °C.
Using this strategy, 73 membrane proteins were identified out of 158 total proteins.