Reverse-phase high-performance liquid chromatography for protein determination
Reverse-phase high-performance liquid chromatography (RP-HPLC) is a commonly used protein analysis method. This method utilizes the differences in proteins under polar and non-polar conditions to achieve separation and detection. This article will explain how to use RP-HPLC to determine proteins.
1. Sample Preparation
First, appropriately process the protein sample, such as concentration, desalination, dissolution, etc.
2. Chromatographic Condition Adjustment
Select suitable mobile phase and stationary phase according to the properties of the protein.
3. Sample Loading
Inject the processed sample into the chromatographic column through an auto-sampler.
4. Protein Separation
By changing the polarity of the mobile phase, proteins will be separated on the chromatographic column according to their hydrophilicity or hydrophobicity.
5. Detection and Analysis
The chromatograph uses a UV detector to convert the separated proteins into electrical signals, which are then transformed into chromatograms.
6. Result Interpretation
Determine the content and purity of proteins by the peaks on the chromatogram.
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