A typical affinity chromatography setup comprises the following components:
Pump:
Ensures a consistent and controlled flow of the mobile phase (sample solution and elution buffer) through the affinity column.
High-pressure pumps are commonly used for faster runs.
Sample Injector:
Allows precise introduction of the sample into the chromatography system.
Auto-samplers can load multiple samples sequentially for high-throughput applications.
Affinity Column:
The core component containing the immobilized ligand specific to the target molecule.
Typically made of materials like agarose, sepharose, or other polymeric beads covalently linked with the ligand.
Detectors:
Monitor eluted compounds from the column.
Common types include:
Ultraviolet (UV) Detector: Detects compounds based on absorbance at specific wavelengths, useful for most organic molecules.
Fluorescence Detector: For molecules that fluoresce or are labeled with fluorescent tags.
Conductivity Detector: Monitors changes in solution conductivity, useful when elution involves changes in ionic strength.
Fraction Collector:
Collects separated compounds into distinct fractions for further analysis or use.
Temperature Control Unit:
Maintains specific temperature conditions required for stability or optimal binding/elution.
Recorder/Data System:
Interfaces with computers for real-time data acquisition, analysis, and visualization.
Essential for peak identification, quantification, and data storage.