Chromatography systems are often defined by their pressure characteristics. Systems that operate at pressures of 50–3,000 psi (3–100 bar) are categorized as medium pressure chromatography systems. This section describes medium pressure chromatography systems, their advantages, general considerations, set up, and flow path, and provides some answers to common questions, and some troubleshooting tips.
Related Topics: Low Pressure Chromatography Systems and Gravity Chromatography.
Medium pressure chromatography is used for the full range of chromatographic separations including all protein purification techniques. Complex gradients, or changes in the proportion of two buffers, are easily possible with medium pressure chromatography systems. These gradients can be generated by a switch/proportioning valve before the pump inlet or by varying the speed of two separate pumps. Both strategies can produce accurate, reproducible gradients. Medium pressure systems can be designed to meet the requirements of the application such as a simple inject and elute strategy to a fully automated system using additional modules and accessories like an autosampler, application specific valves, gradient mixing, UV, conductivity, pH, refractive index and other detectors, and fraction collectors.
Bio-Rad offers two families of medium pressure chromatography systems, the NGC™ platform and the BioLogic DuoFlow™ platform. Both systems are modular, allowing you to easily scale up and add on new capabilities. The NGC system offers new system capabilities and tools to enable a simpler workflow. The Point-to-Plumb™ feature provides a guided light path to allow quick visualization of where to plumb your system and the Tier Rotate™ system feature reduces path length to streamline your separations. Explore the NGC system tour to learn more and simply design your own personalized NGC system using our configurator tool.
Flexibility Medium pressure systems can perform all types of chromatography. They are routinely used for high-resolution ion exchange and high-resolution size exclusion separations. Multi-dimensional purification schemes to simple tagged recombinant proteins purification applications can be easily performed.
High resolution Medium pressure systems produce enough pressure to accommodate higher resolution chromatography media, 5–15 µm.
Automation Most medium pressure systems use software or control systems for unattended operation. Although some low pressure systems offer basic automation, medium pressure systems are designed to operate without user intervention via intuitive software.
Medium pressure chromatography has been used for more than a century to separate and purify molecules. It is typically performed as part of the larger laboratory workflow, either to separate a mixture of molecules — preparative chromatography — or to quantitate highly purified molecules or compounds — analytical chromatography. Medium pressure chromatography has many applications that can be classified either by the type of chromatographic method used (e.g. ion exchange vs affinity) or by the types of molecules being isolated and their characteristics (e.g. antibodies vs organic acids). For further details visit the Chromatography Applications pages.
Bio-Rad offers several medium pressure chromatography systems such as the range of NGC systems and the range of BioLogic DuoFlow™ systems.
Backpressure Medium pressure systems can move the buffer along a flow path when the pressure capabilities of the system exceed the resistance or backpressure after the pump. The following factors will increase backpressure and should be considered in the set up and use of the system:
Sample Application Buffer pumps are used to pump samples onto the column because the column backpressure is relatively high. The plumbing can vary depending on the volume or clarity of the sample. Some examples are:
Medium pressure systems follow the basic flow path: buffer reservoir to pump, to injection valve, to column, to UV detector, to conductivity monitor, to pH monitor, to a fraction collector or outlet valve. Medium pressure systems are often used for high resolution separations so it is important to minimize all tubing lengths and dead volumes. Extra volume in tubing or components will cause "band broadening", the diffusion of distinct peaks into one another.
Fluidics setup and flow path design can be easily achieved using software and instrumentation to indicate the order in which parts should be connected, using the Point-to-Plumb and guided fluidics selection tools on the NGC systems for example. Application-specific flow paths can be easily plumbed using a series of available template designs.
Medium pressure setup and flowpath.
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