Fast Protein Liquid Chromatography

Introduction

Fast protein liquid chromatography is a protein friendly, high-performance liquid chromatography. It uses high resolution small-diameter stationary phase for proper protein separation and characterisation. Apart from proteins, biological samples like oligonucleotides and plasmids can be used as a sample in this method. Fast protein liquid chromatography allows the user to monitor various parameters at a time, like pH, conductance and UV levels. It also allows multiple columns to run in a tandem motion, which minimises the time to isolate pure protein. It is mostly used in biochemistry and enzymology laboratories. The technology can use desired flow rate for the mobile phase.

Basic features of Fast protein liquid chromatography

The basic feature of Fast protein liquid chromatography is to give versatile purification of biomolecules like proteins, peptides, biopolymers, etc. The volume of the sample may range from 100 ml to litres. The whole Fast protein liquid chromatography apparatus is placed in a cold room of about 4 degrees Celsius. This has to be maintained so that the structure of biomolecules or proteins is not denatured or disturbed. The pressure used in technology ranges to a few bars; thus, it is also called medium pressure liquid chromatography. Here the velocity of the solvent is by the microprocessor via software interface to maintain the constant rate of flow of the solvent. Usually, Fast protein liquid chromatography notes from various online sites have their basic features, properties, and principle, making understanding the process easier. The Fast protein liquid chromatography protocol has the procedure for different fast protein liquid chromatography types. The fast protein liquid chromatography finely extracts protein biomolecules from a sample mixture, the process is fast, so it gets it the name. The process can help in the cost-effective separation of protein biomolecules like enzymes, hormones and even venoms. During the process, no decomposition or denaturation of protein is observed, which is the superiority of this technique. 

Principle of Fast protein liquid chromatography

The Fast protein liquid chromatography is of four types: 

• Ion exchange chromatography: This method allows the separation of the molecules based on their polarity or charge. Cations and anions can be separated using this method. The separation occurs by reversible exchange of ions between the ion exchange resin and the ions present in the solution. According to the character, the nature of resin exchange can be of four types: strong cation exchange resin,  strong anion exchange resin, weak cation exchange resin and weak anion exchange resin. The resin used can be natural like dolomite, zeolites or synthetic resins, either organic or inorganic. The ion exchange resin must have some quality for the procedure. It should be chemically firm, not reactive, denser than water, and have sufficient ion-exchange groups. The first step to this procedure is the stationary phase reaching equilibrium before applying the sample. After that, elution is done, and the product is collected. 
• Affinity chromatography: It is generally used for sample purification. It separates the mixture based on a highly specific interaction such as antigen-antibody interaction or enzyme-substrate interactions. The molecule of interest should have known properties and be passed on a typical gel matrix in this process. Affinity chromatography is used for the purification of recombinant proteins. The target molecule is trapped on the stationary phase while the mobile phase with impurities eluted out. The stationary phase will be then washed off to release and collect the target molecules. 
• Reversed-phase chromatography: it is also known as adsorption chromatography. Here the binding of mobile solute to an immobilised aromatic ligand or n-alkyl hydrocarbon occurs by hydrophobic interaction. The elution order is reversed, the polar compounds are eluted first, and then non-polar compounds are retained, so it is called reversed-phase chromatography.
• Size exclusion chromatography: It is also known as gel filtration chromatography. It separates molecules based on their size. Here a mixture of molecules are dissolved in the mobile phase and applied to the chromatographic column. In this type of chromatography, the stationary phase is a porous packing material, usually a bead matrix. The mobile phase is the buffer that has the sample dissolved and passed through the stacked matrix. The matrix bead acts as a sieve and entraps the small molecules temporarily, and the larger molecules are eluted or excluded from the columns. Particles of different sizes will be eluted at a different rate through the stationary phase.

Advantages and Applications of Fast protein liquid chromatography

Fast protein liquid chromatography applications are :

• Assessment of protein foldings
• Separation of plasma protein in urine and Cerebral fluid
• Diagnosis of beta thalassemia
• Rapid purification of RNA
• Separation of lipoprotein 
• To analyse the nitrogen constituents in beer
• It can efficiently purify animal venoms
• Isolation of compounds from erythrocytes

Fast protein liquid chromatography advantages are as follows:

• It has a versatile resolution
• The programming system is simple
• A wide range of column supports is possible since the pressure used in Fast protein liquid chromatography is low.
• Suitable for preparative as well as analytical chromatography.

Conclusion

The Fast protein liquid chromatography system is an efficient system for separating proteins. They can separate proteins based on different factors like their charge, size, affinity and their interaction. This wide range of character-based possibilities helps choose the appropriate method and perform the process successfully. It helps us to extract 100% pure protein extract. We can also check various factors like pH, conductance and UV levels at the time of the experiment. The process for fast protein liquid chromatography systems is different, but the Fast protein liquid chromatography system protocols available online at different biochemistry sites can help in the practical procedures.