With the development of molecular biology, more and more researchers have mastered various experimental techniques of molecular biology, but the upstream work of molecular biology is often not our goal, the key is to obtain pure expression products to study its biological effects. Compared with the upstream work, the protein purification work is very complicated. In addition to ensuring the purity, the protein product must also maintain its biological activity. The purification process of protein purification equipment must be able to produce the same quantity and quality of protein every time, with good repeatability.
Under laboratory conditions, in order to achieve large-scale production and application of protein purification, and to have good repeatability in daily applications, it is necessary to understand the basic principles of protein purification equipment and the principles and advantages of various protein purification technologies And limitations, in order to provide some guidance for the selection of purification methods and the formulation of the overall plan for protein purification equipment.
Protein purification equipment should use the inherent similarities and differences between different proteins, as well as the similarities between various proteins to remove the contamination of non-protein substances, so as to purify the target protein from other proteins. Taking advantage of these differences in size, shape, charge, hydrophobicity, solubility, and biological activity between each protein, proteins can be extracted from mixtures such as E. coli lysates to obtain recombinant proteins.
Protein purification equipment is mainly divided into two stages: rough separation stage and fine purification stage. The rough separation stage of protein purification equipment is mainly to separate the target protein and other cell components. The sample volume is large and the components are miscellaneous. Therefore, the resin used has high capacity, high flow rate, large particles and large particle size, and can quickly separate proteins and pollutants. Prevent target protein from being degraded. The polishing stage of protein purification equipment requires higher resolution to distinguish the protein of interest from those of similar size and physicochemical properties. In order to improve the resolution, it is necessary to use smaller resin particles. Usually, it is also necessary to consider the use of ion exchangers and hydrophobic columns at the same time, and comprehensively consider the two factors of resin selectivity and column efficiency.