1. Purification of biological macromolecules
Gel filtration is separated according to different molecular weights. Due to its separation characteristics and the advantages of simplicity, convenience and no change in the biological activity of samples, gel filtration has become an important means of purifying biological macromolecules, especially for molecules with different sizes but similar physical and chemical properties, it is difficult to separate them by other methods, and gel filtration is undoubtedly a suitable method, For example, for the separation of polymers with different degrees of polymerization, etc.
2. Molecular weight determination
Gel filtration is an effective method for preliminary determination of molecular weight of proteins because it is simple and requires less samples. The disadvantage of this method is that the accuracy of measurement results is affected by many factors. Since this method assumes that neither the standard matter nor the sample has adsorption with the gel, if the standard matter or the sample has certain adsorption with the gel, the measurement error will be relatively large. The calculation formula is valid only when the protein is basically spherical, but not for some elongated proteins such as fibrin. In addition, due to the strong hydration of sugar, the molecular weight of glycoprotein determined by gel filtration is relatively large, while the measured value of ferritin is relatively small.
3. Desalting and removing small molecule impurity
Using gel filtration to desalinate and remove small molecule impurities is a simple, effective and fast method, which is much faster than the general method of dialysis desalting, and generally does not cause large dilution of samples, and biomolecules are not easy to denaturate. The commonly used is Welch Tandex G-25, and there are a variety of desalting column products for sale, easy to use, can be used for many times.
4. Heat removal materials
Pyrogenic substances refer to some polyglycoprotein complexes produced by microorganisms that heat the human body. They are a class of substances with large molecular weight, so they can be separated from other substances with relatively small molecular weight by the exclusion effect of gel filtration. For example, gel filtration is a simple and effective method to remove heat source substances in water, amino acids and some injection.
5. Concentration of solution
The hydroscopicity of gel particles can be used to concentrate macromolecular sample solutions. For example, if dry Tandex (coarse particles) is added to a solution, the Tandex can absorb large amounts of water, and the small molecules in the solution can also penetrate into the pores of the gel, while the large molecules are excluded. The gel particles are removed by centrifugation or filtration to obtain a concentrated sample solution. This concentration method basically does not change the ionic strength and pH value of solution.
6. Protein renaturation
In order to reduce aggregation at high concentrations, gel filtration chromatography can also be used for protein renaturation in vitro. The isolation effect of the chromatography medium reduced the aggregation of protein interaction, so that the renaturation concentration and renaturation rate were greatly improved. Meanwhile, protein can also be purified by gel filtration chromatography.
Two important factors affect the yield of protein renaturation by gel filtration chromatography: the first is the loading of protein under denaturation conditions, and the second is the change of protein structure in the renaturation buffer. In addition, the amount of protein loading also affects the protein renaturation yield because of the aggregation of structures at the tip of the column due to the increase of the loading amount.
In order to improve the efficiency of protein renaturation, a gradient gel filtration chromatography method was developed. Preset in the chromatographic column concentration denaturant gradient, when the renaturation of protein into the post, because of the apparent molecular weight than denaturant protein, and protein after a denaturant concentration gradually reduce environment, protein folding method step by step, so as to effectively reduce the formation of aggregate, and the aggregation and folding of protein separation.
Linear gradients may be frequently encountered in gel filtration chromatography, but sometimes some proteins may be unstable at certain denaturant concentrations, and these concentration processes need to be crossed as soon as possible during the gradient process. Sometimes the speed limit of protein folding is at some denaturant concentration, and this period of time needs to be increased. Therefore, different types of gradient forms can be designed in gel filtration chromatography to meet the renaturation needs of different proteins.
Welch Materials gel filter filler:
- Tandex G series and LH20 multi-mode gel filtration series
- SuperTandex Prep Grade series
- Tanrose Fast Flow series
- Tanrose/Tanrose CL series
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