Protein purification can be a costly and time-consuming process in the manufacture of biomolecules. Either from a tissue or by their overexpression in a model organism, e.g. bacteria, yeast, or mammalian cells in culture, one of the objectives of the process is to obtain the highest functional protein yield while minimizing contaminants.
Many factors can affect the quality and results of the process for protein purification. Thus, to obtain the desired yield and purity, optimization and selectivity of the proteins from the crude cell lysates is a must, as well as ensuring the proper conditions of the solution at every step. A good buffer environment is required to avoid changes in the pH that could affect the solubility, function and folding of the protein. Some of the characteristics of a good buffer are: Chemical stability, water solubility, high capacity at expected pH, and compatibility with analytical and experimental applications.
Despite the purity and integrity quality assessment of protein samples and the different techniques to detect potential contaminants, low amount impurities and degradation products can go unnoticed, especially in low concentration samples or during optimization phases in which minute aliquots are analyzed
To preserve the yield, it is important to differentiate the impurities that must be removed completely and those that can be reduced to acceptable levels, since different types of material will contain different impurities. In any case, the quality of the end-product should never be compromised. A good practice is to work with highly efficient techniques at early stages in the process to capture the product and reduce the contaminant content, and then continue to work with increased selectivity in later steps for optimal separation. Some other basics rules include: Work quickly, eliminate protease to keep your target protein active, minimize sample handling, combine techniques with complementary selectivity, work from large beads to small beads
At CBL we count with extensive hands-on experience and expertise with immunoaffinity purification of proteins.