Bioseparation techniques

This textbook describes both theoretical and practical aspects principles of the most frequently used techniques for purification of proteins, peptides, nucleic acids and other charged biomolecules, the media available and how to select them, application examples and detailed instructions for the most commonly performed procedures. Practical information, with many tips and hints drawn from over forty years of experience in chromatography purification, guides beginners and experts towards obtaining the best possible results from the latest chromatographic media in order to choice the most suitable techniques for capture, intermediate purification or polishing steps in a purification protocol and the techniques that are used for purification and analysis of a wide range from microscales to kilograms of product.
Biomolecules purification varies from simple one-step precipitation procedures to large scale validated production processes. Often more than one purification step is necessary to reach the desired purity. This includes any conditioning steps necessary to transfer the product from one technique into conditions suitable to perform the next technique. Each step in the process will cause some loss of product. For example, if a yield of 80% in each step is assumed, this will be reduced to only 20% overall yield after 8 processing steps. Consequently, to reach the targets for yield and purity with the minimum number of steps and the simplest possible design, it is not efficient to add one step to another until purity requirements have been fulfilled. Occasionally when a sample is readily available purity can be achieved by simply adding or repeating steps. However, experience shows that, even for the most challenging applications, high purity and yield can be achieved efficiently in fewer than four well-chosen and optimized purification steps. Techniques should be organized in a logical sequence to avoid the need for conditioning steps and the chromatographic techniques selected appropriately to use as few purification steps as possible. The key to successful and efficient biomolecules purification is to select the most appropriate techniques, optimize their performance to suit the requirements and combine them in a logical way to maximize yield and minimize the number of steps required.