Laboratory Exercises in Bioseparations

Chapters 3 to 11 of this text are organized around the study of various unit operations in the approximate order of their customary application to bioseparations. In this chapter, five of these operations are singled out for further exploration in the laboratory. These are flocculant screening (Chapter 3), crossflow filtration (Chapter 4), centrifugation of cells and lysate (Chapter 5), aqueous two-phase partitioning of a protein (Chapter 6), and gradient-elution ion exchange chromatography of test proteins (Chapter 7). Each section of this chap­ter is thus an independent laboratory exercise. The instructions can be applied flexibly to the materials and equipment available at a particular laboratory or department. The calculations, reporting, and scale-up applications are applicable to any experiment that follows the generic paradigm of each of the sets of lab instructions. The pattern to be followed consists of becoming acquainted with the equipment and describing it as a unit operation in a report, execution of a predesigned experiment, recording of appropriate data, analysis of the data in the context of this textbook, presenting reduced data in a report, critically analyzing the quality of the results, and, finally applying the actual numerical results to a scale-up to production scale. Process economics may be applied where appropriate. In this laboratory exercise, a flocculant will be evaluated for its ability to flocculate cells or lysate particles. Lysate particles are smaller and require, typically, higher concentration of flocculant than that required to flocculate whole cells. The flocculant concentration required will be determined by observing the persistence of flocculation and clarity of supernatant, measured as a function of flocculant concentration. Flocculants are usually polymers with properties, such as charge, that cause them to interact with cells or lysate particles and bind them together. Their effectiveness depends on molecular weight, charge, solubility, and other properties, and their interactions with cells and particles depend, therefore, on pH, ionic strength, temperature, and dry solids concentration. Unless a great deal is known about the suspended material of interest, or extensive experience has been published, the choice of flocculating conditions usually depends on educated trial and error.