Fractional free volume protein

Equilibrate a 1 x 10 cm Sephadex G-25 column with 50 ml Soln. F. Apply the iodinated sample, elute with Soln. F, and collect 1-ml fractions. The iodinated protein appears in the void volume, and free iodine-125 elutes with the total volume. Estimate its radioactivity and calculate the specific radioactivity (Bequerel per milligram of protein). 

Fine grinding and air classification make possible the production of some cake flour from hard wheat and some bread flour or high-protein fractions from soft wheat. AppHcation of the process theoretically frees the miller from dependence on different wheats, either hard or soft, that change each crop year. The problem is how to market the larger volume of low protein or starch fractions at prices adequate to justify the installation and operation of the special equipment (46). 

In a batch configuration, host cells that contain an expression vector for the recombinant product are added to a predetermined volume of growth medium (Figure 4.12A). The cells are allowed to grow until the nutrients in the medium are depleted or the excreted by-products reach inhibitory levels. At that time, the cells are harvested, and recombinant protein, found in inclusion bodies, cell-membrane fractions, or cytoplasm, are isolated after disruption of the harvested cells. Because the host cells are destroyed at the end of each run, they must be replaced every three to seven days for fermentation or every two weeks for roller-bottle or microcarrier-support production of adherent cells. To ensure uniformity and reproducibility, the FDA requires manufacturers of recombinant proteins to establish and validate a seed stock of recombinant host cells that are validated to contain the characterized expression vector and to be free of contaminants. 

Figure 2 Typical phase diagram of an aqueous polysaccharide (l)-protein (2) dispersion showing the Gibbs free energy as a function of the volume fraction () of each, at different temperatures from Tx, where the dispersion is metastable, to the critical solution temperature (Tc), where the two components are miscible in all proportions. ABC is the spinodal curve DBE (not connected) is the binodal curve.

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