Cycle half-life, immunosorbent

The recovery of protein products from fermentation processes by immunosorbent chromatography can be economically attractive. The amount of immunosorbent required is a major cost factor. The proper selection of antibody, matrix, immobilization method and elution conditions can allow high throughput for a given volume of immunosorbent. The throughput depends mainly on the flow rates through the column and cycle half-life. 

As shown in Equation 1, the capacity per cycle is directly proportional to the amount of antibody immobilized, the immobilization yield, the M.W. of the protein and the column volume and an exponential function of the number of cycles. The amount of antibody immobilized will usually be less than 10 gL l. Higher activation of the matrix required for greater than 10 gL l loading results in a decrease in the immobilization yield. The maximum immobilization yield is 1.0 (100%) while 0.8 (80%) is not difficult to obtain. The M.W. of the protein to be isolated is fixed. The only way to increase the capacity per cycle significantly is to increase the volume of the immunosorbent or increase the number of cycles prior to reaching 50% of initial capacity (cycle half-life). Increasing the volume of immunosorbent increases the amount of monoclonal antibody required. 

Since the cost of the antibody is the major cost, increasing the volume of immunosorbent is the most expensive way to increase capacity per cycle. The least expensive way to increase the capacity of the isolation system is to increase the number of cycles. The number of cycles that can be obtained in any given purification is dependent on the cycle half-life and time-volume constraints. The total amount of protein that can be isolated in a given number of cycles is given in Equation 2. 

As the capacity of the immunosorbent column decreases with increasing cycle number the concentration of the purified protein decreases as the volume in which it is eluted is a function of the column volume. If the concentration of eluted protein is 1 in the first cycle, the average concentration eluted in n = n/2 cycles = 0.75. The average concentration in the second cycle half-life = 0.375 and in the third cycle half-life - 0.1875. As a general rule if the cost of the final isolation is 1 for the first cycle half-life, it will be 1.33 for the second cycle half-life and 1.71 for the third cycle half-life. Regardless of the final process step, the decrease in concentration of eluted protein with cycle number will increase the final isolation costs and must be weighed against the cost of antibody needed to increase column volume and decrease the number of cycles. 

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