Abstract for BASF Poster:

The long-term stability of high-level expression is the most important factor to consider when choosing cell lines for the expression of recombinant proteins. Declining volumetric yields in large-scale fermentation can be caused by changes influencing the cell population as a whole such as loss in viability, depletion of nutrients or accumulation of metabolites. Alternatively, genetic instability may lead to the outgrowth of a less productive, but metabolically favored sub-population. Currently a variety of parameters are measured to monitor the condition of cells in fermenters, including glucose uptake, lactate accumulation and oxygen consumption; in addition, periodic viable cell counts allow the determination of the growth rate and viability of the population. All of these methods measure the condition of the cell population as a whole and changes must involve a significantly large proportion of the total culture in order to be detectable. Here we report on a method that allows the evaluation of the productivity of individual cells. The gel microdrop secretion assay combines the capture of secreted recombinant protein with analysis using flow cytometry to generate an expression profile of a fermenter sample. Using this method, we detected the appearance of two populations of cells with different productivity rates. Analysis of the recovered sub-populations confirmed the existence of low and high productivity cells that differed in vector copy number. Subsequent testing of an alternative producer line showed a uniform expression profile and consistent long-term productivity. Therefore, the single cell secretion assay proved to be a rapid method to analyze population dynamics in bioreactors and isolate sub-populations of the producer cell line.