(93e) Effect of pH on Cell Recovery and Density during Isopycnic Separation of Cells Using Percoll Density Gradient

The ability to separate and recover cells is an important process in biological research. Percoll is a density gradient media used for the fractionation of cells based on the cell density. This mode of separation is known as isopycnic separation. Percoll is a solution containing colloidal silica coated with polyvinylpyrrolidone (PVP). The properties of Percoll are such that it can be customized as desired. In this study, it is hypothesized that osmolality, pH, and cell number play an important role in the determination of cell density and recovery after isopycnic separation. This study attempts to analyze the effect of the basic nature of Percoll and cell loading on the cell recovery and density at constant osmolality.

A continuous density gradient was prepared after Percoll was modified as per the manufacturer's protocol and was also modified as per our protocol. Different types of primary cells as well as immortalized cell lines were utilized to analyze the hypothesis. This study discusses the results with leuckocytes, Chinese Hamster Ovary (CHO), and MCF-7 cell lines.

Preliminary experiments have been conducted using the CHO cell line and initial results indicate that the pH of Percoll does not affect the recovery and density. But the initial loading does have an effect on cell recovery. The results from the manufacturer's protocol and our protocol were compared and basic statistical analysis was performed. Additional experiments are planned to test the effect of pH on primary cells, including leuckocytes and fibroblasts, as primary cells tend to be more sensitive to extra-cellular conditions than immortalized cell lines.