SmartFlow Technologies

Isolation of Secreted Proteins from Mammalian Cell Culture

Optimization Procedure

This Isolation of secreted proteins from mammalian cell lines WORKS Optimization Procedure is intended to isolate recombinant proteins such as monoclonal antibodies (Mab) from cell culture media.

This optimization procedure uses a microfiltration (MF) membrane that retains the cells and cell debris and allows the desired secreted protein to pass freely through the membrane. The passage characteristics of secreted proteins from mammalian cells change with different buffers, temperatures, concentrations, and membranes.

By examining the passage characteristics of the different MF membranes available in the appropriate process conditions, a well defined and executed process development study can identify the most efficient membrane and process conditions to achieve the required performance.


This Isolation of secreted monoclonal antibodies from mammalian cells protocol is intended for isolating, concentrating and diafiltering recombinant proteins such as monoclonal antibodies (Mab) in two simultaneous processes. This process has been repeatedly implemented with consistent success in CHO cell culture.

The initial step isolates the target Mab from the culture medium by using a 0.45 μm modified polyethersulfone membrane (MPS) to pass the Mab freely into the permeate and retain the cells, large molecular weight broth components, and any accumulated cell debris. The protocol calls for the media to be concentrated to 5X prior to starting the diafiltration. The required diafiltration buffer is generated in the second ultrafiltration process.

The target of the second step is to concentrate the target molecule with a 50kD polyethersulfone (PES) ultrafiltration membrane. The permeate from this process is fed back to the recirculation loop of the isolation process to create a closed loop system. The target product is concentrated in the retentate tank of the second loop and recovered when the desired concentration or target yield is achieved.

Case Study

Process development scientists at a major biopharmaceutical company have looked at concentrating CHO cells while passing IgG through a MF as an alternative to centrifugation for the initial step in their downstream processing. Industrial scale cell clarification with traditional TFF formats have been subject to low flux rates and unacceptable product yields. Centrifugation combined with depth filtration is reported to provide acceptable yields but require significantly higher initial capital investment to implement.