Protein Binding in Sterile Filtration of Cell Culture Media
Considerations in Sterile Filtration of Cell Culture Media
Single-use, sterile vacuum filtration devices are commonly used in cell culture media preparation and aqueous liquid processing. For cell culture applications, fast-flowing filter materials such as polyethersulfone (PES) that exhibit low protein binding properties are preferred. Various brands of PES vacuum filters exist with a range of filter surface areas. Larger filter surface areas generally translate to faster speed and higher throughput, but one must carefully consider the potential for unwanted protein binding and loss of critical culture media components during filtration.
Protein loss or binding to filter material during media filtration can significantly affect the reliability and reproducibility of scientific experiments and assays involving cell culture. Culture media, which requires sterile filtration, may contain serum proteins, additives, inhibitors, drugs, and more depending on the research being performed. Reductions in critical media components can interfere with cell health and sample integrity, reduce signal-to-noise ratios, and lead to inconsistent results. During filtration of culture media, proteins and other large biomolecules in sera, including immunoglobulin IgG (approx. 150 kDa),1 can interact with filter surfaces. Protein binding to filter membranes can result from a range of factors including surface charge, hydrophobicity, time exposure, and pore size of the filter as well as the specific characteristics of the protein itself.
To address these challenges, we developed Stericup® and Steritop® vacuum filtration systems with fast-flowing PES Express® PLUS filters that exhibit low protein binding properties, reducing processing times while minimizing protein adsorption. These filtration systems help maintain critical components in media including sera proteins, IgG, antibiotics, growth factors, and other additives, ensuring preservation of the desired levels of these components.2 Researchers seeking even lower protein binding can select our Stericup® filtration systems with 0.22 µm Durapore® PVDF membrane.
IgG-Binding Radioimmunoassay
We compared protein binding across sterile filtration devices from different suppliers using a custom radioimmunoassay for IgG binding. Evaluation of filter materials was performed using 125I-labeled goat IgG to determine the amount of bound protein. Replicate filter discs were extracted from each vacuum unit and placed into borosilicate tubes to minimize differences in device design. Each disc was incubated in tracer solution (1 mL of 1 mg/mL goat IgG in phosphate-buffered saline (PBS) containing 125I-goat IgG at a concentration of 0.1 µCi/mL) for 2 hours with agitation, washed, and assayed for bound radioactive protein.
Comparison of Sterile Filtration Devices for Culture Media Preparation
We compared filters from eight brands of sterile vacuum filtration devices. Filter surface areas were determined by measurement or using published values (see Figure 1). Surface areas ranged between 34 cm2 and 64 cm2 of PES filter material for all devices tested. Low and high filter binding controls run in parallel were consistent with nature of the polymer structure types (very low ≤10 µg/cm2 and high binding ≥150 µg/cm2; data not shown).
Figure 1.Surface areas of filters from sterile vacuum filtration devices using published or measured values. Values ranged from 38 cm2 to 64 cm2. The filter from Brand F had the smallest surface area. Filters from Brand B, C, and E had the largest membrane surface area.
A 125I-labeled goat IgG radioimmunoassay was used to measure the protein binding capacity of each filter. Results indicate that PES Express® PLUS filters from the Stericup® devices exhibited significantly lower protein adsorption rates of IgG compared to filters from the other brands (Figure 2).
Figure 2.Amount of 125I-goat IgG bound per square area of PES filters from different sterile vacuum filtration devices. The amount of bound IgG protein ranged from ~24 µg/cm2 to 170 µg/cm2. The PES filter from the Stericup® sterile vacuum filtration device exhibited the lowest binding of all devices tested.
Total protein binding was then calculated for each filter (Figure 3). These results can be used to determine the expected loss of protein during routine filtration of sera-containing tissue culture media. Results clearly demonstrate fast-flowing PES Express® PLUS filter membranes from the Stericup® vacuum filtration device exhibited lowest protein adsorption. The filter from Brand A also exhibited similarly low levels of protein binding. Filters from devices from other suppliers exhibited up to ~8x more binding, which could lead to reductions of key components in the filtrate and potential undesired impacts on downstream usage of the filtered media.
Figure 3.Total bound protein binding ranged from ~1000 µg to more than 8000 µg across filters from tested sterile vacuum filtration devices. Filters from the Stericup® device and Brand A exhibited the lowest total protein binding. Filters from Brand G exhibited 8x higher binding than the lowest binding filters.
Disposable Vacuum Filtration Units for Culture Media Preparation
Stericup® and Steritop® vacuum filtration systems with low protein binding, fast-flowing 0.22 µm PES Express® PLUS filters offer considerable advantages in sterile filtration of cell culture media and reagents. This study underscores the superior performance of Stericup® and Steritop® devices compared to other brands with higher unwanted binding characteristics, as measured by a radioimmunoassay for IgG binding. The low protein binding properties of Stericup® vacuum filtration devices help to minimize non-specific protein adsorption to the filter material, ensuring preservation of critical levels of media components in tissue culture applications. By utilizing vacuum filtration devices with low protein binding, researchers can achieve more consistent cell cultures and reproducible results in cell-based experiments.
Steritop® Filter Units
Steritop® Bottle-top Filter Units can be used on bottles with 33 mm or 45 mm thread.
Stericup® E and Steritop® E Eco-Friendly Filter Units
References
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