Bioprocess Contaminant Detection Using Simple Western
The Simple Western™ family is made up of automated, capillary-based immunoassay platforms that combine the power of CE-SDS or cIEF with the sensitivity of immunodetection using conventional antibodies. Simple Western assays enable Size and Charge-based screening of complex sample types.
- Separate and analyze proteins by immunoassay or total protein content.
- Analyze crude or purified samples for proteins of interest or bioprocess contaminants.
- Quantitate expression levels, isoform distribution, and fragmentation in a gel- free, blot-free format.
- Choose from five instruments of differing throughput and separation mode options.
Current methods for assessing the concentration of residuals, like ELISAs, flow cytometry and traditional Western blotting are labor-intensive and prone to error. These traditional methods can also be misleading due to a lack of specificity and flexibility required to obtain a comprehensive contaminant profile and may exclude relevant critical information like size-based results, oligomerization state and low limit of detection.
Highly sensitive Simple Western immunoassays deliver accurate detection of residual proteins resulting from therapeutic protein and vaccine development processes like host cell protein (HCP), Protein A, GFP, and BSA.
Measuring empty and full adeno-associated viral vectors (AAVs) is a major challenge in developing gene therapies for which current methods are limited. Now, there is a novel method for quantifying the DNA content of AAV particles with Simple Western that seamlessly combines CE-SDS/cIEF and immunodetection with conventional Western blot antibodies.
The Simple Western method automatically separates AAV samples by Size or Charge followed by specific and sensitive detection using anti-DNA and anti-VP1/2/3 antibodies directly in the capillary for quantitative and reproducible measurement of % full AAVs to total AAVs ratio in a sample, or Content Ratio. This Simple Western assay for empty/full viral vectors can also apply to the capsid content (and stability) of lentivirus!
Traditionally, host cell-related impurities are monitored by ELISA using an antibody targeting HCP (anti-HCP) or by SDS-PAGE with total protein staining. Simple Western does both! Anti-HCP antibodies may be used on Simple Western with the added advantage over ELISA of size separation profiles and minimal matrix interference. Additionally, Simple Western has total protein detection with the 5X Total Protein Labeling Reagent with sensitivity that rivals SYPRO Ruby. A major advantage of the 5X Total Protein Labeling Reagent on Simple Western is that it can label any protein regardless of host cell origin. Therefore, Total Protein Detection on Simple Western eliminates the need for an anti-HCP antibody to detect impurities and may offer more complete coverage of the host cell proteome.