Accelerate Your Cell & Gene Therapy Workflows with Advanced Western Blotting Solutions
Develop and characterize your therapeutic candidates with Simple Western™ systems. The automated sample processing of Simple Western systems gives you reproducible, accurate and precise data with less hands-on time and faster time to market.
THE SIMPLE WESTERN ADVANTAGE IN CELL & GENE THERAPY
Sensitivity: Conserve your precious AAV sample with AAV purity measurements that are more sensitive than SYPRO Ruby. Boost your immunoassay sensitivity using Simple Western which can detect as little as 150 pg of protein in 3 µL of sample.
Quantitation: Dose response curves made easy. Simple Western systems provide quantitative protein measurements with a 3 log dynamic range
Reproducibility: Simple Western eliminates the unavoidable variation from traditional western blotting methods, with intra-assay CV's of <15%.
Throughput: Simple Western can generate up to 96 data points in a single overnight run, all in a fully automated fashion! You can generate quantitative data and screen therapeutic candidates with triplicate datapoints overnight - something that would take days, if not weeks, with traditional Western blotting methods.
Large protein detection: Detect and analyze large molecular weight proteins like dystrophin that can be challenging to transfer and analyze with traditional western blotting.
Method transferability: Take advantage of full assay automation and Compass for Simple Western to get digital run files containing assay conditions and data, allowing for easy analysis, transfer and archiving between sites or from sponsors to CROs.
Compliance: Compass software is 21 CFR Part 11 compliant and is GMP ready for your organization's needs.
Hear from your Peers!
"I used Simple Western because the protein samples I had were small. We only had a small number of [iPSC-derived macrophages] because the cell line does not proliferate." - Elena Navarro-Guerrero, Ph.D., Senior Postdoctoral Researcher, Nuffield Department of Medicine, Target Discovery Institute
Multi-attribute, automated solutions for measuring identity and purity of AAV samples in one run
Measuring the critical quality attributes (CQAs) of viral vectors can be time-consuming. What if you could get all the CQAs you need at once? Simple Western is a multi-attribute method capable of characterizing many quality attributes, like empty/full capsid content ratios, VP protein ratios, identity, purity, and potency. Get to the answers you need faster with Simple Western!
DO YOUR AAVS CONTAIN DNA? THE SIMPLE WESTERN AAV EMPTY/FULL ASSAY
Measuring empty and full AAVs is a major challenge in developing gene therapies for which current methods are limited. Simple Western is a rapid and sensitive assay for accurately quantifying the ratio of % full AAVs to total AAVs in a sample, called the Content Ratio. A major advantage of this method is that Simple Western is a fully automated immunodetection platform with a throughput of up to 96 samples that can be processed overnight. This automation not only decreases labor costs but also enables faster iterations during in-process development. Equally advantageous is the tiny sample volume requirement of Simple Western (as little as 3 μL sample) that minimizes impact on final product titer and allows for the analysis of as few as 7.2 x 107 VP per well.
VP PROTEIN STANDARDS FOR ACCURATE VP PROTEIN RATIO AND VIRAL PROTEIN TITER MEASUREMENTS
Comprehensive monitoring of VP1, VP2, and VP3 protein concentrations and their molar ratios is an effective tool to ensure the optimization and consistent quality of final AAV gene therapy products. In traditional western blots, differences in transfer efficiencies of different proteins can result in misleading signal intensities in a blot analysis. Simple Western, however, shows no significant differences in the intensities of identical amounts in stoichiometric VP mixtures and allows for reproducible quantification of VP protein ratios and titer. Learn how PROGEN´s recombinant AAV2 VP standards are used with Simple Western in fit-for-purpose solutions for VP capsid protein ratio and viral protein titer measurements.
Monitor AAV Capsid Identity, Purity, and Capsid Ratios During Product Purification
Simple Western immuno- and total protein assays can be used to monitor AAV capsid proteins throughout the purification process. From measuring the expression of novel AAV capsids to downstream manufacturing and QC, Simple Western systems can accurately measure product identity and purity where it is imperative. In this Application Note, learn how the Cell and Gene Therapy Consortium Catapult uses highly specific PROGEN antibodies in fully automated Simple Western assays to monitor and characterize AAV capsids during product purification.
Immunodetection of VP1, VP2 and VP3 proteins during purification from whole-cell lysate
Total Protein detection of VP1, VP2 and VP3 proteins during purification from whole-cell lysate.
The Next Generation of AAV Characterization Tools
As the gene therapy field continues to expand and companies increase the number of product batches manufactured per year, quality control requirements for product release have increased significantly. In this webinar, Tony Bou Kheir, PhD, from the Cell and Gene Therapy Catapult in London, showcases an advanced analytical platform for AAV vector characterization, addressing industry challenges in 1) assay precision, 2) standardization and throughput, and 3) enabling in-line product characterization in closed processes.
Limited in AAV sample availability? In this virtual poster, we describe a method to monitor the purification of AAV2 using automated capillary electrophoresis followed by immunoassay and total protein detection directly in the capillary, eliminating the need for SDS-PAGE. In this collaborative poster between the Cell & Gene Therapy Catapult, PROGEN and ProteinSimple, we describe a highly sensitive, fully automated assay which effectively reduces sample size requirements down to 3 µL of AAV sample, corresponding to approximately 400 pg or 1x108 genomic copies loaded per well.
Download AAVAnalysis PosterIdentify Process-Related Impurities
Current methods for assessing the concentration of residuals like enzyme-linked immunosorbent assays (ELISAs), flow cytometry and traditional Western blotting are labor-intensive, prone to error and can also be misleading. Simple Western systems automate the process start to finish, and capture critical information on sizing and oligomerization, giving you more flexibility to build a comprehensive contaminant profile. The methods outlined in this application note for the evaluation and characterization of HCP, Protein A, GFP and BSA offer proof-of-concept methods for adaptation as quantitative and reliable protocols throughout the development process of a biotherapeutic.
E. coli HCP antigen detection on Simple Western. Compass for Simple Western lane and graph views. (A) The concentration of HCP lysate plotted against the average background-subtracted total peak area (B) E. coli HCP antigen was diluted in 0.1X Sample Buffer and titrated from 33.3 µg/mL to 1.23 µg/mL following default Simple Western protocols. Goat anti-E. coli HCP (1:10) was diluted in Antibody Diluent 2 (PN 042-203), followed by the anti-goat secondary HRP conjugate.
Ultra-High Sensitivity AAV Purity Measurements to Conserve Your Precious AAV samples
Many manufacturers of AAVs still rely on traditional SDS-PAGE with SYPRO Ruby staining to monitor purification. Unlike Simple Western, SYPRO Ruby staining is labor-intensive with many manual washing steps, generates large volumes of liquid waste, and requires special imaging equipment. Furthermore, SYPRO Ruby requires at least 1 ng of protein for reliable detection. In contrast Simple Western can reliably detect as little as 150 pg. In this application note, we demonstrate a Simple Western total protein assay workflow for AAV purity measurements that is more sensitive than SYPRO Ruby. Get the purity information you need while conserving your precious AAV samples!
Side-by-side comparison of high sensitivity AAV total protein assay on Simple Western (left) and SYPRO Ruby (right) shows Simple Western outperforms SYPRO Ruby in sensitivity in addition to improved automation and time to results.
REPRODUCIBILITY YOU CAN COUNT ON INCLUDING FOR HIGH MOLECULAR WEIGHT PROTEINS LIKE DYSTROPHIN
Let's face it: some proteins are just tricky to analyze. Take dystrophin, a high value gene therapy target, as an example. The large size of the dystrophin protein and its susceptibility to proteolysis pose challenges with extracting the protein, and there can be further difficulties associated with electrophoretic separation and repeatable membrane transfer, making it difficult to reliably detect, let alone quantify, with traditional techniques like Western blot.
Dystrophin's large molecular weight is no problem for Simple Western which can easily detect and quantify proteins from 6 to 440 kDa. This major advantage over traditional Western blot is accompanied by automation, high throughput, high sensitivity, and low-sample size requirements. In this publication spotlight, we highlight the rapidly increasing contribution of Simple Western to Duchenne Muscular Dystrophy (DMD) research, including in the world's premier journals like Science and Nature Medicine.
Download Spotlight
Dystrophin protein expression was analyzed by Wes and compared with HEK293T cells overexpressing dystrophin cDNA. Figure adapted with permission from Gee et al.
SINGLE-CELL WESTERNS MEASURE GENE EDITING EFFICIENCY FOR SICKLE-CELL RESEARCH
Single-Cell Westerns measure protein expression in thousands of single cells per run. Their high sensitivity combined with the ability to use conventional Western blotting antibodies makes them particularly well-suited for quantifying gene therapy efficacy and tracking differentiation. For example, Milo can resolve hemoglobin subunits in single red blood cells (RBCs), something that is nearly impossible to do with flow cytometry, enabling researchers to track efficacy of sickle cell gene therapies. In addition, Milo offers the ability to quantify editing efficiency of hematopoietic stem cells (HSCs), quantify differentiation efficiency of HSCs into RBCs, and differentiate between hemoglobin subunits, measuring their expression with existing commercial reagents. Read this publication spotlight to learn how Milo's Single-Cell Westerns support clinical trials aimed at advancing gene therapies for Sickle Cell Disease and other hemoglobinopathies.
Single-Cell Western analysis of red blood cells from patients treated with LentiGlobin BB305 Drug Product. Figure adapted with permission from Bonner et al.
Simple Western
Single-Cell Western
Other Cell and Gene Therapy Resources
Cancer &
Immuno-Oncology
SARS-CoV-2
Cell & Gene Therapy
C> Western Blotting Solutions
Targeted Protein Degradation
SARS-CoV-2 Serology
Exosome
Vaccine Development
Neuroscience
