For a list of peer-reviewed articles our products have been cited in, check out the publications page.
Utilizing Waters’ Open Interface Portal (OIP) for multi-vendor hardware control, ProteinSimple’s Maurice Empower® Control Kit enables seamless control of the Maurice platform with all key functions preserved and with full regulatory compliance, including 21 CFR Part 11 controls for industry-leading security and data integrity.
Simple Western was used to separate proteins by charge and size to evaluate the pharmacokinetic properties of adalimumab and two adalimumab biosimilars in human serum.
Viral capsid content can impact gene therapy product efficacy and is therefore considered a Critical Quality Attribute (CQA) that must be properly evaluated during the development and manufacturing of AAVs. Traditional analytical tools such as transmission electron microscopy (TEM), analytical ultracentrifugation (AUC), and ion-exchange chromatography (IEX) can be used to characterize capsid content but are complex, labor-intensive, and pose challenges in data reproducibility, throughput, and scalability. In this application note, we show how imaged-capillary isoelectric focusing (icIEF) technology on Maurice can be used to characterize empty, intermediate and full AAV capsids at native and stability screening conditions, providing robust and reproducible data. With this, Maurice provides crucial data to aid in developing the right formulation for AAV therapeutics.
Therapeutic monoclonal antibodies (mAbs) make up a large portion of the rapidly growing drug market. Ensuring safety and efficacy through comprehensive understanding of these products’ critical quality attributes (CQAs), including charge heterogeneity, is a regulatory requirement. Various charge isoforms of mAbs can result from cell culture or production processes, potentially affecting the mAb structure and function. While imaged capillary isoelectric focusing (icIEF) is the preferred method for charge profiling, ion-exchange chromatography (IEC) has been the major tool for fractionation combined with characterization. However, IEC is not compatible with certain types of molecules, hydrophobic antibody drug conjugated (ADCs) for example, and icIEF typically provides higher separation resolution. Moreover, an individual charge variant obtained from IEC fractionation may not be comparable to the variant peak in the icIEF profile. Therefore, there is an unmet need for IEF-based fractionation of charge variants for characterization.
We have developed a novel icIEF fractionation solution, which involves icIEF separation and collection of charge variants. This solution enables Maurice icIEF-based peak identification followed by downstream analysis. Here we report icIEF fractionation followed by ZipChip-based mass spectrometry (MS) characterization of the NIST mAb and XMT-1535 mAb. ZipChip (CE-ESI) was utilized for mass spectrometry characterization of the fractions due to its broad sample matrix compatibility, easy sample prep, and fast mass spectrometry analysis time. Individual charge variants of each antibody were successfully collected in less than 2 hours with purity > 80% using icIEF separation conditions with or without urea. Rapid analysis using ZipChip chowed the mass spec identification of major and minor isoforms correlated well with reported mass spec data (literature and report). Urea in icIEF separation did not affect the quality of fractionation nor the mass spec result. Multiple fractionation runs of the NIST mAb suggested good reproducibility of the system. We believe this novel icIEF fractionation solution coupled with other analysis methods, such as mass spectrometer, will deliver a powerful charge variant characterization tool for biotherapeutic analytical tool kit.
Keeping track and close control of glycosylation in therapeutic monoclonal antibodies and fusion proteins is crucial to ensure safety and efficacy of these important classes of biotherapeutics. The removal of glycans from biotherapeutics pose a challenge when investigating the changes in physicochemical and pharmacological properties. In this work we used the highly specific enzymes SialEXO®, to enable simplified workflows for glycoprofile analysis by desialylation, OglyZOR® for specific hydrolysis of O-glycans, and FabRICATOR® for digestion of antibodies or Fc-fusion proteins. The results demonstrate simplified workflows to quantitate charge and size heterogeneity associated with the removal of glycans and digestion of antibodies, fusion proteins using capillary iso-electric focusing (cIEF) and CE-SDS. The combination of specific enzymatic sample preparation with robust cIEF and CE-SDS has potential to speed up, increased through-put and simplify routine testing of critical quality attributes when developing or manufacturing biotherapeutics.
icIEF is well-established as the gold standard tool in the biopharmaceutical industry for protein charge characterization. Newer therapeutic modalities are coming to market, including viruses and virus like particles. A challenge associated with icIEF analysis of virus and virus like particle (VLP) samples is sample aggregation during IEF. For most sample (mAbs, proteins), adding solubilizers into the sample solution, such as urea and non-iconic surfactants, can prevent aggregation. However, intact viruses and VLPs may disassociate when these additives are used. While reducing the final sample concentration can help minimize aggregation, UV light absorbance-based detection may not possess sufficient sensitivity to analyze these samples at lower concentrations. In the presentation, we will illustrate the results of using an icIEF-UV fluorescence instrument for charge characterization of viruses and VLP samples. The UV fluorescence method requires no dye labeling and shows significantly higher sensitivity than UV absorption detection.
Since the first therapeutic monoclonal antibody (mAb) was commercialized in the mid-80’s, close to 100 therapeutic mAb products (accounting for around a quarter of all biotech drugs) have hit the market; making it a $125 billion industry that targets critical pathological health conditions – including but not limited to products for antitumor, antiviral, and antiplatelet therapies. From early-stage process development to batch lot release testing, the efficacy, safety, identity, stability, and purity of therapeutic mAb products throughout their shelf life are of crucial importance. Capillary electrophoresis sodium dodecyl sulfate (CE-SDS) has become the gold standard technique for the quality-control of therapeutic mAbs and proteins due to its ease of implementation, robustness, and reproducibility, replacing the more traditional and labor-intensive technique such as SDS-PAGE gel. Successful CE-SDS method development, under both reducing and nonreducing conditions, aims to reduce assay-associated impurities, fragmentations, and aggregations.
Here, we have used the monoclonal IgG System Suitability Reference Standard developed by U.S. Pharmacopeia (USP) to assess the rigor and robustness of an optimized Maurice™ CE-SDS PLUS method compared to the recommended USP protocol provided in monograph <129>. The optimization leveraged Design of Experiments (DOE) to optimize key components in sample preparation, denaturing conditions, and sample injection. The results show that the optimized methods: (1) cause less fragmentation compared to the USP <129> method, (2) are not susceptible to sample injection variations that might differ between instruments, and (3) provide comparable data to the USP <129>
monograph for mAbs.
The studies presented here demonstrate the utility of Maurice platforms in biosimilar development through the characterization of certain monoclonal antibodies, heavily glycosylated species, and PEGylated species.
How Simple Western, Single-Cell Western, Simple Plex and FluorChem Imagers validate biomarkers for neurodegenerative diseases such as Parkinson's disease, dementia, Alzheimer's disease, multiple sclerosis, and epilepsy.
In this Scientific Review, learn how quantitative, high sensitivity Westerns enable cutting-edge Neuroscience research. ProteinSimple offers a range of research tools to study the biological processes that accompany neurological/neurodegenerative disease. Simple Western™ assays on Jess™, Abby™, Peggy Sue™, Sally Sue™, or NanoPro™ 1000 offer fully automated Western analysis by charge or size separation that provide market-leading sensitivity, quantitation, and multiplexing capabilities so you can get the most data out of your precious samples.