Challenges
In the past few years, the use of biomolecules for therapeutic purposes has dramatically increased. For example, the market for monoclonal Antibodies (mAbs) for therapeutic purposes was valued at $44.6 billion in 2011, and is expected to grow 5.3% by 2016. Compared to small molecules, which are synthesised using chemical reactions, production of biomolecules is a more complex procedure, typically involving cultures in bioreactors using specially engineered cell lines.Therefore, it is critical for biopharmaceutical manufacturers to be able to monitor several parameters in every step of their process workflow to ensure consistent and optimal production of biomolecules. Analytical groups supporting biological drug development efforts require rapid, sensitive, and precise methods for quantifying proteins from cell cultures, fermentation harvests, or purification intermediates, and need to be able to monitor bioreactor product titers to determine when to harvest or abort a reactor cycle.
HPLC
To ensure, reliable, real-time and rapid analysis of biomolecules, high-performance liquid chromatography (HPLC) products are used to separate mixtures of compounds and to identify, analyse and quantify the biomolecules in almost any mixture. Successful HPLC-applications depend on the use of columns containing highlyspecific ligands bound to HPLC-resins; the ligands selectively bind the biomolecule and the resins allow for rapid purification and quantitation.The new HPLC products launched as a result of the collaboration between BAC and Life Technologies, combine the selectivity of CaptureSelect® ligands with the benefits of the POROS® resins. CaptureSelect® ligands are based on single-chain antibody fragments with desirable chromatographic characteristics including robustness, high specificity and flexibility and POROS resins demonstrate exceptionally high dynamic binding capacity, capture efficiency, and resolution over a range of flow rates.These two elements have been brought to create a number of different columns that support quantification of a range of biomolecules:• POROS CaptureSelect LC-kappa and POROS CaptureSelect LC-lambda; for the analysis of human antibodies and antibody fragments containing either a kappa or a lambda light chain;• POROS CaptureSelect IgG-Fc; for the detection of any human IgG subclass (even IgG3) in the presence of IgG from other sources (e.g. bovine);• POROS CaptureSelect IgM; for the analysis of human, mouse and rat IgM; and • POROS CaptureSelect HSA; for the analysis of HSA-fusion proteins (specific for human serum albumin in the presence of albumin from other species).
Efficient purification
The introduction of the POROS CaptureSelect products provides researchers with ready-to-use analytical chromatography columns for high-speed quantification or purification of most biological drug classes including immunoglobulins, fusion proteins, Fab fragments and bispecific antibodies. Supporting quantitation of samples taken directly from samples of cell cultures or fermentation harvests without pre-treatment, the columns are compatible with automatable HPLC systems and can achieve high-speed quantification in less than 3 minutes, making them suitable for the most demanding biomolecule production applications. The affinity resins can also be used for hundreds of cycles, providing a costefficient solution for monitoring biomolecule quantities in cells cultures, fermentation harvests or bioreactors. In addition, method development is streamlined as the columns can be used for all of the required analysis. The purity of the resulting elution product is suitable for subsequent analysis, including mass spectroscopy.
Applications
The unique selectivity of these columns support Process Analytical Technology (PAT) for most biopharmaceutical manufacturing processes and are designed to work at the high flow rates necessary to provide the realtime results demanded in biopharmaceutical development and quality assurance. The most common application is the quantitation of therapeutic proteins throughout the manufacturing process and the current line of products support quantitation of most common biomolecules including antibodies, antibody fragments and HSA fusion proteins. A secondary application is the small-scale purification of immunoglobulins, Fabs and fusion proteins for further downstream analysis in research applications.