High Throughput Analytics for Protein Purification Process Development

medicilon
medicilon
Jul 28, 2017 · 4 min read

Medicilon’s protein scientists have been working on protein expression and purification for many years. We can start your project even you have nothing in hand but the name of your protein. In Medicilon’s laboratories, protein purification is performed in scales from micrograms and milligrams. All Protein Purification Services start with the analysis of physico-chemical and biological properties of a target protein resulting in the development of tailored procedures for its extraction, purification and characterization. Email:marketing@medicilon.com.cn web:www.medicilon.com

1.1 Preface — From Quack to Exhaustive Regulation

The business of manufacturing and selling substances with the purpose to bring healing or relief to individuals suffering from illness has undergone vast changes. Until the end of the 19th century, most drugs were prepared in pharmacies and there was no regulation or restriction of claimed efficacy or safety of the drugs. Around this time pharmacies began to develop from small manufacturers into pharmaceutical companies and a new industry was born. Insulin and antibiotics were the first bio-pharmaceutical blockbusters and many of today’s large pharmaceutical companies originated around this business. Soon came the first major scandals and as a reaction to each scandal came new laws and regulations. The early legislation mostly aimed to secure a correlation between label claim and actual content. Since then, continuous addition of laws and guidelines have made the pharmaceutical industry subject to very detailed regulation, both with regard to development, testing, production and admission/licensing. Manufacturing is required to be at state of the art and numerous guidelines from different agencies and organizations have been issued describing what the current state of the art is. State of the art protein purification processes are required to be based on identified product quality attributes and their detailed and documented linkage to process parameters. At the stage of manufacturing a detailed process control strategy must be in place. The developed control strategy is aimed at ensuring very high as well as highly reproducible quality based on different control elements and the knowledge of the links between process parameters and product quality. A control strategy consists of numerous elements including process control elements such as Procedural Controls and Process Parameter Controls and testing control elements such as In-Process Testing, Specifications, and Process Monitoring.

1.2 Therapeutic Proteins

Therapeutic proteins can be divided into three groups: therapeutic proteins with enzymatic or regulatory activity (for replacement therapy e.g. growth hormone, insulin, blood factors), therapeutic proteins with special target activity (e.g. monoclonals antibodies), protein vaccines (against pathogens or treatment of cancer and autoimmune diseases). Together they have an enormous marked value and today it is impossible to imagine a pharmaceutical market without therapeutic proteins. Compared to small molecules therapeutics, protein therapeutics are challenging to manufacture and due to their size and complexity also elaborate to analyze and characterize. Due to the biologic origin and the complexity of the therapeutic proteins, a final drug product administered to a patient will never consist only of numerous identical protein molecules in an appropriate matrix. Much rather it is a well defined population of protein species: Some related to the main product such as charge variants, aggregates, and glycoforms and some which are a reminiscence of the biological host in form of host cell proteins. Purification of therapeutic proteins from their biological source must be performed under gentle conditions in order to preserve the biological function and mostly rely on a set of unit operations including but not limited to filtration, chromatography, centrifugation, precipitation, and crystallization. The introduction of high throughput techniques has modernized the field of protein purification process development over the past decade. While high throughput techniques such as automation, parallelization, and scale miniaturization has long been applied for screenings in the field of bioprocess development, this was not the case for purification process development at the turn of the millennium. The main driving force in this progress is the need for very short purification process development times and the requirement for a deeper knowledge and understanding of the processes. 1.3 The Dawn of Modern Protein Purification Process Development As it has been for decades, liquid chromatography is still the central unit operation for biomolecule purification. When developing a purification process step based on liquid chromatography, there is a vast number of solid and liquid phases to choose from and the total number of possible parameters influencing any given separation process exceeds by far what can be tested in a sequential packed column mode. Hence, it is not a surprise that the first reported application of high throughput techniques for protein purification process development was related to chromatography. This first report described a parallel micro scale experimentation in micro titer plate format with automated liquid handling and was published by Thiemann et al. in 2004. The authors suggested that due to high number of experiments necessary for a systematic variation of the relevant parameters in the optimization of chromatographic process step, automation of the liquid handling would bring substantial benefit with regard to experimental throughput. Where this initial work published by Thiemann et al. was aimed at the optimization of single purification step, Rege et al. in 2006 proposed a micro plate based screening method to identify the best sequence of chromatography steps and hence named their approach high throughput process development. Hereby they had named a new field within in protein purification research and became the herald of a new era in protein purification process development.

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