Rapid and Straightforward Quantification of Recombinant Proteins Using Fluorescence Polarization

摘要： Biotechnology enables the production and commercialization of recombinant proteins for diagnostic, therapeutic and research applications. For instance, monoclonal antibodies nowadays represent a fast growing market that accounts for almost half of the biopharmaceutical compounds business with $75 billion turnover in 2013. Quantification of proteins is an important task in order to optimize cell culture conditions and ensure that every step of the production process is in line with the specifications. For therapeutics, precise and reliable dosage is mandatory to avoid adverse effects for the patients. Current analytical techniques such as ELISA or HPLC are time-consuming and often require purification prior to analysis. In this context, there is a strong demand for a technology that would be to protein quantification what a balance is to weight measurement: rapid, precise, accurate, robust, and straightforward. To this effect, we have developed an assay for the rapid quantification of recombinant proteins. It is based on fluorescence polarization and requires little or no pretreatment of samples. Fluorescence anisotropy of a small specific fluorescent ligand increased upon binding to its recombinant target protein, e.g. a monoclonal antibody, enabling quantification. The assay did not require prior separation of biotechnological cultures making it as straightforward as weighing using a balance. Monitoring of the production of a representative His-tagged protein during E.coli fermentation was obtained by regular sampling and assaying using the fluorescence polarization assay.