A novel approach for oxidation analysis of therapeutic proteins.

Citation:

Iva Turyan, Khatwani, Nikhil , Sosic, Zoran , Jayawickreme, Shiranthi , and Mandler, Daniel. . 2016. “A Novel Approach For Oxidation Analysis Of Therapeutic Proteins.”. Anal. Biochem.analytical Biochemistry, 494, Pp. 108 - 113.

Abstract:

Measuring and monitoring of protein oxidn. modifications is important for biopharmaceutical process development and stability assessment during long-term storage. Currently available methods for biomols. oxidn. anal. use time-consuming peptide mapping anal. Therefore, it is desirable to develop high-throughput methods for advanced process control of protein oxidn. Here, we present a novel approach by which oxidative protein modifications are monitored by an indirect potentiometric method. The method is based on adding an electron mediator, which enhances electron transfer (ET) between all redox species and the electrode surface. Specifically, the procedure involves measuring the sharp change in the open circuit potential (OCP) for the mediator system (redox couple) as a result of its interaction with the oxidized protein species in the soln. Application of Pt and Ag/AgCl microelectrodes allowed for a high-sensitivity protein oxidn. anal. We found that the Ru(NH3)2+/3+6 redox couple is suitable for measuring the total oxidn. of a wide range of therapeutic proteins between 1.1 and 13.6%. Accuracy detd. by comparing with the known percentage oxidn. of the ref. std. showed that percentage oxidn. detd. for each sample was within ±20% of the expected percentage oxidn. detd. by mass spectrometry. [on SciFinder(R)]