Application Notes

Thermal stability buffer screening of therapeutic antibodies

Issue link:

Contents of this Issue


Page 0 of 3

1 Thermal Unfolding of Antibodies Application Note NT-PR-003 Thermal Stability Buffer Screening of Therapeutic Antibodies Lea Martin, Melanie Maschberger and Dennis Breitsprecher NanoTemper Technologies GmbH, Munich, Germany Abstract The development of therapeutic antibodies requires optimal formulations for long-term antibody stability. For this, buffer screening approaches are routinely used, in which the thermal stability of a given antibody in different buffers are tested. The buffers typically vary regarding buffer substances, pH values, salt concentrations and other excipients. The prerequisite for such screening approaches are high measurement precision, low sample consumption, and high throughput. Moreover, it is highly desirable to measure under native conditions, without dilution of the antibody or the requirement to use reporter dyes or other modifications. Here we present nanoDSF, a label-free, native DSF technique, as the method of choice to perform rapid and precise buffer screening projects for therapeutic antibodies during the drug development process. Introduction Monoclonal antibodies have been used for 30 years to treat a number of diseases, and their popularity and clinical application spectrum are growing constantly. Especially the development of novel antibody entities, such as antibody-drug conjugates or bispecific antibodies, further increases the efficacy and specificity of antibodies (Adair et al, 2012; Yang & Ambrogelly, 2014). Since the number of novel antibodies and antibody conjugates increases each year, methods are needed to address the requirements in the pharmaceutical industry for quick and precise screening approaches to identify optimal buffer conditions, or the most stable antibody variants. Here we present a label-free differential scanning fluorimetry approach (nanoDSF) using the Prometheus NT.48 to determine the thermal stability of therapeutic antibodies in a variety of different formulations. Figure 1 Crystal structures of two antibodies The Prometheus NT.48 uses a dual-UV detection system that is specifically designed to monitor shifts in the fluorescence emission of the amino acid tryptophan. In order to determine thermal unfolding transition points, the shift in tryptophan emission (expressed as the ratio of the tryptophan emission at 350 and 330 nm) is plotted versus time and fitted automatically. The high density of data points allows for precise identification and quantification of multiple transition points with unprecedented accuracy. Combined with the minimal sample consumption, speed and ease of use, the Prometheus NT.48 is perfectly suited for antibody formulation screens. Results Two therapeutic monoclonal antibodies (mAbs), mAb1 and mAb2, where subjected to a thermal unfolding formulation screen. For each antibody, 45 different buffer conditions were tested. The

Articles in this issue

view archives of Application Notes - Thermal stability buffer screening of therapeutic antibodies