Refereed journal article or data article (A1)

Sensitive, homogeneous, and label-free protein-probe assay for antibody aggregation and thermal stability studies




List of Authors: Valtonen Salla, Vuorinen Emmiliisa, Eskonen Ville, Malakoutikhah Morteza, Kopra Kari, Härmä Harri

Publisher: TAYLOR & FRANCIS INC

Publication year: 2021

Journal: mAbs

Journal name in source: MABS

Journal acronym: MABS-AUSTIN

Volume number: 13

Issue number: 1

Number of pages: 11

ISSN: 1942-0862

DOI: http://dx.doi.org/10.1080/19420862.2021.1955810

URL: https://doi.org/10.1080/19420862.2021.1955810

Self-archived copy’s web address: https://research.utu.fi/converis/portal/detail/Publication/67270415


Abstract
Protein aggregation is a spontaneous process affected by multiple external and internal properties, such as buffer composition and storage temperature. Aggregation of protein-based drugs can endanger patient safety due, for example, to increased immunogenicity. Aggregation can also inactivate protein drugs and prevent target engagement, and thus regulatory requirements are strict regarding drug stability monitoring during manufacturing and storage. Many of the current technologies for aggregation monitoring are time- and material-consuming and require specific instruments and expertise. These types of assays are not only expensive, but also unsuitable for larger sample panels. Here we report a label-free time-resolved luminescence-based method using an external Eu3+-conjugated probe for the simple and fast detection of protein stability and aggregation. We focused on monitoring the properties of IgG, which is a common format for biological drugs. The Protein-Probe assay enables IgG aggregation detection with a simple single-well mix-and-measure assay performed at room temperature. Further information can be obtained in a thermal ramping, where IgG thermal stability is monitored. We showed that with the Protein-Probe, trastuzumab aggregation was detected already after 18 hours of storage at 60 degrees C, 4 to 8 days earlier compared to SYPRO Orange- and UV250-based assays, respectively. The ultra-high sensitivity of less than 0.1% IgG aggregates enables the Protein-Probe to reduce assay time and material consumption compared to existing techniques.

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Last updated on 2022-29-06 at 08:39