title: Orthogonal high-throughput thermal scanning method for rank ordering protein formulations.
abstract: A high-throughput thermal-scanning method to rank-order formulation conditions for therapeutic proteins is described. Apparent transition temperatures for unfolding and aggregation of four different proteins are determined using the dyes SYPRO Orange and thioflavin T (ThT) under a variety of buffer conditions. The results indicate that the ThT-based thermal scanning method offers several advantages over the previously described SYPRO Orange-based thermal scanning and allows rapid rank ordering of solution conditions relevant toward long-term storage of therapeutic molecules. The method is also amenable to high protein concentration and does not require sample dilution or extensive preparation. Additionally, this parallel use of SYPRO Orange and ThT can be readily applied to the screening of mutants for their unfolding and aggregation propensities.
authors.affiliation: Drug Product Science and Technology, Bristol-Myers Squibb, 1 Squibb Drive, New Brunswick, New Jersey, 08903, USA, vishal.nashine@bms.com.
authors.forename: Vishal C
authors.initials: VC
authors.lastname: Nashine
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Kroetsch
authors.forename: Erinc
authors.initials: E
authors.lastname: Sahin
authors.forename: Rong
authors.initials: R
authors.lastname: Zhou
authors.forename: Monica L
authors.initials: ML
authors.lastname: Adams
mesh_terms: D000465:Algorithms; D000911:Antibodies, Monoclonal / Q000008:administration & dosage / Q000737:chemistry; D052160:Benzothiazoles; D002021:Buffers; D002626:Chemistry, Pharmaceutical / Q000379:methods*; D002919:Chymotrypsinogen; D057166:High-Throughput Screening Assays / Q000379:methods*; D010455:Peptides / Q000008:administration & dosage / Q000737:chemistry; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry*; D060888:Real-Time Polymerase Chain Reaction; D012995:Solubility; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013844:Thiazoles
chemical_list: D000911:Antibodies, Monoclonal; D052160:Benzothiazoles; D002021:Buffers; D010455:Peptides; D011506:Proteins; D013844:Thiazoles; C009462:thioflavin T; D002919:Chymotrypsinogen