Methods for determining ligand binding to a target protein using a thermal shift assay

US9523693B2 · kind B2 · utility

Assignee

• Biotarget Engagement Interest Group AB · SE

Inventors

• Pär Nordlund · Singapore, SG
• Daniel Martinez Molina · Stockholm, SE
• Thomas Lundback · Nacka, SE

Key dates

• Filing date: Jan 16, 2015
• Grant date: Dec 20, 2016
• Expiry date: Feb 14, 2035

Classification

• Technology area (CPC G): Physics
• CPC primary: G01N33/5306
• WIPO field: Measurement
• WIPO sector: Instruments

Abstract

The present invention concerns a method of determining whether a non-purified sample contains a target protein bound to a ligand of interest comprising the steps of a) exposing the non-purified sample to a temperature which is capable of causing or enhancing precipitation of the unbound target protein to a greater extent than it is capable of causing or enhancing precipitation of the target protein bound to the ligand; and b) analyzing said sample for the presence of soluble or native target protein using two or more affinity reagents capable of binding to said soluble or native target protein with a higher affinity than to an unfolded and/or insoluble form of said target protein. The invention particularly concerns the use of two affinity reagents (e.g. antibodies) which are capable of distinguishing between soluble or native, and unfolded and/or insoluble forms of a target protein and whose detection e.g. by FRET based technology, allows the performance of the method without a separation step.