Application Notes

Protein Labeling – Improved quantitation of biomolecular interactions by MicroScale Thermophoresis using the RED-NHS 2nd Generation labeling kit

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APPLICATION NOTE Monolith is a trademark of NanoTemper Technologies GmbH, Munich, Germany. NanoTemper is a registered trademark of NanoTemper Technologies GmbH, Munich, Germany. ©2018 NanoTemper Technologies, Inc. South San Francisco, CA, USA. All Rights Reserved. NT-MO-033 nanotempertech.com MST binding experiments Samples were centrifuged at 14 000 x g for 10 min at 4 °C, then loaded into Monolith NT.115 Premium Capillaries (cat# MO-K025). Experiments with p38α were carried out using a Monolith NT.115 system at LED settings of 20% (RED-NHS) and 40% (RED-NHS 2nd Generation) and high MST power. Experiments with TEM1 were carried out using a Monolith NT.115Pico system at LED settings of 20% (RED-NHS) and 5% (RED-NHS 2nd Generation) and medium MST power. Experiments with Hsp90 were carried out using a Monolith NT.115 system at LED settings of 40% (RED-NHS) and 40% (RED-NHS 2nd Generation) and medium MST power. All Monolith systems were running MO.Control so ware v1.6, and data were analyzed at default MST-on times using MO.Affinity Analysis so ware v2.3. All samples were prepared and measured in triplicate. References 1. Duhr, S., & Braun, D. (2006). Why molecules move along a temperature gradient. Proc Natl Acad Sci U S A, 103(52), 19678– 19682. 2 P. Baaske, C. Wienken, P. Reineck, S. Duhr, D. Braun (2010). Optical Thermophoresis for Quantifying the Buffer Dependence of Aptamer Binding. Angew Chem Int Ed Engl, 2238-2241: 3 Underwood, D. C. et al (2000). SB 239063, a potent p38 MAP kinase inhibitor, reduces inflammatory cytokine production, airways eosinophil infiltration, and persistence. J. Pharmacol. Exp. Ther. 293, 281–288.

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