Determination of binding constant of malE-A1(Cys) for MBP by microscale thermophoresis


Overview
Abstract
The dissociation constant (Kd) of the interaction between malE-A1(Cys) and MBP was measured by microscale thermophoresis. malE-A1(Cys) was conjugated to an Alexafluor 568 and held at 5 nM while the concentration of MBP was varied from 5 µM to 0.153 nM and the migration of the fluorescent protein was measured upon local heating using a Monolith NT.115 Pico with 80% Laser Power, 3% LED power, at 25C.
Type
  • Biophysical Constant Measurement
Target
Scores
Outcome
Successful
Signal
Acceptable
Results
Figure

Image 

MST determination of dissociation constant for nanoCLAMP malE-A1 and MBP

Legend 

Analysis of the interaction between malE-A1(Cys)-AF and MBP by microscale thermophoresis. Binding of malE-A1(Cys)-AF to MBP was quantified in PBS-T (0.05% Tween). malE-A1(Cys)-AF was held at 5 nM while the concentration of MBP was varied from 5 µM to 0.153 nM while the migration of the fluorescent protein was measured upon local heating using a Monolith NT.115 Pico with 80% Laser Power, 3% LED power, at 25C.