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Standard operation procedure for switchSENSE DRX systems

A Correction to this article was published on 29 May 2021

This article has been updated

Abstract

There is currently a large panel of technologies available to address molecular interactions in vitro. Each technology presents individual advantages and drawbacks, and it becomes challenging to choose which technology will be best suited for a molecular interaction of interest. Approaches can be broadly categorized as either microfluidic surface-bound methods (such as Surface Plasmon Resonance (SPR) or switchSENSE) or in-solution methods (such as Isothermal Titration Calorimetry (ITC) or MicroScale Thermophoresis (MST)). In-solution methods are advantageous in terms of sample preparation and ease of use as none of the binding partners are subjected to immobilization. On the other hand, surface-based techniques require only small amounts of immobilized interaction partner and provide off-rate characterization as unbound analytes can be removed from the surface to observe analyte dissociation. Here, a standard operating procedure (SOP) for the switchSENSE method is presented, which aims to guide new users through the process of a switchSENSE measurement, covering sample preparation, instrument and biochip handling as well as data acquisition and analysis. This guide will help researchers decide whether switchSENSE is the right method for their application as well as supporting novice users to get the most information out of a switchSENSE measurement. switchSENSE technology offers the unique advantage of a controlled DNA-based ligand surface within a microfluidic channel which allows the user to distribute specifically up to two different ligand molecules on the surface at a customized density and ratio. The technology offers multi-parameter characterization of binding kinetics, affinity, enzymatic activity, and changes in protein conformation.

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Acknowledgements

We thank ARBRE-MOBIEU network for proposing us to publish this SOP.

Funding

ARC program (SLS220120605310), French Infrastructure for Integrated Structural Biology (FRISBI) ANR-10- INSB-05, ANR-18-CE44-0008, ANR-20-CE11-0026, INCA 2016-1-PL BIO-11 and INCA SLX4 INCA 2016-159.

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Correspondence to Paloma Fernández Varela.

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Special issue: COST Action CA15126, MOBIEU: Between atom and cell.

The original online version of this article was revised: Section name “Covalent coupling” repeated twice by mistake. Now the second section name updated as “Capture method”.

Appendices

Appendix

1. DRX buffer compatibility

Ingredient Concentration
Ethylenediaminetetraacetic acid (EDTA) 0–5 mM
Dimethyl sulfoxide (DMSO) 0–5% v/v
Polysorbate 20 (Tween20®) 0–1% v/v
Triton® X-100, Nonidet® P40, Tergitol® NP-40 0–1% v/v
Glycerol 0–70 wt%
Urea 0–8 M
Guanidine hydrochloride 0–6 M
tris(2-carboxyethyl)phosphine (TCEP) 0–0.1 mM (dynamic mode), 0–1 mM (static mode)
β-mercapto-ethanol, dithiothreitol (DTT) 0–10 µM (dynamic mode), 0–1 mM (static mode)
n-Dodecyl-beta-d-maltoside (DDM) 0–20 mM
Human serum albumin (HSA) 0–0.5 mM

2. DNA sequences on a DRX biochip

48 nt nanolever:

NL-A48: 5′-TAG TGC TGT AGG AGA ATA TAC GGG CTG CTC GTG TTG ACA AGT ACT GAT-3′.

NL-B48: 5′-TAG TCG TAA GCT GAT ATG GCT GAT TAG TCG GAA GCA TCG AAC GCT GAT-3′.

96 nt nanolever:

NL-A96: 5′-TAG TGG AAC TGG TTT GAC TTG AAC GTA GGA AAA CAC TTG GCT TAC AGG GGG ATT GCT CCG TAG GGT AGG CTC TTG ATT ACA CGA CAC GGA ACT GAT-3′.

NL-B96: 5′-TAG TCG TAA GCT GAT ATG GCT GAT TAG TCG GAA GCA TCG AAC GCT GAT TAG TTA CAG TAC CTC CGA GAG CAA GTA GGG CAC CCT GTA GTT CCT GAT-3′.

80 nt nanolever (enzyme biochip):

NL-T80: 5′-TCT CCA CGC CGA CTC TAA TGC TGG TTA CAT CAC TCT TTA TCT CGT TAC ATC ACT CTT TAT GCT CAC ACT TCA GGT TAC TC-3′.

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Müller-Landau, H., Varela, P.F. Standard operation procedure for switchSENSE DRX systems. Eur Biophys J 50, 389–400 (2021). https://doi.org/10.1007/s00249-021-01519-3

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Keywords

  • Biophysical analysis
  • Biomolecular interactions
  • Surface biosensors
  • DNA-encoded addressing
  • SOP
  • SwitchSENSE