Literature/202212301234 biggest challenge of single-molecule methods is non-specific binding

• Source: [@walt2013Optical Methods for Single Molecule Detection and Analysis]
• Tags: #single-molecule #non-specific-binding

The biggest problem of single-molecule detection methods that rely on labeling specific proteins is that there will always be a "background floor" of labels that bound to the wrong protein or to the substrate.

Non-specific binding (NSB) can be worked around by coating the substrates in such a way that there can be a washing step that limits the amount of background. However, it still poses the biggest limitation for single-molecule methods.

Pushing the detection limit down to single-molecules (see: why pushing sensitivity of assays) is very promising, but must be done carefully. The interpretation of the results must be done carefully.

This point is what makes the label-free methods such a relevant tool to characterize samples at low concentrations, focusing on the intrinsic properties of the analytes.