An Effective Surface Passivation Assay for Single-Molecule Studies of Chromatin and Topoisomerase II.

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For complete details on the use and execution of this protocol, please refer to Le et al. (2019).

SUMMARY

A.For single-molecule studies requiring surface anchoring of biomolecules, a poorly passivated surface can result in alterations of biomolecule structure and function that can result in artifacts. This protocol describes surface passivation and sample chamber preparation for mechanical manipulation of chromatin fibers and characterization of topoisomerase II activity in physiological buffer conditions. The method employs enhanced surface hydrophobicity and purified blocking proteins to reduce non-specific surface adsorption. This method is accessible, cost-effective, and potentially widely applicable to other biomolecules.For a complete list of publications that employ this protocol, see the paper references.