Protocol for effective surface passivation for single-molecule studies of chromatin and topoisomerase II.

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Howard Hughes Medical Institute, Cornell University, Ithaca, NY 14853, USA; Physics Department & LASSP, Cornell University, Ithaca, NY 14853, USA. Electronic address:

Published: December 2024

For single-molecule studies requiring surface anchoring of biomolecules, poorly passivated surfaces can result in alterations of biomolecule structure and function that lead to artifacts. Here, we present a surface passivation assay for single-molecule studies of chromatin and topoisomerase II. We detail steps for preparing a nucleosome array and hydrophobic nitrocellulose-coated flow cell. We then describe procedures for chromatin stretching with an angular optical trap (AOT) and performing a chromatin-topoisomerase experiment. This method is cost effective and potentially applicable to other biomolecules. For complete details on the use and execution of this protocol, please refer to Le et al. .

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http://dx.doi.org/10.1016/j.xpro.2024.103500DOI Listing

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