Protocol for machine-learning-based 3D image analysis of nuclear envelope tubules in cultured cells

Anisha Hundal; Defne Urman; Mia Stanic; Razqallah Hakem; Karim Mekhail

doi:10.1016/j.xpro.2024.103214

Protocol for machine-learning-based 3D image analysis of nuclear envelope tubules in cultured cells

STAR Protoc. 2024 Sep 20;5(3):103214. doi: 10.1016/j.xpro.2024.103214. Epub 2024 Jul 31.

Authors

Anisha Hundal¹, Defne Urman², Mia Stanic², Razqallah Hakem³, Karim Mekhail⁴

Affiliations

¹ Department of Laboratory Medicine and Pathobiology, Temerty Faculty of Medicine, University of Toronto, Toronto, ON, Canada; Princess Margaret Cancer Research Centre, University Health Network, Toronto, ON, Canada.
² Department of Laboratory Medicine and Pathobiology, Temerty Faculty of Medicine, University of Toronto, Toronto, ON, Canada.
³ Department of Laboratory Medicine and Pathobiology, Temerty Faculty of Medicine, University of Toronto, Toronto, ON, Canada; Princess Margaret Cancer Research Centre, University Health Network, Toronto, ON, Canada; Department of Medical Biophysics, Temerty Faculty of Medicine, University of Toronto, Toronto, ON, Canada. Electronic address: razq.hakem@uhn.ca.
⁴ Department of Laboratory Medicine and Pathobiology, Temerty Faculty of Medicine, University of Toronto, Toronto, ON, Canada; Temerty Centre for AI Research and Education in Medicine, University of Toronto, Toronto, ON, Canada; College of New Scholars, Artists and Scientists, The Royal Society of Canada, Ottawa, ON, Canada. Electronic address: karim.mekhail@utoronto.ca.

Abstract

The nuclear envelope can form complex structures in physiological and pathological contexts. Current approaches to quantify nuclear envelope structures can be time-consuming or inaccurate. Here, we present a protocol to measure nuclear envelope tubules induced by DNA double-strand breaks using a mid-throughput approach. We describe steps for the induction of these nuclear envelope structures and 3D image analysis using machine-learning-based image segmentation. This protocol can be applied to analyze various nuclear envelope structures in contexts beyond DNA repair. For complete details on the use and execution of this protocol, please refer to Shokrollahi et al.¹.

Keywords: Cell Biology; Cell culture; Genetics; Microscopy; Molecular/Chemical Probes; Single Cell.

MeSH terms

Cells, Cultured
DNA Breaks, Double-Stranded
Humans
Image Processing, Computer-Assisted / methods
Imaging, Three-Dimensional* / methods
Machine Learning*
Nuclear Envelope* / metabolism