Protocol for in vitro co-culture, proliferation, and cell cycle analyses of patient-derived leukemia cells

Jessica Parker; Sean Hockney; Carly Knill; David McDonald; Andrew Filby; Deepali Pal

doi:10.1016/j.xpro.2024.103202

Protocol for in vitro co-culture, proliferation, and cell cycle analyses of patient-derived leukemia cells

STAR Protoc. 2024 Sep 20;5(3):103202. doi: 10.1016/j.xpro.2024.103202. Epub 2024 Jul 20.

Authors

Jessica Parker¹, Sean Hockney¹, Carly Knill², David McDonald², Andrew Filby², Deepali Pal³

Affiliations

¹ Department of Applied Sciences, Northumbria University, Newcastle upon Tyne NE1 8ST, UK.
² Flow Cytometry Core Facility (FCCF), Newcastle University, Newcastle upon Tyne NE1 7RU, UK; Biosciences Institute, Innovation, Methodology and Application (IMA) Research Theme, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK.
³ Department of Applied Sciences, Northumbria University, Newcastle upon Tyne NE1 8ST, UK; Wolfson Childhood Cancer Research Centre, Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, Herschel Building Level 6, Brewery Lane, Newcastle upon Tyne NE1 7RU, UK; School of Cellular and Molecular Medicine, University of Bristol, Biomedical Sciences Building, University Walk, Bristol BS8 1TD, UK. Electronic address: deepali.pal@bristol.ac.uk.

Abstract

Leukemia niche impacts quiescence; however, culturing patient-derived samples ex vivo is technically challenging. Here, we present a protocol for in vitro co-culture of patient-derived xenograft acute lymphoblastic leukemia (PDX-ALL) cells with human mesenchymal stem cells (MSCs). We describe steps for labeling PDX-ALL cells with CellTrace Violet dye to demonstrate MSC-primed PDX-ALL cycling. We then detail procedures to identify MSC-primed G0/quiescent PDX-ALL cells via Hoechst-33342/Pyronin Y live cell cycle analysis. For complete details on the use and execution of this protocol, please refer to Pal et al.¹^,².

Keywords: Cell Differentiation; Cell-based Assays; Stem Cells.

MeSH terms

Animals
Cell Cycle* / physiology
Cell Proliferation* / physiology
Coculture Techniques* / methods
Humans
Mesenchymal Stem Cells* / cytology
Mice
Precursor Cell Lymphoblastic Leukemia-Lymphoma / pathology