Opto-magnetic capture of individual cells based on visual phenotypes
- Maisonneuve-Rosemont Hospital, Canada
Abstract
The ability to isolate rare live cells within a heterogeneous population based solely on visual criteria remains technically challenging, due largely to limitations imposed by existing sorting technologies. Here we present a new method that permits labeling cells of interest by attaching streptavidin-coated magnetic beads to their membranes using the lasers of a confocal microscope. A simple magnet allows highly-specific isolation of the labeled cells, which then remain viable and proliferate normally. As proof of principle, we tagged, isolated, and expanded individual cells based on three biologically-relevant visual characteristics: i) presence of multiple nuclei, ii) accumulation of lipid vesicles, and iii) ability to resolve ionizing radiation-induced DNA damage foci. Our method constitutes a rapid, efficient, and cost-effective approach for isolation and subsequent characterization of rare cells based on observable traits such as movement, shape, or location, which in turn can generate novel mechanistic insights into important biological processes.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files
Article and author information
Author details
Jean François Lemay
Santiago Costantino
Funding
Canadian Institutes of Health Research
- El Bachir Affar
- Elliot Drobetsky
- Hugo Wurtele
Natural Sciences and Engineering Research Council of Canada
- El Bachir Affar
- Elliot Drobetsky
- Hugo Wurtele
- Santiago Costantino
Genome Canada
- Santiago Costantino
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2019, Binan et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 2,215
- views
-
- 328
- downloads
-
- 13
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Opto-magnetic capture of individual cells based on visual phenotypes
eLife 8:e45239.
https://doi.org/10.7554/eLife.45239
Further reading
-
- Cancer Biology
Why does a normal cell possibly harboring genetic mutations in oncogene or tumor suppressor genes becomes malignant and develops a tumor is a subject of intense debate. Various theories have been proposed but their experimental test has been hampered by the unpredictable and improbable malignant transformation of single cells. Here, using an optogenetic approach we permanently turn on an oncogene (KRASG12V) in a single cell of a zebrafish brain that, only in synergy with the transient co-activation of a reprogramming factor (VENTX/NANOG/OCT4), undergoes a deterministic malignant transition and robustly and reproducibly develops within 6 days into a full-blown tumor. The controlled way in which a single cell can thus be manipulated to give rise to cancer lends support to the ‘ground state theory of cancer initiation’ through ‘short-range dispersal’ of the first malignant cells preceding tumor growth.
-
- Cancer Biology
Most human pancreatic ductal adenocarcinoma (PDAC) are not infiltrated with cytotoxic T cells and are highly resistant to immunotherapy. Over 90% of PDAC have oncogenic KRAS mutations, and phosphoinositide 3-kinases (PI3Ks) are direct effectors of KRAS. Our previous study demonstrated that ablation of Pik3ca in KPC (KrasG12D; Trp53R172H; Pdx1-Cre) pancreatic cancer cells induced host T cells to infiltrate and completely eliminate the tumors in a syngeneic orthotopic implantation mouse model. Now, we show that implantation of Pik3ca−/− KPC (named αKO) cancer cells induces clonal enrichment of cytotoxic T cells infiltrating the pancreatic tumors. To identify potential molecules that can regulate the activity of these anti-tumor T cells, we conducted an in vivo genome-wide gene-deletion screen using αKO cells implanted in the mouse pancreas. The result shows that deletion of propionyl-CoA carboxylase subunit B gene (Pccb) in αKO cells (named p-αKO) leads to immune evasion, tumor progression, and death of host mice. Surprisingly, p-αKO tumors are still infiltrated with clonally enriched CD8+ T cells but they are inactive against tumor cells. However, blockade of PD-L1/PD1 interaction reactivated these clonally enriched T cells infiltrating p-αKO tumors, leading to slower tumor progression and improve survival of host mice. These results indicate that Pccb can modulate the activity of cytotoxic T cells infiltrating some pancreatic cancers and this understanding may lead to improvement in immunotherapy for this difficult-to-treat cancer.
