Primed track: reliable volumetric single-cell tracking and lineage tracing of living specimen with dual-labeling approaches
File(s)
Published version
Author(s)
Welling, Maaike
Kalyviotis, Konstantinos
Pantazis, Periklis
Type
Journal Article
Abstract
Mammalian embryonic development starts with a single fertilized zygote that develops into
a blastocyst embryo consisting of three cell types that evolve into either embryonic or extra-embryonic
tissues. Lineage tracing of these cells can provide important information about the molecular and cellular
dynamics contributing to fate allocation during early development. While global labeling techniques allow
for visualization of all cells at the same time, lineage tracing of cells over several divisions can become
complicated due to embryo movement and rotation as well as increasing cell densities. Here, we use
green-to-red photoconvertible proteins for both global and sparse labeling of cells of interest in the
developing murine embryo. We use primed conversion to achieve precise photoconversion of single
nuclei in 4-cell stage embryos followed by volumetric live imaging to capture development up to the
blastocyst stage. We developed an image analysis pipeline, called primed Track, that uses the dual
labeling strategy for both straightforward segmentation and registration of all cells in the embryo as well
as correction of rotational and spatial drift. Together, this strategy allows for reliable and fast tracking
and lineage tracing of individual cells, even over increased imaging time intervals that result in a major
reduction in data volume, all essential conditions for volumetric long-term imaging techniques
a blastocyst embryo consisting of three cell types that evolve into either embryonic or extra-embryonic
tissues. Lineage tracing of these cells can provide important information about the molecular and cellular
dynamics contributing to fate allocation during early development. While global labeling techniques allow
for visualization of all cells at the same time, lineage tracing of cells over several divisions can become
complicated due to embryo movement and rotation as well as increasing cell densities. Here, we use
green-to-red photoconvertible proteins for both global and sparse labeling of cells of interest in the
developing murine embryo. We use primed conversion to achieve precise photoconversion of single
nuclei in 4-cell stage embryos followed by volumetric live imaging to capture development up to the
blastocyst stage. We developed an image analysis pipeline, called primed Track, that uses the dual
labeling strategy for both straightforward segmentation and registration of all cells in the embryo as well
as correction of rotational and spatial drift. Together, this strategy allows for reliable and fast tracking
and lineage tracing of individual cells, even over increased imaging time intervals that result in a major
reduction in data volume, all essential conditions for volumetric long-term imaging techniques
Date Issued
2020-06-05
Date Acceptance
2020-06-01
Citation
Bio-protocol, 2020, 10 (11), pp.1-10
ISSN
2331-8325
Publisher
Bio-Protocol
Start Page
1
End Page
10
Journal / Book Title
Bio-protocol
Volume
10
Issue
11
Copyright Statement
© 2020 Welling et al.
This article is distributed under the terms of the Creative Commons Attribution License (CC BY 4.0)
This article is distributed under the terms of the Creative Commons Attribution License (CC BY 4.0)
License URL
Sponsor
The Royal Society
Identifier
http://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000583105200014&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=1ba7043ffcc86c417c072aa74d649202
Grant Number
WRM\FT\180010
Subjects
Science & Technology
Life Sciences & Biomedicine
Biology
Life Sciences & Biomedicine - Other Topics
Pre-implantation embryos
Mouse
Lineage tracing
Tracking
Primed conversion
Photoconvertible proteins
Live imaging
Dual color fluorescent labeling
IN-VIVO
FLUORESCENT PROTEINS
BLUE
CONVERSION
MECHANISM
Publication Status
Published
Article Number
ARTN e3645
Date Publish Online
2020-06-05