Overview of the reporter genes and reporter mouse models

Li, Shun; Chen, Lixiang; Peng, Xinjun; Wang, Chao; Qin, Boyin; Tan, Dan; Han, Chengxiao; Yang, Hua; Ren, Xu; Liu, Fang; Xu, Chunhua; Zhou, Xiaohui

doi:10.1002/ame2.12008

Cited by 41 publications

(24 citation statements)

References 77 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In the case of animal studies, it is also very useful to be able to integrate antibody staining with imaging of fluorescent marker proteins expressed by engineered cells transferred into animals or expressed in situ. We therefore next investigated whether the IBEX method could be used to image tissues from animals expressing fluorescent proteins (25). To this end, high-dimensional imaging was performed on thymic tissues from transgenic animals expressing the following fluorescent proteins (FPs): cyan FP (CFP), green FP (GFP), yellow FP (YFP), and red FP (RFP) (26).…”

Section: Ibex Builds and Improves Upon Existing Iterative Imaging Techniquesmentioning

confidence: 99%

IBEX: A versatile multiplex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues

Radtke

Kandov

Lowekamp

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

141

142

View full text Add to dashboard Cite

The diverse composition of mammalian tissues poses challenges for understanding the cell–cell interactions required for organ homeostasis and how spatial relationships are perturbed during disease. Existing methods such as single-cell genomics, lacking a spatial context, and traditional immunofluorescence, capturing only two to six molecular features, cannot resolve these issues. Imaging technologies have been developed to address these problems, but each possesses limitations that constrain widespread use. Here we report a method that overcomes major impediments to highly multiplex tissue imaging. “Iterative bleaching extends multiplexity” (IBEX) uses an iterative staining and chemical bleaching method to enable high-resolution imaging of >65 parameters in the same tissue section without physical degradation. IBEX can be employed with various types of conventional microscopes and permits use of both commercially available and user-generated antibodies in an “open” system to allow easy adjustment of staining panels based on ongoing marker discovery efforts. We show how IBEX can also be used with amplified staining methods for imaging strongly fixed tissues with limited epitope retention and with oligonucleotide-based staining, allowing potential cross-referencing between flow cytometry, cellular indexing of transcriptomes and epitopes by sequencing, and IBEX analysis of the same tissue. To facilitate data processing, we provide an open-source platform for automated registration of iterative images. IBEX thus represents a technology that can be rapidly integrated into most current laboratory workflows to achieve high-content imaging to reveal the complex cellular landscape of diverse organs and tissues.

show abstract

Section: Ibex Builds and Improves Upon Existing Iterative Imaging Techniquesmentioning

confidence: 99%

IBEX: A versatile multiplex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues

Radtke

Kandov

Lowekamp

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

141

142

View full text Add to dashboard Cite

show abstract

“…Knocking Cre into the gene locus instead of following the more conventional strategy of inserting a specific fluorescent reporter [ 14 ] presents several advantages. Firstly , it allows for greater flexibility in the choice of reporter.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of TH-Cre knock-in cell lines for detection and specific targeting of stem cell-derived dopaminergic neurons

Fiorenzano

Birtele

Wahlestedt

et al. 2021

Heliyon

View full text Add to dashboard Cite

The focal and progressive degeneration of dopaminergic (DA) neurons in ventral midbrain has made Parkinson's disease (PD) a particularly interesting target of cell-based therapies. However, ethical issues and limited tissue availability have so far hindered the widespread use of human fetal tissue in cell-replacement therapy. DA neurons derived from human pluripotent stem cells (hPSCs) offer unprecedented opportunities to access a renewable source of cells suitable for PD therapeutic applications. To better understand the development and functional properties of stem-cell derived DA neurons, we generated targeted hPSC lines with the gene coding for Cre recombinase knocked into the TH locus. When combined with flexed GFP, they serve as reporter cell lines able to identify and isolate TH þ neurons in vitro and after transplantation in vivo. These TH-Cre lines provide a valuable genetic tool to manipulate DA neurons useful for the design of more precise DA differentiation protocols and the study of these cells after transplantation in pre-clinical animal models of PD.

show abstract

“…In the case of animal studies, it is also very useful to be able to integrate antibody staining with imaging of fluorescent marker proteins expressed by engineered cells transferred into animals or expressed in situ. We therefore next investigated whether the IBEX method could be used to image tissues from animals expressing fluorescent proteins (25). To this end, high dimensional imaging was performed on thymic tissues from transgenic animals expressing the following fluorescent proteins (FPs): cyan (CFP), green (GFP), yellow (YFP), and red (RFP) (26).…”

Section: Ibex Is a Versatile Imaging Methodsmentioning

confidence: 99%

IBEX – A versatile multi-plex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues

Radtke

Kandov

Lowekamp³

et al. 2020

Preprint

View full text Add to dashboard Cite

The diverse composition of mammalian tissues poses challenges for understanding the cell-cell interactions required for organ homeostasis and how spatial relationships are perturbed during disease. Existing methods such as single-cell genomics, lacking a spatial context, and traditional immunofluorescence, capturing only 2-6 molecular features, cannot resolve these issues. Imaging technologies have been developed to address these problems, but each possesses limitations that constrain widespread use. Here we report a new method that overcomes major impediments to highly multi-plex tissue imaging. Iterative Bleaching Extends multi-pleXity (IBEX) uses an iterative staining and chemical bleaching method to enable high resolution imaging of >65 parameters in the same tissue section without physical degradation. IBEX can be employed with various types of conventional microscopes and permits use of both commercially available and user-generated antibodies in an ‘open’ system to allow easy adjustment of staining panels based on ongoing marker discovery efforts. We show how IBEX can also be used with amplified staining methods for imaging strongly fixed tissues with limited epitope retention and with oligonucleotide-based staining, allowing potential cross-referencing between flow cytometry, Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-Seq), and IBEX analysis of the same tissue. To facilitate data processing, we provide an open source platform for automated registration of iterative images. IBEX thus represents a technology that can be rapidly integrated into most current laboratory workflows to achieve high content imaging to reveal the complex cellular landscape of diverse organs and tissues.Significance StatementSingle cell flow cytometry and genomic methods are rapidly increasing our knowledge of the diversity of cell types in metazoan tissues. However, suitably robust methods for placing these cells in a spatial context that reveal how their localization and putative interactions contribute to tissue physiology and pathology are still lacking. Here we provide a readily accessible pipeline (IBEX) for highly multi-plex immunofluorescent imaging that enables a fine-grained analysis of cells in their tissue context. Additionally, we describe extensions of the IBEX workflow to handle hard to image tissue preparations and a method to facilitate direct integration of the imaging data with flow cytometry and sequencing technologies.

show abstract

Overview of the reporter genes and reporter mouse models

Cited by 41 publications

References 77 publications

IBEX: A versatile multiplex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues

IBEX: A versatile multiplex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues

Evaluation of TH-Cre knock-in cell lines for detection and specific targeting of stem cell-derived dopaminergic neurons

IBEX – A versatile multi-plex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues

Contact Info

Product

Resources

About