Brainbow 101

Chris Potter, John's Hopkins School of Medicine

Chris Potter, Ph.D., Assistant Professor, Department of Neuroscience, Johns Hopkins University School of Medicine.
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Complex tissues, like the brain, are composed of thousands to millions to billions of similar cell-types (e.g., neurons or glia). The general labeling of a cell-type, such as all neurons in the brain, does not allow identification of individual cells within the same tissue. The ability to distinguish one neuron from another allows tracing of neural circuits, the characterization of cell-cell interactions, and the ability to follow changes to the cell and its progenitors throughout development.

Basic idea behind Brainbow lineage marking showing how stochastic deletion by site specific recombination of fluorescent protein-encoding modules leads to different fluorescent outputs.

Brainbow technology, first described by Livet and Lichtman for the mouse model (Livet et al, Nature, 2007) presented a breakthrough in cell labeling. Instead of labeling a complex tissue in the traditional single color (e.g, green by GFP), the Brainbow technology allowed different neurons in the tissue to be colored stochastically and independently in up to 3 different colors (CFP, YFP, RFP). The different combinations of colors (cyan, yellow, and red) allowed ~90 different distinguishable hues, enabling many individual neurons to be simultaneously identified. Brainbow technology colorfully revealed the beauty inherent to complex tissues.

Brainbow. Mouse hippocampus.
Image Credit: Tamily Weissman, Harvard University,
on Cell Picture Show

Since 2007, there have been many improvements to Brainbow, increasing its utility, and adapting the multi-color labeling technology to other organisms like fly, fish, and plants. Following these improvements, and understanding which system is best to address one’s questions, can be daunting. Fortunately, a new review by Weissman and Pan (Genetics, 2015) helps explain the recent improvements, what types of questions can be addressed, and how Brainbow imaging can be optimized to the greatest effect.

The authors adapted Brainbow technology to the zebrafish system (Zebrabow; Pan et al, Development, 2013) and are well versed in the intricacies of multi-color labeling. In their review, they satisfyingly cover both the history and up-to-date variants of Brainbow technology. They explain, with numerous examples and schematics, how Brainbow technology works, from the genetically encoded approaches used for multi-color labeling to more recent viral-based approaches. They also explain how to make the best of Brainbow labeling, and describe how colors variations might be best represented to yield the greatest degree of separation.

Brainbow technology applied to Drosophila melanogaster. Image from “Fly Over the Brainbow” by Veronique Greenwood

Weissman and Pan explain and explore Brainbow technologies in the many different model organisms, and help clarify the differences between available Brainbow variants. For example, in Drosophila, there are many Brainbow versions: dBrainbow, Flybow, TIE-DYE, and LOLLibow. The review by Weissman and Pan helps guide the reader through the strengths and weaknesses to each approach.

Weissman and Pan have written an excellent primer on Brainbow. It will help the interested reader make sense of what is available, and how multi-color labeling might be applied to one’s own research. Also worthy of mention is another outstanding Brainbow review by Richier and Salecker (WIREs Developmental Biology, 2015). This review provides further explanation of Brainbow technologies, and includes comprehensive descriptive tables highlighting available transgenic lines and fluorescent proteins. With these Brainbow reviews in hand, the reader can quickly become well versed in the scientific art of multi-color labeling.

Tamily A. Weissman and Y. Albert Pan (2015)Brainbow: New Resources and Emerging Biological Applications for Multicolor Genetic Labeling and Analysis  Genetics February 2015 199:293-306; dpi: 10.1534/genetics.114.172510

Richier, B. and Salecker, I. (2015), Versatile genetic paintbrushes: Brainbow technologies. WIREs Dev Biol, 4: 161–180. doi: 10.1002/wdev.166

Livet J, Weissman TA, Kang H, Draft RW, Lu J, Bennis RA, Sanes JR, Lichtman JW. (2007) Transgenic strategies for combinatorial expression of fluorescent proteins in the nervous system. Nature. 2007 Nov 1;450(7166):56-62. doi:10.1038/nature06293

Y. Albert Pan, Tom Freundlich, Tamily A. Weissman, David Schoppik, X. Cindy Wang, Steve Zimmerman, Brian Ciruna, Joshua R. Sanes, Jeff W. Lichtman, and Alexander F. Schier  (2013) Zebrabow: multispectral cell labeling for cell tracing and lineage analysis in zebrafish  Development 2013 140:2835-2846; doi:10.1242/dev.094631

 

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