Fluorescent genetic barcoding for cellular multiplex analyses

Maetzig, Tobias; Morgan, Michael; Schambach, Axel

doi:10.1016/j.exphem.2018.08.001

Cited by 5 publications

(3 citation statements)

References 50 publications

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“…After sample mixing, each sample becomes traceable in longitudinal studies through the detection of its color code by flow cytometry. In general, FGB vector systems either make use of a "one color code per vector" approach or utilize combinatorial transductions of differentially color-coded vectors to generate 6 (=6xFGB) to >100 unique fluorescent identifiers [25]. The 6xFGB system appeared most reliable in our hands, since each vector produces a unique color code, optionally accommodates sgRNA expression cassettes, and enables the fate tracking of mixed hematopoietic samples in vitro and in vivo [21].…”

Section: Introductionmentioning

confidence: 99%

A Multiplex CRISPR-Screen Identifies PLA2G4A as Prognostic Marker and Druggable Target for HOXA9 and MEIS1 Dependent AML

Hassan

Lieske

Dörpmund

et al. 2021

IJMS

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HOXA9 and MEIS1 are frequently upregulated in acute myeloid leukemia (AML), including those with MLL-rearrangement. Because of their pivotal role in hemostasis, HOXA9 and MEIS1 appear non-druggable. We, thus, interrogated gene expression data of pre-leukemic (overexpressing Hoxa9) and leukemogenic (overexpressing Hoxa9 and Meis1; H9M) murine cell lines to identify cancer vulnerabilities. Through gene expression analysis and gene set enrichment analyses, we compiled a list of 15 candidates for functional validation. Using a novel lentiviral multiplexing approach, we selected and tested highly active sgRNAs to knockout candidate genes by CRISPR/Cas9, and subsequently identified a H9M cell growth dependency on the cytosolic phospholipase A2 (PLA2G4A). Similar results were obtained by shRNA-mediated suppression of Pla2g4a. Remarkably, pharmacologic inhibition of PLA2G4A with arachidonyl trifluoromethyl ketone (AACOCF3) accelerated the loss of H9M cells in bulk cultures. Additionally, AACOCF3 treatment of H9M cells reduced colony numbers and colony sizes in methylcellulose. Moreover, AACOCF3 was highly active in human AML with MLL rearrangement, in which PLA2G4A was significantly higher expressed than in AML patients without MLL rearrangement, and is sufficient as an independent prognostic marker. Our work, thus, identifies PLA2G4A as a prognostic marker and potential therapeutic target for H9M-dependent AML with MLL-rearrangement.

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Section: Introductionmentioning

confidence: 99%

A Multiplex CRISPR-Screen Identifies PLA2G4A as Prognostic Marker and Druggable Target for HOXA9 and MEIS1 Dependent AML

Hassan

Lieske

Dörpmund

et al. 2021

IJMS

Self Cite

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“…Fluorescence-based encoding is the most common technique, because of the easily tunable emission spectra, based on the structure of the fluorophore, and the easy readout. Examples include fluorescent-dye-doped micro- and nanoparticles, , fluorescent nanorods, DNA-based fluorescence nanobarcodes and genetically encoded fluorescent barcodes for cellular multiplexing . Among the fluorescent materials, it is mostly organic dyes that are used for fluorescent barcodes, due to their widespread availability, but quantum dots and fluorescent proteins are also commonly used, especially in biological applications. , The main disadvantages of fluorescent barcodes are the photobleaching and the relatively broad emission spectra, which limits their encoding capacity.…”

Section: Types Of Optical Microbarcodesmentioning

confidence: 99%

Microcavity- and Microlaser-Based Optical Barcoding: A Review of Encoding Techniques and Applications

2023

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Optical microbarcodes have recently received a great deal of interest because of their suitability for a wide range of applications, such as multiplexed assays, cell tagging and tracking, anticounterfeiting, and product labeling. Spectral barcodes are especially promising because they are robust and have a simple readout. In addition, microcavity- and microlaser-based barcodes have very narrow spectra and therefore have the potential to generate millions of unique barcodes. This review begins with a discussion of the different types of barcodes and then focuses specifically on microcavity-based barcodes. While almost any kind of optical microcavity can be used for barcoding, currently whispering-gallery microcavities (in the form of spheres and disks), nanowire lasers, Fabry–Pérot lasers, random lasers, and distributed feedback lasers are the most frequently employed for this purpose. In microcavity-based barcodes, the information is encoded in various ways in the properties of the emitted light, most frequently in the spectrum. The barcode is dependent on the properties of the microcavity, such as the size, shape, and the gain materials. Various applications of these barcodes, including cell tracking, anticounterfeiting, and product labeling are described. Finally, the future prospects for microcavity- and microlaser-based barcodes are discussed.

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“…With the advent of new experimental methods to track clones in vivo (e.g. (Sun, Ramos et al 2014), (McKenna, Findlay et al 2016), (Alemany, Florescu et al 2018), (Raj, Wagner et al 2018), (Spanjaard, Hu et al 2018), (Wagner, Weinreb et al 2018), (Maetzig, Morgan et al 2018) and the rapid expansion in the size of datasets, many statistical and mathematical modelling approaches have been developed to conduct inferences from the data. These approaches are becoming extremely important to deriving information about the dynamics of hematopoiesis in vivo.…”

Section: Seeing the Forest Using The Treesmentioning

confidence: 99%

A track of the clones: new developments in cellular barcoding

Lyne

Kent

Laurenti

et al. 2018

Experimental Hematology

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International experts from multiple disciplines gathered at Homerton College in Cambridge, UK from September 12-14, 2018 to consider recent advances and emerging opportunities in the clonal tracking of hematopoiesis in one of a series of StemCellMathLab workshops. The group included thirty-five participants with experience in the fields of theoretical and experimental aspects of clonal tracking, and ranged from PhD students to senior professors. Data from a variety of model systems as well as from clinical gene therapy trials were discussed alongside strategies for data analysis and sharing, as well as challenges arising due to underlying assumptions in data interpretation and communication. Recognizing the power of this technology underpinned a group consensus of a need for improved mechanisms for sharing data and analytical protocols to maintain reproducibility and rigor in its application to complex tissues.

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Fluorescent genetic barcoding for cellular multiplex analyses

Cited by 5 publications

References 50 publications

A Multiplex CRISPR-Screen Identifies PLA2G4A as Prognostic Marker and Druggable Target for HOXA9 and MEIS1 Dependent AML

A Multiplex CRISPR-Screen Identifies PLA2G4A as Prognostic Marker and Druggable Target for HOXA9 and MEIS1 Dependent AML

Microcavity- and Microlaser-Based Optical Barcoding: A Review of Encoding Techniques and Applications

A track of the clones: new developments in cellular barcoding

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