Cell Biology Application of Acumen eX3 High Content Cell Analyzer (III)——Stem Cell Related Research
High-throughput cell biology refers to the integration of automated instrumentation and cell biology techniques to answer questions that traditional methods cannot solve. It requires advanced imaging techniques, chemical reagents, and biological methods to obtain information on cell phenotypes, functions, and interactions in a fast, multi-channel manner. Its core idea is to get as much data as possible—whether it comes from a large number of samples or a “omics” approach without compromising the quality of the data.
The Acumen eX3 high-content cell analysis platform from TTP LabTech UK is equipped with three different wavelength solid-state laser generators at 405nm, 488nm and 633nm, which are compatible with many fluorescent dyes. Acumen eX3 has the capability of full-hole high-speed imaging and multiple fluorescence analysis. It combines the advantages of CCD imaging system and flow cytometry to complete the whole plate scanning of microplates (including 96, 384, 1536 well plates) in 10 minutes. . Currently, the platform has been successfully applied to various cell-level studies, including screening studies on human genomic RNAi libraries 1-3, cancer-related research 2 (including cell cycle, cell proliferation, clonal formation, cytotoxicity, apoptosis) , cell adhesion, angiogenesis, protein kinases, etc., and stem cell differentiation studies3. In particular, the high-quality imaging advantages of AcumeneX3 are also highly regarded in the field of systems biology such as tissue sections and animal models such as nematodes, fruit flies and zebrafish. In the past few years, Acumen eX3 has become one of the most highly regarded high-throughput cell biology research tools in the research and pharmaceutical field.
Stem cells play an important role in the body repair process because of their ability to differentiate into other cell types. Because of the potential of stem cells to treat human diseases, research on stem cells is considered one of the most important and potential directions in today's medical research.
Since stem cell proliferation and differentiation are small-probability events, capturing such events requires expanding the screening volume and imaging and analyzing all cells as much as possible. Acumen eX3 is the high-throughput full-hole imaging capability that is ideal for detecting compound stimulation and RNAi-induced proliferation and differentiation of stem cells. Through a variety of molecular markers and dyes, Acumen eX3 accurately counts the ratio of differentiated stem cells to total cell count.
Figure 1 Determination of stem cell proliferation using Brdu-Alexa 488 antibody and PI staining
Neural stem cells are stem cells that are present in brain tissue and can differentiate into neurons, astrocytes, oligodendrocytes, and the like. Its self-proliferation has a relatively stable source of cell banks. In addition to its multi-directional differentiation potential, neural stem cells have lower immunogenicity and do not express mature cell antigens. They have good tissue fusion and can survive in the host for a long time. Our users have also successfully used Acumen eX3 for the detection of neural stem cell differentiation experiments.
Figure 2. After induction of differentiation of mNPC cells, DAPI staining and Tuj1 antibody labeling were performed and scanning imaging was performed using Acumen eX3 405 nm laser and 488 nm laser.
references
1. Kittler, R. et al. Genome-scale RNAi profiling of cell division in human tissue culture cells. Nat. Cell Biol. 9, 1401-12 (2007).
2. Kittler, R. et al. An endoribonuclease-prepared siRNA screen in human cells identification genes essential for cell division. Nature 432, 1036–1040 (2004).
3.Cheng et al. Antisense inhibition of human miRNAs and indications for an involved for miRNA in cell growth and apoptosis. Nucleic Acids Res. 33(4):1290-7 (2005)
4. Chresta et al., AZD8055 is a potent, selective, and orally bioavailable ATP-competitive mammalian target of rapamycin kinase inhibitor with in vitro and in vivo antitumor activity.,, Cancer Res. Jan 1;70(1):288- 98 (2010)
5.Phong et al., The p38 MAPK promotes cell survival in response to DNA damage butis not required for G2 DNA damage checkpoint in human cancer cells Mol Cell Biol.Jun 1 (2010)
6.Wylie & Bowen Determination of cell colony formation in a high-content screening assay Clin Lab Med 27(1): 193-9 (2007)
7.Wedge et al. AZD2171: A highly potent, orally bioavailable, vascular endothelial growth factor receptor-2 tyrosine kinase inhibitor for the treatment of cancer. Cancer Res 65(10): 4389-400 (2005)
8. Kunz-Schughart, LA et al., Potential of fibroblasts to regulate the formation of three-dimensionalvessel-like structures from endothelial cells in vitro Am J Physiol Cell Physiol 290 (5), C1385 (2006)
9. Hulkower K, Perr M, Quantifying Cell Migration and Invasion. GEN, 2008;
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10.Zhang JH, Chung TD, Oldenburg KR, "A Simple Statistical Parameter for Use in
Evaluation and Validation of High Throughput Screening Assays."J Biomol Screen.
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May 26, 2022
