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Rediject coelenterazine h

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RediJect Coelenterazine h is a chemical compound used as a substrate for bioluminescent reactions. It is a key component in various laboratory techniques, such as reporter gene assays and live-cell imaging, that rely on the generation of luminescent signals.

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Lab products found in correlation

D luciferin, by GoldBio (3 mentions) Bca protein assay, by Thermo Fisher Scientific (2 mentions) Ivis 100 system, by PerkinElmer (1 mentions) D luciferin, by Promega (1 mentions) Ganciclovir, by Roche (1 mentions) In vivo f pro, by Bruker (1 mentions) In vivo xtreme 4mp, by Bruker (1 mentions)

Close spelling variants of this product

Xenolight RediJect Coelenterazine h Coelenterazine h Coelenterazine

4 protocols using rediject coelenterazine h

1

Notch Pathway Modulation Assay

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The γ-secretase inhibitor Segamacestat (Selleck Biochem) was dissolved in DMSO as 10mM stock and used at 10μM final concentration. Controls received same amount of DMSO. For assays of notch pathway activation, the notch reporter plasmid pCBFRE-luc, a gift from N. Gaiano (Addgene plasmid # 26897), was used. The notch reporter was co-transfected with a constitutive expression (EF1α promoter-driven) renilla-luciferase plasmid. Firefly luciferase (Notch reporter) and Renilla luciferase bioluminescence activity were measured by in vivo imaging system (IVIS) and determined consecutively by adding respective substrates (D-luciferin, Gold Biotechnology and RediJect Coelenterazine h, PerkinElmer, respectively) and included background measurements before and between reporter analyses. Notch reporter signal was normalized to the signal from constitutive plasmid. Dominant negative forms of RBPJ (DN-RBPJ) and MAML (DN-MAML) were applied to inhibit the intracellular notch pathway. This was done either by co-transfection in the notch reporter assays or using lentiviral transduction followed by GFP-based flow cytometric sorting to remove non-transduced cell population.
Welte T., Kim I.S., Tian L., Gao X., Wang H., Li J., Holdman X.B., Herschkowitz J.I., Pond A., Xie G., Kurley S., Nguyen T., Liao L., Dobrolecki L.E., Mo Q., Edwards D.P., Huang S., Xin L., Xu J., Li Y., Lewis M.T., Wang T., Westbrook T.F., Rosen J.M, & Zhang X.H. (2016). Oncogenic mTOR signaling recruits myeloid-derived suppressor cells to promote tumor initiation. Nature cell biology, 18(6), 632-644.
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2

Notch Pathway Modulation Assay

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The γ-secretase inhibitor Segamacestat (Selleck Biochem) was dissolved in DMSO as 10mM stock and used at 10μM final concentration. Controls received same amount of DMSO. For assays of notch pathway activation, the notch reporter plasmid pCBFRE-luc, a gift from N. Gaiano (Addgene plasmid # 26897), was used. The notch reporter was co-transfected with a constitutive expression (EF1α promoter-driven) renilla-luciferase plasmid. Firefly luciferase (Notch reporter) and Renilla luciferase bioluminescence activity were measured by in vivo imaging system (IVIS) and determined consecutively by adding respective substrates (D-luciferin, Gold Biotechnology and RediJect Coelenterazine h, PerkinElmer, respectively) and included background measurements before and between reporter analyses. Notch reporter signal was normalized to the signal from constitutive plasmid. Dominant negative forms of RBPJ (DN-RBPJ) and MAML (DN-MAML) were applied to inhibit the intracellular notch pathway. This was done either by co-transfection in the notch reporter assays or using lentiviral transduction followed by GFP-based flow cytometric sorting to remove non-transduced cell population.
Welte T., Kim I.S., Tian L., Gao X., Wang H., Li J., Holdman X.B., Herschkowitz J.I., Pond A., Xie G., Kurley S., Nguyen T., Liao L., Dobrolecki L.E., Mo Q., Edwards D.P., Huang S., Xin L., Xu J., Li Y., Lewis M.T., Wang T., Westbrook T.F., Rosen J.M, & Zhang X.H. (2016). Oncogenic mTOR signaling recruits myeloid-derived suppressor cells to promote tumor initiation. Nature cell biology, 18(6), 632-644.
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3

Multimodal Monitoring of Transduced Cells

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For the hMultistem cells, eGFP expression was used for visual confirmation of transduction after which fLuc and HSV-tk expression were assessed. For fLuc expression, 100.000 cells were seeded in triplicate in a 24-well plate and were allowed to attach before BLI measurements were performed. An amount of 75 μg of D-luciferin (Promega, Madison, WI, USA) was added per well prior to BLI experiments using an IVIS 100 system (Perkin Elmer, Waltham, MA, USA). The temperature was maintained at 37°C. Scanning parameters included medium binning, f stop = 1, time = 10 s or 1 min. Data were analyzed by using the living image 2.50.1 software. Similarly, mCherry expression was used for the tumor cell lines in order to confirm transduction by fluorescence microscopy. BLI was performed using the rLuc substrate coelenterazine-h (Rediject Coelenterazine-h, Perkin Elmer, 0.25 μg/well).
HSV-tk expression was confirmed with a GCV killing experiment. Hereby, 20.000 cells were seeded in a 24-well plate and were allowed to grow. The following day, ganciclovir (Cymevene, Roche, Basel, Switzerland) was added in different concentrations (100 μM, 1 μM, and 0.01 μM) for four consecutive days, after which BLI was performed. Cells were subsequently collected and a BCA protein assay (Thermo Scientific, Rockford, USA) was performed.
Leten C., Trekker J., Struys T., Roobrouck V.D., Dresselaers T., Vande Velde G., Lambrichts I., Verfaillie C.M, & Himmelreich U. (2016). Monitoring the Bystander Killing Effect of Human Multipotent Stem Cells for Treatment of Malignant Brain Tumors. Stem Cells International, 2016, 4095072.
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4

Xenograft ATC Model for Metastasis

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All animal studies were approved by Weill Cornell Medicine’s Institutional Animal Care and Use Committee. 8–10-week-old female and male NOD-scidIL2Rgnull (NSG) mice (Jackson laboratory) were used for xenograft experiments with ATC cells. GFP and fLuc-expressing 8505C (0.75×106 cells per mouse) and patient-derived ATC cells (0.36×106 cells per mouse) were i.v. injected to establish systemic metastasis establishment (28 (link)). Longitudinal measurements of 8505C tumor burdens were taken using a whole-body optical imager (In-Vivo Xtreme 4MP, Bruker) 15 minutes after intraperitoneal injection of 100 µl of 150 µg/ml D-luciferin (GoldBio). Total tumor burden was measured by generating full body region of interests (ROIs) and integrating total flux of luminescence using Bruker Analysis software. GFP-expressing livers, spleen, heart, and lungs were extracted from mice and imaged with a whole-body optical imager (In-vivo F-Pro, Bruker). To track ICAM-1-CAR T cells expressing rLuc, mice were injected i.v. with 100 µl of RediJect Coelenterazine H (Perkin Elmer) and then immediately imaged using the In-Vivo Xtreme 4MP (Bruker).
Min I.M., Shevlin E., Vedvyas Y., Zaman M., Wyrwas B., Scognamiglio T., Moore M.D., Wang W., Park S., Park S., Panjwani S., Gray K.D., Tassler A.B., Zarnegar R., Fahey TJ I.I.I, & Jin M.M. (2017). CAR T therapy targeting ICAM-1 eliminates advanced human thyroid tumors. Clinical cancer research : an official journal of the American Association for Cancer Research, 23(24), 7569-7583.
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