Download e-book for kindle: Apoptosis and Cancer: Methods and Protocols by Gil Mor, Ayesha B. Alvero

By Gil Mor, Ayesha B. Alvero

ISBN-10: 1493916602

ISBN-13: 9781493916603

ISBN-10: 1493916610

ISBN-13: 9781493916610

In Apoptosis and melanoma: tools and Protocols, Second Edition, professional researches within the box element the functionality of molecular and mobile biology innovations for learning and detecting the activation of the apoptotic pathway. Chapters specialise in assays constructed to notice its activation not just in vitro but in addition in vivo, optimized multiplex research, medium- to high-throughput displays, and the mobile procedure. Written within the hugely profitable Methods in Molecular Biology series structure, chapters comprise introductions to their respective subject matters, lists of the required fabrics and reagents, step by step, easily reproducible laboratory protocols, and key tips about troubleshooting and keeping off identified pitfalls.

Authoritative and functional, Apoptosis and melanoma: tools and Protocols, moment version aids scientists as a stand-alone source for the execution and research of the defined protocols and as a reference for the research and detection of apoptosis inside of and out of doors the realm of melanoma examine.

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Extra info for Apoptosis and Cancer: Methods and Protocols

Example text

Multichannel pipette: 12-channel, 30–300 μL. 4. 5 mL microfuge tubes. 2 Cell Line Components 1. CellPlayer™ NucLight Red Cell Line (see Note 2). 2. Low riboflavin media: F-12 or MEM (see Note 3). 3. 10 % FBS. 4. Puromycin (see Note 4). 3 Reagents and Compounds 1. Caspase-3/7 Reagent (Essen BioScience, Ann Arbor, MI, USA, Cat # 4440). 2. 53 g/mol. Make working stock of 500 μM in 100 % DMSO. 35 g/mol. Make working stock of 1 mM in 100 % DMSO. 35 g/mol. Make working stock of 1 mM in 100 % DMSO. 1 Cell Culture and Seeding Assay Plate 1.

If using a different NucLight Red cell line, it is the user’s Kinetic Measure of Apoptosis 41 responsibility to optimize the effective puromycin or bleomycin concentration for that cell type according to the Essen BioScience’s lentivirus reagent instructions. Puromycin or bleomycin should be removed during the assay. 5. Any drug compound can be tested in this assay. 4 %. The protocol outlined above tests three common apoptotic and cytostatic compounds. If testing new drug compounds, we recommend including one known apoptotic compound (staurosporine) and one known cytostatic compound (cycloheximide).

11. Condensation will form on the plate when transferred from ambient temperature to the 37 °C, humidified environment. Scanning right away will cause the images to appear out of focus. Allow the plate to equilibrate in the incubator for 15–30 min to avoid condensation. The effects of very toxic compounds, such as staurosporine, can be seen immediately, so it is imperative to begin imaging less than 30 min–1 h after addition to cells. 12. The first time the apoptosis experiment is performed, a new image collection and processing definition need to be created.

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