By Michael C. Dovey, Leonard I. Zon (auth.), John S. Yu (eds.)

Through the progressive thought of melanoma stem cells, melanoma examine has been reinvigorated to check the position of those specified cells in melanoma propagation and as objectives of leading edge cures. In Cancer Stem Cells: tools and Protocols, preeminent researchers have compiled melanoma stem mobile study suggestions and protocols to advertise fit pageant, discourse, and collaboration during this very important box. the amount covers vast subject matters similar to id and isolation of melanoma stem cells, animal types of melanoma stem cells, methylation profiling, the contribution of the area of interest within the rules of melanoma stem cells, immunologic focusing on, and using general stem cells as a therapy, between different matters. Written within the hugely profitable Methods in Molecular Biology™ sequence structure, chapters comprise short introductions to their respective issues, lists of the mandatory fabrics and reagents, step by step, effortlessly reproducible laboratory protocols, and professional notes on troubleshooting and averting identified pitfalls.

Comprehensive and state-of-the-art, Cancer Stem Cells: tools and Protocols will relief scientists around the globe within the furthering of our knowing of melanoma initiation and propagation and, most significantly, within the improvement of novel ambitions for melanoma therapy.

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3. Mouse Irradiation 1. Irradiate the syngenic or congenic mice (8–12 weeks old) in which the leukemic cells are to be injected at lethal doses of radiation (800–850 cGy of whole body irradiation) (see Note 11) ∼2–3 h prior to injection. 4. Injection of Mice 1. Transfer the lethally irradiated mouse from the cage and place it gently into a mouse-restraint device exposing the tail. 2. Prewarm the tail of the restrained mouse for vasodilation with a safe heat source such as warm water or heat lamp for 30–60 s (see Note 12).

Perform the staining for 30 min at 4°C. 3. Wash the labeled cells twice with PBS 2% FBS. 4. Finally, resuspend the cells in FACS buffer at a concentration of 2–5 × 107 per ml. 5. Define the quadrants on the FACS Vantage cell sorter, using isotype controls. If sorting of CD34+ CD38− population is desired, define the population by strictly selecting for cells with high CD34 expression and a CD38 expression less than half of the maximum CD38 APC fluorescence of the isotype control. 6. Sort the cells at a maximum rate of 3,000–6,000 cells per second in sterile FACS tubes containing IMDM 50% FCS (collection medium).

These brain tumor stem cells are a kind of cancer stem cell, competent to form tumors that mimic the parent tumor in experimental animals. Studies from our laboratory and others have demonstrated that brain tumor stem cells and NSC share similar mechanisms and pathways for proliferation. For example, we have identified that one of the AMPK/snf1 kinases, maternal embryonic leucine zipper kinase (MELK), is highly expressed in NSC and malignant brain tumors, as well as in brain tumor stem cell-enriched cell cultures.

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