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Extra resources for Adipose Tissue Protocols (Methods in Molecular Biology Vol 155)

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The labeled probe, which can be employed in several ISH reactions, is then stored at –20°C until use. Probe concentration depends on the type of reaction and of tissue. 2. PREHYBRIDIZATION Before performing a hybridization reaction, paraffin needs to be removed (by two passages in xylol, each of at least 10 min). Sections are rehydrated in decreasing EtOH concentrations (100, 95, 75, 50, and 30%), then in H2O DEPC. 075 M Na-citrate). Sections can be prehybridized for 2 h at 58°C in the hybridization mixture of 50% formamide, 5X SSC, and denaturated salmon sperm DNA (32).

Sewter, C. , Byrne, C. , and O’Rahilly, S. (1998) Depot-related gene expression in human subcutaneous and omental adipocytes. Diabetes 47, 1384–1391. 71. Gemmell, R. , Bell, A. , and Alexander, G. (1972) Morphology of adipose cells in lambs at birth and during subsequent transition of brown to white adipose tissue in cold and in warm conditions. Am. J. Anat. 133, 143–164. 72. , and Ricquier, D. (1993) Beta 3-adrenoceptor agonist ICI-D7114 is not as efficient on reinduction of uncoupling protein mRNA in sheep as it is in dogs and smaller species.

5–2 nm) with a sputter coater. This coating is thinner than that used for conventional SEM observation, and allows study of cells in their finest ultrastructural detail, but requires such careful tissue handling that, for instance, specimens cannot even be moved from one laboratory to another. 7. 1. ). 2. 2, for 2 h at 4°C, followed by washing in the same buffer. 3. 4, for 1–2 h at RT and frozen in freon cooled with liquid nitrogen. Specimens are fractured, and replicated at –112°C in a Balzers freeze-fracturing device (Balzers, Lichenstein).

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