By Trygve O. Tollefsbol (auth.), Trygve O. Tollefsbol (eds.)
Biological getting older: equipment and Protocols investigates many of the tactics which are stricken by the age of an organism. numerous new instruments for the research of organic getting older were brought lately, and this quantity presents tools and protocols for those new suggestions as well as its insurance of validated approaches. The editors have conscientiously chosen merely these themes which are thought of mainstays of the sphere or are displaying promise in revolutionizing this particularly new technology. the 3 major components of concentration during this state of the art compendium of organic getting older learn are: equipment which are simple to knowing the elemental mechanisms of mobile getting older; concepts used to intrude within the getting older method; and ways to interpreting the various molecular tactics of organic getting older. Researchers looking new expertise and methods will locate this quantity of large profit as they stream in the direction of new instructions within the interesting and increasing box of organic getting older.
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Extra info for Biological Aging: Methods and Protocols
Figure 1 outlines an equation developed in our lab that takes into account all the cells counted and results in a more exact population doubling. 4. Notes 1. 9% (w/v) NaCl to detach cells and chelate Ca2+ and Mg2+ ions that could hinder the action of trypsin. 2. With the exception of BJ fibroblasts and other primary cell cultures, 1–2 min submerged in trypsin at room temperature or 37°C should be sufficient to disperse cells. BJs and other difficult cell types may require 5–15 min of incubation at 37°C, with occasional checks under an inverted microscope to ensure that cells are detached.
The results illustrated that IGF-1 mRNA production 02_Tollefsbol 1/25/07 12:57 PM Page 16 16 Phipps et al. , 1995 (26) Itahana et al. (Chapter 4, this volume) was lowered to undetectable levels in senescent cells whereas IGF-1 receptor mRNA production remained at detectable levels. Another biological marker that is differentially regulated in senescent cells is epidermal growth factor (EGF). EGF signaling has been postulated to be impaired in nonproliferating senescent human diploid fibroblasts downstream of receptor binding.
1. DHR 123 STAINING OF PEROXIDES (NOTE 11) 1. ) (final DHR123 concentration 30 μM). 2. Cells are incubated at 37°C for 30 min in the dark. 3. After incubation, cells are centrifuged (400g, 5 min) and supernatant discarded. 4. The pellet is resuspended in 3 mL of serum-free DMEM and is immediately used for analysis and sorting (Note 12). 5. ). Sorting gates are defined in Fl1 (logarithmic), conveniently in a FSC/FL1 dotplot. Use the left and right quartiles with respect to FL1 to sort cells with highest (high FL1) and lowest (low FL1) peroxide levels.