Download e-book for iPad: The Golgi Complex: Methods and Protocols by William J. Brown
By William J. Brown
Targeting new tools and strategies constructed to handle longstanding questions about the Golgi complicated, this quantity explores a various set of chapters, starting from stay and stuck telephone imaging ideas to in vitro biochemical reconstitution structures. each one bankruptcy presents a close set of particular directions, which should still permit someone to effectively entire the assays. Written for the hugely profitable Methods in Molecular Biology sequence application, chapters comprise short introductions to their respective subject matters, lists of the mandatory fabrics and reagents, step by step, comfortably reproducible laboratory protocols, and tips from the specialists on troubleshooting and warding off identified pitfalls.
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Extra info for The Golgi Complex: Methods and Protocols
If transfecting more than two 12-well plates, it may be necessary to wash each plate once prior to the 5-min Lipofectamine incubation step. After washing once, return plates to incubator and resume washing during the 20-min incubation of the Lipofectamine–DNA mixture. 5. Primary neurons are fragile and susceptible to detachment of neurites if media are added too forcefully. Examine neurons after initial washes and again after transfection to determine whether cell survival is affected by either step.
5 ml polyallomer tube. Gently mix by flicking the tube (see Note 12). 2. Allow the exchange reaction to continue for 15–30 min at room temperature. 3. Stop the exchange reaction by adding 2 μl of MgCl2 stock solution to a final concentration of 2 mM. 4. Add exomer or other Arf1 effector to the reaction at a final concentration of 1 μM and mix by flicking the tube. 5. 1). Do not run the dye front off of the gel. Ensure that virtually all of the lipids are present in the pellet fractions and not in the supernatant fractions.
Conventional confocal microscopes: Zeiss Pascal or Olympus FV300. 2. Spectral confocal microscope: Olympus FV1000. 3. Spinning disk confocal microscope: Olympus IX81 plus Disk Spinning Unit (DSU). , poly-lysine, laminin, and tenascin). Careful preparation of glass coverslips that serve as substrates for neuronal growth is a key step to assure appropriate development of axons and dendritic arbors. For most experiments we coated glass coverslips with polyL-lysine (molecular weight 30,000–70,000).
The Golgi Complex: Methods and Protocols by William J. Brown