Microinjection as a tool to explore small GTPase function

Microinjection overcomes the plasma membrane barrier to the introduction of charged or large nonlipid soluble molecules into cells by the direct insertion of a hollow capillary micropipette into the cell. With the application of pressure, aqueous solution is then directly transferred into either the cytosol or the nucleus. I give specific examples of the application of this approach to the functional study of small GTPases of the Sar1, ARF, and rab family in membrane trafficking between the Golgi apparatus and endoplasmic reticulum (ER). The principles illustrated by these examples should be generally applicable to other small GTPases. Detailed protocols for capillary microinjection using semiautomated equipment are given.