Microinjection of Proteins into Somatic Cells: Needle Microinjection and Scrape Loading

Abstract

This chapter focuses on the needle microinjection and scrape loading of protein molecules into cultured somatic cells. The presence of a fluorescent marker in the injection solution can greatly facilitate microinjection. It allows the examination of flow and easy identification of injected cells. It is critical to obtain an optimal shape of the microneedle by adjusting the temperature and force of the pipette puller. Each needle can be used for more than 50 cells, assuming that the solution for microinjection is reasonably clean. The volume delivered by needle microinjection is usually in the range 1 to 10% cell volume. With some practice, the variability can be maintained to within a factor of 2. A number of factors can affect the volume, including the pressure, tip size, accumulation of proteins at the tip, and presence of any air bubbles in the needle. Compared with needle microinjection, the amount of delivery varies more extensively among scrape-loaded cells. The yield of injected cells and the extent of cell damage also vary among cell types. Cell damage often manifests as the appearance of large vacuoles.