Previously we reported on a free-hand dissection guide for proteomics samples, where the major focus was on dissection of fresh brain tissue. Although the fresh brain provides clear advantages, such as being able to see structures in very great detail by differences in color and shade, it also has some disadvantages. This particularly becomes evident when processing multiple samples within a short time-window, e.g., after a physiological stimulus. In this chapter, we discuss some of the current methods to take tissue samples from frozen tissue, keeping in mind the requirements for subcellular isolation (e.g., synaptosomes) of proteomics samples. In addition, we provide a step-by-step protocol to improve standardization of tissue dissection in your lab.