Methodology for Studying the Compartments of the Human Breast

Abstract

A shortcoming of using human breast tissue for molecular analysis is the heterogeneous nature of the sample. To resolve this, various microdissection techniques have been employed to obtain homogeneous cells, such as manual scraping of tissue with scalpel blades, needles, or other probes to positively select cells of interest. These techniques are limited because of poor delineation of tissue and high susceptibility to contamination from dissimilar cells. Infrared laser capture microdissection (LCM), developed at the National Institute סf Health, has become increasingly commercially available in the last two decades of the twentieth century [1–3]. Laser capture microdissection, also called microdissection, laser microdissection (LMD), is a method for isolating cells or specific regions of interest from cells, tissue, or organisms. This technique enables researchers to investigate DNA/RNA and proteins from specific cells or regions of tissue. LCM consists of an LMD system, camera, and software used to select and collect the areas of interest. The basic principle of LCM involves a laser which fuses the desired material onto a specialized cap which can be then close over a 0.5 mL microcentrifuge tube. Before LCM can be performed, a paraffin-embedded tissue (PET) is sectioned with a regular microtome, or a cryostat is used to create histological sections from frozen tissue. The histological sections are placed onto a membrane slide which is then freshly hemotoxylin and eosin (H&E) stained. After the slide has been stained, the sample is set in 100 % EtOH followed by air drying.