What happens to a specimen in pathology?

At first sight of a histology slide, you might doubt how difficult it can be to prepare the slide, but in fact, you cannot even imagine where the slide starts with. Indeed, it is a black box even for some medical specialists how a tissue specimen is prepared into a histology slide. There are quite several steps along the complex process that the pathologist relies on to figure out an accurate diagnosis. I am going to break it down in a simple way that even non-medical specialists can get a glimpse of.

A tissue specimen is obtained in a variety of clinical scenarios, from a doctor's office to an operating room. A fresh specimen is initially placed in a specimen container or bag depending on the size and the degree of fragmentation. Primary fixation with formalin may be required for certain specimens, including malignancy and tissues rich in enzymes. The specimen container or bag is then preserved in the refrigerator until it is collected and transported to pathology.

Upon arrival at pathology, the specimen is weighted and measured. The color and the texture are recorded as well. The surgical margin is inked if there is clinical suspicion or evidence of malignancy. Inking helps identify the invasion of cancer cells beyond the margin. Afterward, the specimen is cut manually at 5 mm intervals. Representative sections that appear heterogeneous in nature or contain lesions are collected and trimmed to fit into one or more cassettes. Extended fixation with formalin usually takes additional 4-8 hours before further processing.

An adequately fixed specimen is dehydrated and embedded in the medium, in most cases, paraffin. Until this stage, we describe that the tissue specimen is formalin-fixed paraffin-embedded (PPFE). The PPFE tissue is sectioned with a rotary microtome or other cutting devices at a 3-5 μm interval. The section floats on the water surface and is placed onto a slide. The embedding medium is then replaced with formalin again for subsequent staining. As a reminder, stains are mostly water-soluble, including hematoxylin and eosin. Finally, the corresponding patient label is printed on the slide, and the pathologist is ready to review it under the microscope.

Preparation of a histology slide is indeed a complex process. Every step is crucial for the quality of the slide and definitely can influence the pathologic diagnosis if this process is not carefully carried out. Therefore, quite a few automatic machines have been invented to eliminate potential human errors during the process. These machines do not only automate tissue embedding, serial sectioning, and staining but also standardize the steps to obviate the variability of technicians. Anyway, never underestimate the effort it takes to prepare even just a histology slide. It is never easy as it seems!