Next Lesson - How to Read a Histology Slide
Core
Pathology comes from the Greek words pathos (suffering) and logos (study), so literally means the study of suffering. Webster dictionary defines pathology more fully as:
“The study of the essential nature of diseases and especially of the structural and functional changes produced by them”
Pathology can be seen as the disease-focused counterpart to biology. While biology is concerned with the study of life and cellular function, pathology is concerned with disease and cellular dysfunction.
Diagnostic pathology involves studying the structural and functional changes in cells and tissues in order to arrive at a diagnosis. Ultimately, many diseases can be understood as consequences of failed homeostasis, with associated changes in cell morphology (shape and structure) and function. Thus, the cell can be seen to be at the centre of many diseases.
Modern pathology is commonly credited as having stemmed from the work of German physician Rudolf Virchow (1821-1902). Virchow was a strong critic of the theory of the four humours (the old idea that disease came from an imbalance of four body fluids). This theory still had supporters in the 19th century. He was one of the first physicians to examine disease at the cellular level and is regarded as a founder of cellular pathology. He encouraged his students to use microscopes and to "think microscopically", and had a major impact on medical education in Germany. He also coined terms that are still in use today: "leukaemia", "embolism" and "thrombosis".
You will find yourself confronted with the works of Virchow throughout your medical journey.
Image - Rudolf Virchow, the father of modern pathology
Public Domain Source [Public Domain]
- Medical microbiology
- This is the study of microorganisms such as bacteria, viruses, fungi, and parasites. It aims to understand their role in disease and symptom presentation.
- Chemical pathology
- This field combines clinical skills with laboratory work. Chemical pathologists use biochemical tests to diagnose disease and to manage patients.
- Haematology
- Haematology is the branch of medicine concerned with the study and treatment of diseases of the blood.
- Immunology
- This branch of pathology is concerned with the immune system (the body’s defences) and how it protects against, or causes, disease.
- Histopathology and cytopathology
- Histopathology is the study of the cellular changes within tissues that are caused by disease. Cytopathology is a diagnostic technique that examines sampled cells from various body sites to determine the cause or the nature of disease. An example of a practical use of cytopathology is cervical screening (which used to be called a smear test). The sample is first tested for the types of human papillomavirus (HPV) known as high-risk HPV, which can cause cervical cancer. If high-risk HPV is found, the cells from the same sample are examined under the microscope for abnormal changes.
Pathologists provide information to decide the best path forward for patient recovery, so their role is essential in every aspect of patient care. Lesions (areas of abnormal tissue) can be biopsied before surgery, to help decide whether surgery is needed and to guide what surgery is done. Microscopic analysis can provide further insight into which drugs or other treatments will be the most suitable.
There are two main types of sample for diagnostic microscopy: tissue samples (histology) and cell samples (cytology).
Histology (the study of tissues) involves the removal of a tissue sample to be able to assess both tissue and cell morphology.
Cytology (the study of cells) examines individual cells and small cell groups, often sampled by fine-needle aspiration (drawing cells out through a thin needle), rather than an intact tissue section. Cytology sampling is generally less invasive (it involves less cutting into the body) than taking a larger tissue sample for histology. Reliability depends on the diagnostic question, the lesion and whether the sample is adequate (good enough to give an answer). Cytology can sometimes be as accurate as histology. In some cases, such as the thyroid nodule below, it can give a diagnosis. An inadequate or indeterminate sample (one that does not give a clear answer) may need repeat sampling or histology. Histology is more likely to be needed when tissue architecture (how the cells are arranged within the tissue) matters for the diagnosis.
In the assessment of a thyroid nodule (a lump in the thyroid gland), the cytology result guides what happens next. An inadequate result usually calls for repeat sampling. Indeterminate findings (results that do not clearly show whether the nodule is benign or malignant) may need further sampling or surveillance (repeat checks over time). Some people with indeterminate findings need diagnostic surgery (surgery to remove part of the thyroid so that it can be examined). The choice depends on the exact cytology result and on clinical assessment. The NICE thyroid cancer guideline (NG230), recommendation 1.2.11 and table 1, sets out the recommended next steps after initial fine-needle aspiration cytology.
Microscopic diagnosis is based on the pattern recognition skills of the pathologist, who must answer these questions:
- Is the sample normal or abnormal?
- Is it inflammatory or neoplastic (a new, abnormal growth)?
- Is it benign or malignant?
- Is it a primary tumour or a metastasis?
The Process of a Histological Report
Each step prepares the tissue so that it keeps its structure and can be cut thin, stained and examined under a microscope.
- Fixation
- The preservation of biological tissues from decay due to autolysis (the cell’s own enzymes breaking it down) or putrefaction (rotting caused by bacteria). Usually a solution of formaldehyde (a preservative chemical) is used, and the process often takes between about 6 and 72 hours, depending on the specimen.
- Cut-up
- The tissues are cut up and put into a cassette (a small slotted holder for the tissue). The cassettes are then placed in racks of formalin (a solution containing formaldehyde). Formaldehyde fixes cells by cross-linking proteins, which means it joins protein molecules to one another so that the tissue keeps its structure.
- Processing
- The samples are dehydrated (water is removed) using alcohol in a vacuum. The alcohol is then replaced with xylene (a clearing agent, which can mix with both alcohol and wax). The xylene is finally replaced with molten paraffin wax, which soaks into the tissue.
- Blocking (embedding)
- The processed tissues are removed from their cassettes and put into metal moulds, which are then filled with more molten paraffin wax. The wax is allowed to cool and the metal mould is removed. The tissues are now embedded in blocks of wax and are hard enough to be cut.
- Microtomy
- The blocks are cut into very thin sections using a machine called a microtome. The sections are usually about 3-5 micrometres (millionths of a metre) thick, which is thin enough to be seen through with a light microscope.
- Staining
- Before staining, the wax is dissolved out of the sections and they are rehydrated (water is put back). Staining is usually done with two dyes: haematoxylin and eosin (H&E staining). Haematoxylin stains nuclei purple and eosin stains the cytoplasm and connective tissues pink.
- Mounting
- The stained section is already on the slide. A mounting medium is used to attach a coverslip (a thin sheet of glass) over it. The mounting medium will dry and harden, preserving the tissue and firmly attaching the coverslip.
- Microscopy
- The slides are ready to be examined by a pathologist (a specialist in diagnosing disease from samples).
- Report writing
- Reports follow a set format: clinical history, macroscopic appearance (what the specimen looks like to the naked eye), microscopic appearance and conclusion.
If a diagnosis is needed urgently, for example during an operation, frozen sections can be used. Here, a cryostat (a machine that freezes and cuts tissue) is used to rapidly freeze the tissue after embedding it in a freezing medium, allowing rapid sectioning. Most frozen section results are given to the surgeon within about 20 minutes of the pathologist receiving the specimen.
Edited by: Dr. Bethany Turner
Reviewed by: Dr. Marcus Judge
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