What is a biopsy?

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The removal and examination of tissue and cells from the body, which is performed to establish a precise diagnosis and to determine proper treatment and prognosis.
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Indications and Procedures

Biopsy is one of the most common diagnostic tools in medicine. Illness or disease can be caused by biological agents (such as viruses, bacteria, fungi, and parasites), by physical agents (such as radiation, heat, extreme cold, and trauma), by genetic and metabolic abnormalities (such as diabetes), or by cancer, which is a new, abnormal growth commonly called a tumor. Often, however, the cause of an illness or disease may not be known. Nevertheless, the structural changes caused by the disease are characteristic enough so that the study of these alterations can give a clear picture of the nature and course of the disease. Diseases may primarily affect one organ at a time, such as hepatitis (the inflammation of the liver), or may involve many organ systems at once, as in Acquired immunodeficiency syndrome (AIDS).

The signs and symptoms of disease are not specific and are often shared by many conditions. For example, all diseases of the liver can result in jaundice, or yellowing of the skin, and abnormal blood tests. The clinician, who may be an expert in liver disease, may not be able to tell for certain whether the underlying liver condition is caused by a virus, by a toxic substance such as alcohol, or by both. A needle biopsy of the liver may then be obtained. The sample is examined by an expert surgical pathologist, and a specific diagnosis is rendered.

Similarly, a lump in the breast may be innocuous (benign) or cancerous (malignant). Only a biopsy of such a lesion can determine this conclusively. Such a biopsy can be obtained by fine needle aspiration, which is a simple procedure that can be performed in a clinic, or by excision in the operating room. A frozen section is then made for the purpose of rapid diagnosis and management.

Once a biopsy is obtained, it is placed in a special fixative, such as formalin. This solution will preserve—or fix—the internal structure of the tissue and its cells. Expert technicians in histology, called histotechnologists, will then embed the tissue in waxlike paraffin to obtain a “block.” The tissue block is then placed in a microtome, and extremely thin pieces (about 5 micrometers thick in width) are cut from it. The slices are placed on glass slides and stained with different dyes, the most common of which is hematoxylin and eosin (H & E), to delineate the cellular substructures. The slides are examined by a surgical pathologist, who renders a pathology report in which the gross and microscopic features are described, and a diagnosis is made. A differential diagnosis may also be made, in which other possible causes of disease that may give a similar histologic picture are discussed. In addition to the routine study described above, a much more extensive and expensive workup of the biopsy may be done, depending on the anticipated complexity of the condition and organ.

The study of a biopsy requires diligent preparation and staining of the tissue, which is the realm of histotechnologists. Staining refers to the application of artificial dyes to tissue sections and cells to facilitate their microscopic study. Certain tissues and cell parts have different chemical and biological affinities for dyes which, when properly applied, help demarcate and differentiate the properties of these cells. A huge battery of special stains exists that can be used to examine every aspect of cell function in both health and disease. For example, specific enzymes can be evaluated; this technique is called enzyme histochemistry.

Immunologic stains, which help evaluate the status of immune system cells, are expensive and extremely tedious, and their proper interpretation requires considerable expertise. Many antibodies are commercially available for such testing. When directed against specific antigenic cell markers, they form immune complexes that can be targeted with immunological stains. Such stains can then be evaluated by immunofluorescence or immunoperoxidase techniques. Both types employ as their principle of action the forming of complexes between antigens and antibodies and the staining of these complexes. Immunofluorescence staining techniques involve the use of special stains that cause the tissue to shine when it is viewed under a fluorescent microscope; such procedures are performed on frozen section tissues. With immunoperoxidase stains, fixed tissues are used, and the stains are permanent.

Tissue samples can also be studied with an electron microscope, in which electron beams greatly magnify subcellular structures. In this way, the alterations of specific cellular components such as cell membranes, mitochondria, and intracellular viruses can be visualized and analyzed. This ultrastructural study is especially valuable in needle biopsies of the kidneys, as well as in the study of certain unusual cancer cells.

Another highly sophisticated method used to evaluate tissue and cell function in a biopsy is the application of molecular genetics and molecular biopsy techniques. The polymerase chain reaction (PCR) involves the splitting (splicing) of a specific section of genetic material in a cell and its amplification through a chemical chain reaction into innumerable folds, so that it can be visualized through a light microscope. This type of evaluation allows for the examination of specific microorganisms in a cell and can determine the presence of certain genetic markers of unusual diseases or cancer.

Another way to study the properties of cells is by examining their genetic makeup. Karyotyping is a technique in which the actual chromosomes in a cell are photographed during mitotic divisions; the chromosomes appear as patterns of bands. Genetic abnormalities can be identified by the number of chromosomes and their appearance. This procedure is often used in the study of cancer cells. An even more sophisticated study of cellular genetic makeup is called gene rearrangement, in which the order of gene stacking is examined for specific markers of certain cancerous growths, especially of white blood cells. Other techniques that are used to evaluate cell functions and morphology are cellular imaging, in which the contours of cell membranes and surfaces are compared using computers, and the use of flow cytometers, in which cells are targeted immunologically and then counted. Both techniques are employed in cancer studies, and the second is also used for patients with abnormal immune systems, such as those with AIDS.

The aforementioned studies are expensive and available only at large medical centers and research institutes. The diagnostic workup in most hospitals, however, does not require the use of these sophisticated methods. Usually, routine H & E stains are applied. The Papanicolaou stain is commonly used with fine needle aspiration biopsies.

A department of pathology in a large medical center usually has one or more surgical pathologists, who are closely affiliated with the clinical and surgical departments and with their many branches and specialties. Interpreting biopsies obtained by any of the surgical or medical specialties is the most important duty of the surgical pathologist, and it requires great expertise and diligence. Because of the complexity of this task, specialized experts in pathology are becoming the norm. For example, a dermatopathologist is a surgical pathologist trained to interpret skin biopsies. Similarly, hematopathologists, neuropathologists, and nephropathologists are experts in the interpretation of blood-related, nerve- and brain-related, and kidney-related biopsies, respectively.

In incisional biopsies, only a portion of the lesion is sampled, and the procedure is strictly of a diagnostic nature. In excisional biopsy, the entire lesion is removed, usually with a rim of normal tissue, and therefore the procedure serves both a diagnostic and a therapeutic function. The decision whether to perform an incisional or an excisional biopsy depends primarily on the size and location of the lesion; the smaller the lesion, the more logical it is to remove it completely. It is preferable, however, to sample a deeply seated large tumor first because the type and extent of the excision varies considerably depending on the tumor type. For example, a small skin mole is usually excised completely, whereas a large soft tissue or bone tumor should be sampled.

Biopsies are also classified according to the instrument used to obtain them: cold knife versus cautery, needle, or endoscope. Of these the one usually least suitable for microscopic study is that obtained with a cautery, which uses a hot knife that burns, chars, and distorts tissue.

An endoscope is a tubelike fiber-optic instrument that is inserted into an orifice or small incision in order to view the contents of a body cavity. The instrument can be rigid or flexible and is equipped with a light source (usually a laser) and a small cutting tool at its tip to allow for the removal of small samples of tissue. Endoscopic biopsies are frequently used to obtain tissue and cell samples from the lungs and the airways, mainly to diagnose laryngeal and lung cancers; this procedure is usually done by a lung specialist. The endoscope is also used to sample lesions in the esophagus, stomach, intestines, and the rest of the intestinal tract, including the rectum. Such procedures are usually performed by a gastroenterologist, a specialist in the stomach and the gastrointestinal tract. Endoscopic biopsies of the urinary bladder and the prostate are done by urologists.

Needle biopsies are commonly used to obtain samples from superficial or deep-seated lumps. A slender, cylindrical core of tissue, corresponding to the open diameter of the needle, is obtained. The needle biopsy is commonly used to obtain tissue samples from kidneys, bone, and the deep viscera such as the liver. The modified technique of aspiration cytology, commonly called fine needle aspiration, employs a fine-caliber needle (0.6 to 0.9 millimeter in open diameter) and is widely used to obtain cytologic and minute tissue samples, especially for lesions of the lymph glands, breasts, thyroid gland, salivary glands, lungs, and prostate. Fine needle aspiration is often inexpensive, safe, quick, and, when performed and interpreted by experienced workers, quite accurate. Because of the ready availability and relative inexpense of the endoscopic and fine needle aspiration biopsy techniques, they have become popular; almost every part of the body is now within reach of one or another of these two techniques.

Frozen section biopsy requires great expertise because this biopsy is usually a form of consultation done during surgery. A tissue sample is instantly frozen, sectioned, stained, and examined—all within about fifteen minutes—in order to render a specific diagnosis. The implications of this diagnosis are far reaching and will influence the surgical procedure and the long-term therapy and outcome for the patient. A frozen section report, for example, may determine whether an organ such as a breast, lung, or kidney must be removed and whether long-term radiation therapy or chemotherapy will be administered; such would be the case if the diagnosis is read as malignant.

There are two indications, other than establishing a diagnosis, for performing a frozen section: determining the adequacy of the margins of surgical excision (for example, to remove a malignant tumor completely) and establishing whether the tissue obtained contains an ample diagnosable sample to carry out other specialized tissue studies.

Uses and Complications

The following examples illustrate the practical use of the various biopsy techniques.

An excisional biopsy is performed on a pigmented dark lesion on a sun-exposed surface of the body of a young man and is diagnosed as malignant melanoma, which is a tumor of the pigment-producing cells of the body. This diagnosis is confirmed through the use of specialized immunological stains employing specific antibodies against melanin. The pathologist also comments that surgical margins of excision of that tumor are safe and do not contain tumor, and that the tumor is only superficial in nature and does not show deep invasion into the tissue. These two points imply that the patient will probably have a complete cure.

A fine needle aspiration biopsy is applied on a lump on the breast of a young woman. The material obtained is spread on slides, stained with a Papanicolaou stain, and evaluated within hours of its removal. The lump is diagnosed as a fibroadenoma, which is a benign tumor that is completely innocuous and of no further consequence to the young patient.

An elderly patient has an endoscopic biopsy of a visualized mass in the colon, which proves to be cancer. The patient is taken to the operating room, and the colon is resected. A frozen section is performed on the margin on the surgical excision to make sure that it contains no tumor. The stains used in this example are the simple and routine H & E stains.

A liver biopsy is performed on a patient with jaundice (yellowing of the skin), and a diagnosis of viral hepatitis B is made. This diagnosis is made following the study of the liver biopsy by routine stains and by stains that use immunological antibodies against the viral antigen. This is a specific and highly accurate diagnostic study.

A lymph gland excisional biopsy is performed on a patient who feels lumps all over his body. The biopsy is examined with routine and special stains, immunological marker studies, and gene rearrangement. Such extensive studies are performed to make sure that his condition is completely benign and is not neoplastic—that is, that he does not have malignant lymphoma (cancer of lymph tissue).

The biopsy, in its varied forms and techniques, has become an essential component of quality medical care. The biopsy report is both a medical and a legal document. Tissue slides and blocks are often stored for many years, in some places indefinitely. Peer slide reviews and consultations are common and are used as gauges for quality control and management. There are some limitations with histologic diagnosis, which mainly revolve around recognizing a specialist’s own limitations and the need to seek a consultation by another expert pathologist as needed.

Perspective and Prospects

The first attempts to affirm the role of biopsy in medical practice were made by Carl Ruge and Johann Veit of the University of Berlin, who in the 1870s introduced surgical biopsy as an essential diagnostic tool. Despite the inevitable controversies that followed, Johann F. A. von Esmarch, a professor of surgery and a leading military surgeon of his time, presented forceful arguments at the German Surgical Congress of 1889 on the need to establish a microscopic diagnosis before operating in suspected cases of malignant tumors requiring extensive mutilating procedures. Shortly thereafter, the freezing microtome was introduced for the purpose of creating frozen sections, which hastened the acceptance of this recommendation. In the United States, the specialty of surgical pathology, which deals with biopsy interpretations, was created by a collaboration between surgeons and a gynecologist.

It is said that the first full-fledged American surgical pathologist was Joseph Colt Bloodgood, in the division of surgical pathology created by W. S. Halsted of Johns Hopkins Hospital in Baltimore. The founders of modern American surgical pathology and biopsy interpretation are Arthur Purdy Stout of Columbia Presbyterian Hospital in New York, James Ewing and his successors Fred Stewart and Frank Foote of Memorial Hospital in New York City, and Pierre Masson of the University of Montreal.

The biopsy remains the cornerstone for modern medical management, especially in cancer therapy. It will continue to enjoy that role in diagnostics as more venues become established to obtain, study, and evaluate minute tissue and cytologic samples.

Bibliography

Bancroft, John D., and Marilyn Gamble, eds. Theory and Practice of Histological Techniques. 6th ed. New York: Churchill Livingstone/Elsevier, 2008.

Dubowitz, Victor, Caroline A. Sewry, and Anders Oldfors. Muscle Biopsy: A Practical Approach. Oxford: Saunders, 2013.

Fisher, Cyril, Elizabeth A. Montgomery, and Khin Thway. Biopsy Interpretation of Soft Tissue Tumors. Philadelphia: Wolters Kluwer/Lippincott, Williams and Wilkins, 2011.

Koss, Leopold G., Stanisław Woyke, and Włodzimierz Olszewski. Aspiration Biopsy: Cytologic Interpretation and Histologic Basis. 2d ed. New York: Igaku-Shoin, 1992.

Mills, Stacey E., et al., eds. Sternberg’s Diagnostic Surgical Pathology. 5th ed. 2 vols. Philadelphia: Wolters Kluwer Health/Lippincott Williams & Wilkins, 2010.

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Shah, Rajal B., and Ming Zhou. Prostate Biopsy Interpretation: An Illustrated Guide. New York: Springer-Verlag, 2012.

Sloan, John P. Biopsy Pathology of the Breast. 2d ed. New York: Oxford University Press, 2001.

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Yazdi, Hossein M., and Irving Dardick. Diagnostic Immunocytochemistry and Electron Microscopy: Guides to Clinical Aspiration Biopsy. New York: Igaku-Shoin, 1992.