Histology, also known as microscopic anatomy or microanatomy, is the branch of
biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecular interactions, Physiology, physiological mechanisms, Development ...

which studies the microscopic
anatomy Anatomy (Greek ''anatomē'', 'dissection') is the branch of concerned with the study of the structure of s and their parts. Anatomy is a branch of natural science which deals with the structural organization of living things. It is an old scie ...

of biological tissues. Histology is the microscopic counterpart to
gross anatomy Gross anatomy is the study of anatomy Anatomy (Greek ''anatomē'', 'dissection') is the branch of biology concerned with the study of the structure of organism In biology, an organism (from Ancient Greek, Greek: ὀργανισμός, ' ...
, which looks at larger structures visible without a
microscope A microscope (from grc, μικρός ''mikrós'' 'small' and ''skopeîn'' 'to look (at); examine, inspect') is a used to examine objects that are too small to be seen by the . is the of investigating small objects and structures using a ...

. Although one may divide microscopic anatomy into ''organology'', the study of organs, ''histology'', the study of tissues, and ''
cytology Cell biology (also cellular biology or cytology) is a branch of biology studying the structure and function of the cell, also known as the basic unit of life Life is a characteristic that distinguishes physical entities that have bi ...
'', the study of
cells Cell most often refers to: * Cell (biology), the functional basic unit of life Cell may also refer to: Closed spaces * Monastic cell, a small room, hut, or cave in which a monk or religious recluse lives * Prison cell, a room used to hold peopl ...
, modern usage places all of these topics under the field of histology. In
medicine Medicine is the science Science () is a systematic enterprise that builds and organizes knowledge Knowledge is a familiarity, awareness, or understanding of someone or something, such as facts ( descriptive knowledge), skills (proced ...

histopathology Histopathology (compound of three Greek language, Greek words: ''histos'' "tissue", πάθος ''pathos'' "suffering", and -λογία ''-logy, -logia'' "study of") refers to the light microscope, microscopic examination of Tissue (biology), tiss ...
is the branch of histology that includes the microscopic identification and study of diseased tissue. In the field of
paleontology Paleontology (), also spelled palaeontology or palæontology, is the scientific study of life that existed prior to, and sometimes including, the start of the Holocene epoch (geology), epoch (roughly 11,700 years before present). It includes th ...
, the term paleohistology refers to the histology of
fossil A fossil (from Classical Latin Classical Latin is the form of Latin, Latin language recognized as a Literary language, literary standard language, standard by writers of the late Roman Republic and early Roman Empire. It was used from 75 ...


Biological tissues

Animal tissue classification

There are four basic types of animal tissues:
muscle tissue Muscle tissues are soft tissue of a tendon A tendon or sinew is a tough band of fibrous connective tissue that connects muscle to bone A bone is a Stiffness, rigid tissue (anatomy), tissue that constitutes part of the vertebrate skeleton i ...

muscle tissue
nervous tissue Nervous tissue, also called neural tissue, is the main tissue component of the nervous system In Biology, biology, the nervous system is a Complex system, highly complex part of an animal that coordinates its Behavior, actions and Sense, se ...
connective tissue Connective tissue is one of the many basic types of animal Animals (also called Metazoa) are multicellular eukaryotic organisms that form the Kingdom (biology), biological kingdom Animalia. With few exceptions, animals Heterotroph, consum ...
, and
epithelial tissue Epithelium () is one of the four basic types of animal Animals (also called Metazoa) are multicellular eukaryotic organisms that form the Kingdom (biology), biological kingdom Animalia. With few exceptions, animals Heterotroph, consume ...
. All animal tissues are considered to be subtypes of these four principal tissue types (for example, blood is classified as connective tissue, since the blood cells are suspended in an
extracellular matrix In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecular interactions, Physiology, physiological mechanisms, ...
, the
plasma Plasma or plasm may refer to: Science * Plasma (physics), one of the four fundamental states of matter * Plasma (mineral) or heliotrope, a mineral aggregate * Quark–gluon plasma, a state of matter in quantum chromodynamics Biology * Blood plasma ...

Plant tissue classification

For plants, the study of their tissues falls under the field of
plant anatomy Plant anatomy or phytotomy is the general term for the study of the internal structure A structure is an arrangement and organization of interrelated elements in a material object or system A system is a group of Interaction, interacting or ...
, with the following four main types: * Dermal tissue *
Vascular tissue stalk, showing vascular bundles, which include both phloem and xylem. Image:BrambleLeaf_CrossPolarisedLight_Diagram.jpg">250px, Detail of the vasculature of a tissue Tissue may refer to: Biology * Tissue (biology), an ensemble of similar cells t ...
Ground tissue The ground tissue of plants includes all tissues that are neither dermal The dermis or corium is a layer of skin Skin is the layer of usually soft, flexible outer tissue covering the body of a vertebrate animal, with three main functions: prot ...
* Meristematic tissue

Medical histology

Histopathology is the branch of histology that includes the microscopic identification and study of diseased tissue. It is an important part of anatomical pathology and surgical pathology, as accurate diagnosis of cancer and other diseases often requires histopathological examination of tissue samples. Trained physicians, frequently licensed pathologists, perform histopathological examination and provide diagnostic information based on their observations.


The field of histology that includes the preparation of tissues for microscopic examination is known as histotechnology. Job titles for the trained personnel who prepare histological specimens for examination are numerous and include histotechnicians, histotechnologists, histology technicians and technologists, Medical laboratory assistant, medical laboratory technicians, and biomedical scientists.

Sample preparation

Most histological samples need preparation before microscopic observation; these methods depend on the specimen and method of observation.


Chemical Fixation (histology), fixatives are used to preserve and maintain the structure of tissues and cells; fixation also hardens tissues which aids in cutting the thin sections of tissue needed for observation under the microscope. Fixatives generally preserve tissues (and cells) by irreversibly cross-linking proteins. The most widely used fixative for light microscopy is 10% neutral buffered formalin, or NBF (4% formaldehyde in phosphate buffered saline). For electron microscopy, the most commonly used fixative is glutaraldehyde, usually as a 2.5% solution in phosphate buffered saline. Other fixatives used for electron microscopy are osmium tetroxide or uranyl acetate. The main action of these aldehyde fixatives is to cross-link amino groups in proteins through the formation of methylene bridges (-CH2-), in the case of formaldehyde, or by C5H10 cross-links in the case of glutaraldehyde. This process, while preserving the structural integrity of the cells and tissue can damage the biological functionality of proteins, particularly enzymes. Formalin fixation leads to degradation of mRNA, miRNA, and DNA as well as denaturation and modification of proteins in tissues. However, extraction and analysis of nucleic acids and proteins from formalin-fixed, paraffin-embedded tissues is possible using appropriate protocols.

Selection and trimming

''Selection'' is the choice of relevant tissue in cases where it is not necessary to put the entire original tissue mass through further processing. The remainder may remain fixated in case it needs to be examined at a later time. ''Trimming'' is the cutting of tissue samples in order to expose the relevant surfaces for later sectioning. It also creates tissue samples of appropriate size to fit into cassettes.


Tissues are embedded in a harder medium both as a support and to allow the cutting of thin tissue slices. In general, water must first be removed from tissues (dehydration) and replaced with a medium that either solidifies directly, or with an intermediary fluid (clearing) that is miscible with the embedding media.

Paraffin wax

For light microscopy, paraffin wax is the most frequently used embedding material. Paraffin is immiscible with water, the main constituent of biological tissue, so it must first be removed in a series of dehydration steps. Samples are transferred through a series of progressively more concentrated ethanol baths, up to 100% ethanol to remove remaining traces of water. Dehydration is followed by a ''clearing agent'' (typically xylene although other environmental safe substitutes are in use) which removes the alcohol and is miscible with the wax, finally melted paraffin wax is added to replace the xylene and infiltrate the tissue. In most histology, or histopathology laboratories the dehydration, clearing, and wax infiltration are carried out in ''tissue processors'' which automate this process. Once infiltrated in paraffin, tissues are oriented in molds which are filled with wax; once positioned, the wax is cooled, solidifying the block and tissue.

Other materials

Paraffin wax does not always provide a sufficiently hard matrix for cutting very thin sections (which are especially important for electron microscopy). Paraffin wax may also be too soft in relation to the tissue, the heat of the melted wax may alter the tissue in undesirable ways, or the dehydrating or clearing chemicals may harm the tissue. Alternatives to paraffin wax include, epoxy, Poly(methyl methacrylate), acrylic, agar, gelatin, Micro technique#Celloidin Method, celloidin, and other types of waxes. In electron microscopy epoxy resins are the most commonly employed embedding media, but acrylic resins are also used, particularly where immunohistochemistry is required. For tissues to be cut in a frozen state, tissues are placed in a water-based embedding medium. Pre-frozen tissues are placed into molds with the liquid embedding material, usually a water-based glycol, Optimal cutting temperature compound, OCT, Tris-buffered saline, TBS, Cryogel, or resin, which is then frozen to form hardened blocks.


For light microscopy, a knife mounted in a microtome is used to cut tissue sections (typically between 5-15 micrometre, micrometers thick) which are mounted on a glass microscope slide. For transmission electron microscopy (TEM), a diamond or glass knife mounted in an ultramicrotome is used to cut between 50-150 nanometre, nanometer thick tissue sections.


Biological tissue has little inherent contrast in either the light or electron microscope. Staining is employed to give both contrast to the tissue as well as highlighting particular features of interest. When the stain is used to target a specific chemical component of the tissue (and not the general structure), the term histochemistry is used.

Light microscopy

Hematoxylin and eosin (H&E stain) is one of the most commonly used stains in histology to show the general structure of the tissue. Hematoxylin stains cell cell nucleus, nuclei blue; eosin, an Acid (chemistry), acidic dye, stains the cytoplasm and other tissues in different stains of pink. In contrast to H&E, which is used as a general stain, there are many techniques that more selectively stain cells, cellular components, and specific substances. A commonly performed histochemical technique that targets a specific chemical is the Perls' Prussian blue reaction, used to demonstrate iron deposits in diseases like hemochromatosis. The Nissl method for Nissl substance and Golgi's method (and related silver stains) are useful in identifying neurons are other examples of more specific stains.


In historadiography, a slide (sometimes stained histochemically) is X-rayed. More commonly, autoradiography is used in visualizing the locations to which a radioactive substance has been transported within the body, such as cells in S phase (undergoing DNA replication) which incorporate tritiated thymidine, or sites to which radiolabeled nucleic acid probes bind in in situ hybridization. For autoradiography on a microscopic level, the slide is typically dipped into liquid nuclear tract emulsion, which dries to form the exposure film. Individual silver grains in the film are visualized with dark field microscopy.


Recently, antibodies have been used to specifically visualize proteins, carbohydrates, and lipids. This process is called immunohistochemistry, or when the stain is a fluorescent molecule, immunofluorescence. This technique has greatly increased the ability to identify categories of cells under a microscope. Other advanced techniques, such as nonradioactive ''in situ'' hybridization, can be combined with immunochemistry to identify specific DNA or RNA molecules with fluorescent probes or tags that can be used for immunofluorescence and enzyme-linked fluorescence amplification (especially alkaline phosphatase and tyramide signal amplification). Fluorescence microscopy and confocal microscopy are used to detect fluorescent signals with good intracellular detail.

Electron microscopy

For electron microscopy heavy metals are typically used to stain tissue sections. Uranyl acetate and lead citrate are commonly used to impart contrast to tissue in the electron microscope.

Specialized techniques


Similar to the frozen section procedure employed in medicine, cryosectioning is a method to rapidly freeze, cut, and mount sections of tissue for histology. The tissue is usually sectioned on a cryostat or freezing microtome. The frozen sections are mounted on a glass slide and may be stained to enhance the contrast between different tissues. Unfixed frozen sections can be used for studies requiring enzyme localization in tissues and cells. Tissue fixation is required for certain procedures such as antibody-linked immunofluorescence staining. Frozen sections are often prepared during surgical removal of tumors to allow rapid identification of tumor margins, as in Mohs surgery, or determination of tumor malignancy, when a tumor is discovered incidentally during surgery.


Ultramicrotomy is a method of preparing extremely thin sections for transmission electron microscope (TEM) analysis. Tissues are commonly embedded in epoxy or other plastic resin. Very thin sections (less than 0.1 micrometer in thickness) are cut using diamond or glass knives on an ultramicrotome.


Artifacts are structures or features in tissue that interfere with normal histological examination. Artifacts interfere with histology by changing the tissues appearance and hiding structures. Tissue processing artifacts can include pigments formed by fixatives, shrinkage, washing out of cellular components, color changes in different tissues types and alterations of the structures in the tissue. An example is mercury pigment left behind after using Zenker's fixative to fix a section. Formalin fixation can also leave a brown to black pigment under acidic conditions.


In the 17th century the Italian Marcello Malpighi used microscopes to study tiny biological entities; some regard him as the founder of the fields of histology and microscopic pathology. Malpighi analyzed several parts of the organs of bats, frogs and other animals under the microscope. While studying the structure of the lung, Malpighi noticed its membranous alveoli and the hair-like connections between veins and arteries, which he named capillaries. His discovery established how the oxygen breathed in enters the blood stream and serves the body. In the 19th century histology was an academic discipline in its own right. The French anatomist Xavier Bichat introduced the concept of Tissue (biology), tissue in anatomy in 1801, and the term "histology" ( de , Histologie), coined to denote the "study of tissues", first appeared in a book by August Franz Josef Karl Mayer, Karl Meyer in 1819. Bichat described twenty-one human tissues, which can be subsumed under the four categories currently accepted by histologists. The usage of illustrations in histology, deemed as useless by Bichat, was promoted by Jean Cruveilhier. In the early 1830s Jan Evangelista Purkyně, Purkynĕ invented a microtome with high precision. During the 19th century many Fixation (histology), fixation techniques were developed by Adolph Hannover (solutions of Chromate and dichromate, chromates and chromic acid), Franz Eilhard Schulze, Franz Schulze and Max Schultze (osmic acid), Alexander Butlerov (formaldehyde) and Benedikt Stilling (Frozen section procedure, freezing). Microscope slide#Mounting, Mounting techniques were developed by Rudolf Heidenhain (1824-1898), who introduced gum Arabic; Salomon Stricker (1834-1898), who advocated a mixture of wax and oil; and Andrew Pritchard (1804-1884) who, in 1832, used a gum/isinglass mixture. In the same year, Canada balsam appeared on the scene, and in 1869 Edwin Klebs (1834-1913) reported that he had for some years embedded his specimens in paraffin. The 1906 Nobel Prize in Physiology or Medicine was awarded to histologists Camillo Golgi and Santiago Ramon y Cajal. They had conflicting interpretations of the neural structure of the brain based on differing interpretations of the same images. Ramón y Cajal won the prize for his correct theory, and Golgi for the silver staining, silver-staining Golgi's method, technique that he invented to make it possible.

Future directions

''In vivo'' histology

Currently there is intense interest in developing techniques for ''in vivo'' histology (predominantly using MRI), which would enable doctors to non-invasively gather information about healthy and diseased tissues in living patients, rather than from fixed tissue samples.



{{Authority control Histology, Histotechnology Staining Histochemistry Anatomy