Low-grade squamous intraepithelial lesion

Papanicolaou stain (also Papanicolaou's stain and Pap stain) is a multichromatic (multicolored)

cytological Cell biology (also cellular biology or cytology) is a branch of biology that studies the structure, function, and behavior of cells. All living organisms are made of cells. A cell is the basic unit of life that is responsible for the living and ...

staining Staining is a technique used to enhance contrast in samples, generally at the microscopic level. Stains and dyes are frequently used in histology (microscopic study of biological tissues), in cytology (microscopic study of cells), and in the ...

technique developed by George Papanicolaou in 1942. The Papanicolaou stain is one of the most widely used stains in

cytology Cell biology (also cellular biology or cytology) is a branch of biology that studies the structure, function, and behavior of cells. All living organisms are made of cells. A cell is the basic unit of life that is responsible for the living and ...

, where it is used to aid

pathologists A list of people notable in the field of pathology. A * John Abercrombie, Scottish physician, neuropathologist and philosopher. * Maude Abbott (1869–1940), Canadian pathologist, one of the earliest women graduated in medicine, expert in co ...

in making a diagnosis. Although most notable for its use in the detection of

cervical cancer Cervical cancer is a cancer arising from the cervix. It is due to the abnormal growth of cells that have the ability to invade or spread to other parts of the body. Early on, typically no symptoms are seen. Later symptoms may include abnormal ...

in the

Pap test The Papanicolaou test (abbreviated as Pap test, also known as Pap smear (AE), cervical smear (BE), cervical screening (BE), or smear test (BE)) is a method of cervical screening used to detect potentially precancerous and cancerous processes in t ...

Pap smear The Papanicolaou test (abbreviated as Pap test, also known as Pap smear (AE), cervical smear (BE), cervical screening (BE), or smear test (BE)) is a method of cervical screening used to detect potentially precancerous and cancerous processes in t ...

, it is also used to stain non-

gynecological Gynaecology or gynecology (see spelling differences) is the area of medicine that involves the treatment of women's diseases, especially those of the reproductive organs. It is often paired with the field of obstetrics, forming the combined area ...

specimen preparations from a variety of bodily secretions and from small needle biopsies of organs and tissues. Papanicolaou published three formulations of this stain in 1942, 1954, and 1960.

Usage

Pap staining is used to differentiate cells in smear preparations (in which samples are spread or smeared onto a glass microscope slide) from various bodily secretions and needle biopsies; the specimens may include gynecological smears (

s),

sputum Sputum is mucus that is coughed up from the lower airways (the trachea and bronchi). In medicine, sputum samples are usually used for a naked eye examination, microbiological investigation of respiratory infections and cytological investigations ...

, brushings, washings,

urine Urine is a liquid by-product of metabolism in humans and in many other animals. Urine flows from the kidneys through the ureters to the urinary bladder. Urination results in urine being excretion, excreted from the body through the urethra. Cel ...

cerebrospinal fluid Cerebrospinal fluid (CSF) is a clear, colorless body fluid found within the tissue that surrounds the brain and spinal cord of all vertebrates. CSF is produced by specialised ependymal cells in the choroid plexus of the ventricles of the bra ...

, abdominal fluid,

pleural fluid The pleural cavity, pleural space, or interpleural space is the potential space between the pleurae of the pleural sac that surrounds each lung. A small amount of serous pleural fluid is maintained in the pleural cavity to enable lubrication be ...

synovial fluid Synovial fluid, also called synovia, elp 1/sup> is a viscous, non-Newtonian fluid found in the cavities of synovial joints. With its egg white–like consistency, the principal role of synovial fluid is to reduce friction between the articular ...

seminal fluid Semen, also known as seminal fluid, is an organic bodily fluid created to contain spermatozoa. It is secreted by the gonads (sexual glands) and other sexual organs of male or hermaphroditic animals and can fertilize the female ovum. Semen i ...

fine needle aspiration Fine-needle aspiration (FNA) is a diagnostic procedure used to investigate lumps or masses. In this technique, a thin (23–25 gauge (0.52 to 0.64 mm outer diameter)), hollow needle is inserted into the mass for sampling of cells that, aft ...

s, tumor touch samples, or other materials containing loose cells. The pap stain is not fully standardized and comes in several formulations, differing in the exact dyes used, their ratios, and timing of the process. Pap staining is usually associated with

cytopathology Cytopathology (from Greek , ''kytos'', "a hollow"; , ''pathos'', "fate, harm"; and , '' -logia'') is a branch of pathology that studies and diagnoses diseases on the cellular level. The discipline was founded by George Nicolas Papanicolaou in 1 ...

in which loose cells are examined, but the stain has also been modified and used on tissue slices.

Pap test

Pap staining is used in the

(or Pap test) and is a reliable technique in

screening Screening may refer to: * Screening cultures, a type a medical test that is done to find an infection * Screening (economics), a strategy of combating adverse selection (includes sorting resumes to select employees) * Screening (environmental), a ...

gynecology Gynaecology or gynecology (see spelling differences) is the area of medicine that involves the treatment of women's diseases, especially those of the reproductive organs. It is often paired with the field of obstetrics, forming the combined are ...

Generalized staining method

The classic form of the Papanicolaou stain involves five

stains A stain is an unwanted localized discoloration, often in fabrics or textiles. Stain(s) or The Stain(s) may also refer to: Color * Stain (heraldry), a non-standard tincture * Staining, in biology, a technique used to highlight contrast in samples ...

in three solutions. * The first staining solution contains

haematoxylin Haematoxylin or hematoxylin (), also called natural black 1 or C.I. 75290, is a compound extracted from heartwood of the logwood tree (''Haematoxylum campechianum'') with a chemical formula of . This naturally derived dye has been used as a ...

which stains

cell nuclei The cell nucleus (pl. nuclei; from Latin or , meaning ''kernel'' or ''seed'') is a membrane-bound organelle found in eukaryotic cells. Eukaryotic cells usually have a single nucleus, but a few cell types, such as mammalian red blood cells, h ...

. Papanicolaou used Harris's hematoxylin in all three formulations of the stain he published. * The second staining solution (designated OG-6), contains

Orange G Orange G also called C.I. 16230, Acid Orange 10, or orange gelb is a synthetic azo dye used in histology in many staining formulations. It usually comes as a disodium salt. It has the appearance of orange crystals or powder. Staining Orange G ...

in 95%

ethyl alcohol Ethanol (abbr. EtOH; also called ethyl alcohol, grain alcohol, drinking alcohol, or simply alcohol) is an organic compound. It is an alcohol with the chemical formula . Its formula can be also written as or (an ethyl group linked to a hyd ...

with a small amount of

phosphotungstic acid Phosphotungstic acid (PTA) or tungstophosphoric acid (TPA), is a heteropoly acid with the chemical formula . It forms hydrates . It is normally isolated as the ''n'' = 24 hydrate but can be desiccated to the hexahydrate (''n'' = 6). EPTA is the na ...

. In the OG-6, the OG signifies Orange G and the '6' denotes the concentration of

added; other variants are OG-5 and OG-8). * The third staining solution is composed of three dyes,

Eosin Y Eosin Y, also called C.I. 45380 or C.I. Acid Red 87, is a member of the triarylmethane dyes. It is produced from fluorescein by bromination. Use Eosin Y is commonly used as the red dye in red inks. It is commonly used in histology, most nota ...

Light Green SF yellowish Light green SF, also called C.I. 42095, light green SF yellowish, is a green triarylmethane dye. Uses Biomedical It is used in histology for staining collagen; for that purpose it is a standard dye in North America. In Masson's trichrome i ...

, and

Bismarck brown Y Bismarck brown Y also called C.I. 21000 and C.I. Basic Brown 1, is a diazo dye with the idealized formula H2N)2C6H3N2sub>2C6H4. The dye is a mixture of closely related compounds. It was one of the earliest azo dyes, being described in 1863 by ...

in 95% ethyl alcohol with a small amount of phosphotungstic acid and

lithium carbonate Lithium carbonate is an inorganic compound, the lithium salt (chemistry), salt of carbonate with the chemical formula, formula . This white Salt (chemistry), salt is widely used in the processing of metal oxides. It is listed on the World Health O ...

. This solution, designated EA, followed by a number which denotes the proportion of the dyes, other formulations include EA-36, EA-50, and EA-65. The counterstains are dissolved in 95% ethyl alcohol which prevents cells from over staining which would obscure nuclear detail and cell outlines especially in the case when cells are overlapping on the slide. Phosphotungstic acid is added to adjust the pH of counterstains and helps to optimize the color intensity. The EA counterstain contains Bismarck brown and phosphotungstic acid, which when in combination, cause both to

precipitate In an aqueous solution, precipitation is the process of transforming a dissolved substance into an insoluble solid from a super-saturated solution. The solid formed is called the precipitate. In case of an inorganic chemical reaction leading ...

out of solution, reducing the useful life of the mixture.

Results

The stain should result in cells that are fairly transparent so even thicker specimens with overlapping cells can be interpreted. Cell nuclei should be crisp, blue to black on color and the

chromatin Chromatin is a complex of DNA and protein found in eukaryotic cells. The primary function is to package long DNA molecules into more compact, denser structures. This prevents the strands from becoming tangled and also plays important roles in r ...

patterns of the nucleus should be well defined. Cell

cytoplasm In cell biology, the cytoplasm is all of the material within a eukaryotic cell, enclosed by the cell membrane, except for the cell nucleus. The material inside the nucleus and contained within the nuclear membrane is termed the nucleoplasm. The ...

stains blue-green and

keratin Keratin () is one of a family of structural fibrous proteins also known as ''scleroproteins''. Alpha-keratin (α-keratin) is a type of keratin found in vertebrates. It is the key structural material making up scales, hair, nails, feathers, ho ...

stains orange in color.

stains the superficial epithelial

squamous cell Epithelium or epithelial tissue is one of the four basic types of animal tissue, along with connective tissue, muscle tissue and nervous tissue. It is a thin, continuous, protective layer of compactly packed cells with a little intercellula ...

nucleoli The nucleolus (, plural: nucleoli ) is the largest structure in the nucleus of eukaryotic cells. It is best known as the site of ribosome biogenesis, which is the synthesis of ribosomes. The nucleolus also participates in the formation of s ...

cilia The cilium, plural cilia (), is a membrane-bound organelle found on most types of eukaryotic cell, and certain microorganisms known as ciliates. Cilia are absent in bacteria and archaea. The cilium has the shape of a slender threadlike projecti ...

, and

red blood cell Red blood cells (RBCs), also referred to as red cells, red blood corpuscles (in humans or other animals not having nucleus in red blood cells), haematids, erythroid cells or erythrocytes (from Greek ''erythros'' for "red" and ''kytos'' for "holl ...

confers a blue staining for the

of active cells such as columnar cells, parabasal squamous cells, and intermediate squamous cells. Superficial cells are orange to pink, and intermediate and parabasal cells are turquoise green to blue.

Ultrafast Papanicolaou stain

Ultrafast Papanicolaou stain is an alternative for the fine needle aspiration samples, developed to achieve comparable visual clarity in significantly shorter time. The process differs in rehydration of the air-dried smear with saline, use 4%

formaldehyde Formaldehyde ( , ) (systematic name methanal) is a naturally occurring organic compound with the formula and structure . The pure compound is a pungent, colourless gas that polymerises spontaneously into paraformaldehyde (refer to section F ...

in 65%

ethanol Ethanol (abbr. EtOH; also called ethyl alcohol, grain alcohol, drinking alcohol, or simply alcohol) is an organic compound. It is an Alcohol (chemistry), alcohol with the chemical formula . Its formula can be also written as or (an ethyl ...

fixative, and use of Richard-Allan

Hematoxylin Haematoxylin or hematoxylin (), also called natural black 1 or C.I. 75290, is a compound extracted from heartwood of the logwood tree (''Haematoxylum campechianum'') with a chemical formula of . This naturally derived dye has been used as a h ...

-2 and Cyto-Stain, resulting in a 90-second process yielding transparent polychromatic stains.

Examples of Papanicolaou stain

Papers by George N. Papanicolaou describing his stain

* Papanicolaou, George N. "A new procedure for staining vaginal smears." Science 95.2469 (1942): 438–439. * Papanicolaou, George N. "The cell smear method of diagnosing cancer." American Journal of Public Health and the Nation's Health 38.2 (1948): 202–205. * Papanicolaou, George N. "Atlas of exfoliative cytology." Published for the commonwealth fund by Harvard University Press. (1954). * Papanicolaou, George N. "Memorandum on staining." Atlas of exfoliative cytology. Cambridge, MA: Harvard University Press, Supplement II (1960): 12.

References

{{Human papillomavirus Staining Cytopathology Microbiology techniques Laboratory techniques Histopathology Staining dyes sr:Papanikolau test