Pap Smear

Updated: May 31, 2018
Author: Nicole W Karjane, MD; Chief Editor: Christine Isaacs, MD 


Worldwide, approximately 500,000 new cases of cervical cancer and 274,000 deaths are attributable to cervical cancer yearly, making cervical cancer the second most common cause of death from cancer in women.[1] Fortunately, the incidence of cervical cancer has decreased by more than 50% in the past 30+ years, largely due to the increasing use of cervical cancer screening with cervical cytology.[2]

The American Cancer Society estimates about 13,240 new cases of invasive cervical cancer will be diagnosed and about 4170 women will die from cervical cancer in 2018.[3] Although worldwide cervical cancer rates have decreased dramatically with the increase in screening efforts, incidence and prevalence in developing countries remains high due to lack of screening programs, with approximately 80% of all cervical cancer deaths occurring in the developing world.[1]

The mainstay of cervical cancer screening for the last 60+ years has been the Papanicolaou test. The Papanicolaou test, also known as the Pap test or the Pap smear, was developed in the 1940s by Georgios Papanikolaou. It involves exfoliating cells from the transformation zone of the cervix to enable examination of these cells microscopically for detection of cancerous or precancerous lesions.

An image depicting a Pap smear can be seen below.

Pap smear. Pap smear.

In the technique known as liquid-based cytology, these collected cells are released into a vial of liquid preservative that is then used in the cytology lab to produce a slide for microscopic evaluation of the cells. The older, traditional Pap technique involves direct transfer of the cervical cells to a microscope slide for evaluation. Although the traditional method may introduce confounders such as blood and other debris to the slide, which may make interpretation more difficult, both conventional cytology and liquid-based cytology have been shown to have similar sensitivity and specificity for moderate dysplasia or worse lesions when using a threshold of LSIL or higher. In addition, both types of cytological screening are considered acceptable by the American College of Obstetricians and Gynecologists.[2]

When abnormal cells are detected on the Pap Test, diagnostic testing in the form of colposcopy is often indicated. This testing may be followed by diagnosis of dysplasia via colposcopic biopsies. Subsequent cervical cancer may be prevented through the diagnosis and treatment of these cervical cancer precursors.

Evidence shows that approximately 99-100% of cervical cancers are attributable to infection by high-risk types of the human papillomavirus (HPV). HPV represents a family of double-stranded, circular DNA viruses that can infect skin or mucosal cells, including the anogenital region and the oral cavity, and may be transmitted easily via sexual intercourse or direct contact.[4, 5]

More than 100 types of HPV exist, 12 of which can involve the anogenital region and are considered "high risk" or oncogenic in nature. These include HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, and 59. Of these, HPV 16 is responsible for the largest number of CIN 3 and cervical cancer cases, and HPV 16 and 18 combined are thought to be responsible for nearly 70% of all cases of cervical cancer.[4] Although HPV is a necessary factor in the development of cervical dysplasia that can eventually lead to cervical cancer, most women infected with HPV will not develop cervical dysplasia.[6] The presence of high-risk HPV DNA is accompanied by cytologic abnormalities approximately one third of the time. Whether an HPV infection will progress relates to the persistence of the infection and also possibly to the immune response and smoking status of the woman.[7]

Relevant Anatomy

The female reproductive organs can be subdivided into the internal and external genitalia. The internal genitalia are those organs that are within the true pelvis. These include the vagina, uterus, cervix, uterine tubes (oviducts or fallopian tubes), and ovaries. The external genitalia lie outside the true pelvis. These include the perineum, mons pubis, clitoris, urethral (urinary) meatus, labia majora and minora, vestibule, greater vestibular (Bartholin) glands, Skene glands, and periurethral area.

The cervix is the inferior portion of the uterus, separating the body of the uterus from the vagina. The cervix is cylindrical in shape, with an endocervical canal located in the midline, allowing passage of semen into the uterus. The external opening into the vagina is termed the external os, and the internal opening into the endometrial cavity is termed the internal os. The internal os is the portion of a female cervix that dilates to allow delivery of the fetus during labor. The average length of the cervix is 3-5 cm.

For more information about the relevant anatomy, see Female Reproductive Organ Anatomy. Also see Uterus Anatomy and Vaginal Anatomy.



The Pap test is indicated to screen for malignant and premalignant lesions of the cervix. The recommended age at initiation of cervical cancer screening has undergone significant revision over time as the natural history of HPV infection and subsequent cervical dysplasia has been elucidated. Although former guidelines recommended starting Pap smear screening at age 18 or the onset of sexual activity, these guidelines were revised in 2006 to recommend initiation 3 years after the onset of sexual activity or age 21, whichever comes first. In 2009, these were further revised to recommend that cervical cancer screening begin at age 21, regardless of sexual history. This recommendation was confirmed in 2012 and again in January 2016.[2]

Abnormal cervical cytology is very common in young women, and most abnormal cytology resolves without treatment in adolescents. In addition, women under the age of 21 account for only 0.1% of all cervical cancers, and no evidence exists that cervical cancer screening in this age group reduces cervical cancer incidence, morbidity, or mortality.

Recognizing these facts and the likelihood of cervical cancer screening leading to unnecessary and potentially harmful evaluation and treatment in women at very low risk for malignancy, the 2009 ACOG guideline revision recommended cervical cancer screening beginning at age 21 years of age, regardless of sexual history, and that recommendation remains unchanged.[2]

Both the U.S. Preventive Services Task Force (USPSTF) and the American Cancer Society (ACS) agreed with this recommendation and issued age-appropriate screening strategies for cytology (Pap tests) and HPV testing for cervical cancer screening in 2012 (see Table).[8, 9]

Table. Summary of 2012 Screening Guidelines from the American Cancer Society, American Society for Colposcopy and Cervical Pathology, and American Society for Clinical Pathology (Open Table in a new window)


ACS Recommendations

Age to start screening

Begin screening with cytology at 21 years old, regardless of sexual history

Screening interval age 21–29

Screen with cytology alone every 3 years.* HPV testing should not be used for screening in this age group.

Screening interval age 30-65

Screen with a combination of cytology and HPV testing every 5 years (preferred) or cytology alone every 3 years. Screening by HPV testing alone is generally not recommended.*

Age to stop screening

Age 65, if the woman has adequate negative prior screening and is not otherwise at high risk for cervical cancer

Screening after hysterectomy

Not indicated for women without a cervix and without a history of a high-grade precancerous lesion (eg, CIN2 or CIN3) in the past 20 years or cervical cancer ever

HPV-vaccinated women

Screen according to the same recommendations as for unvaccinated women

These guidelines do not address special populations (eg, women with a history of cervical cancer, women who were exposed in utero to diethylstilbestrol, women who are immunocompromised) who may require more intensive or alternative screening.

*If abnormal test results are present, further testing and management should be performed according to the ASCCP Guidelines as noted below under Management of Abnormal Cytology.

The ACS and USPSTF 2012 guidelines did not address special populations (eg, women with a history of cervical cancer, women who were exposed in utero to diethylstilbestrol (DES), women who are immunocompromised) who may require more intensive or alternative screening. ACOG recommends annual screenings for immunocompromised patients, women with history of CIN2, CIN3, or cancer; and women who were exposed to DES in utero. According to the latest ACOG recommendations,[2] HIV-positive women should be screened with cervical cytology at the onset of sexual activity or age 21 years, whichever comes first, and should undergo annual screening until they have at least 3 consecutive normal annual cytology results. HIV-infected women age 30 years and older can undergo cytology alone or cotesting. Once there are 3 consecutive negative annual cytology results or one negative cotest, screening can be extended to 3 years. Screening should continue for at least 20 years after treatment of CIN2 or more.[2]

Testing for the high-risk HPV types, often referred to as HPV DNA testing, has become increasingly used in cervical cancer screening and may be performed from any liquid-based cytology specimen. Currently, there are four FDA-approved HPV DNA tests on the market used for reflex testing with ASC-US cytology and for co-testing with the Pap test in women age 30 years or older. Type-specific testing for HPV 16/18, also known as HPV genotyping, is also FDA approved for the same indications; however, it is currently only recommended by the ASCCP for triage of women age 30 years and older who have negative cytology and are HPV DNA positive.  

Note that HPV DNA testing for the purpose of primary cervical cancer screening is currently not yet recommended by major societies in women younger than age 30 years due to the high prevalence of HPV infections in this age group as well as the transient nature of these infections. One HPV test was approved by the FDA in 2014 for primary HPV screening in women age 25 years and older. This test may be used as an alternative to cytology in women in this age group; however, appropriate algorithms should be followed according to the interim guidance by the ASCCP and Society for Gynecologic Oncologists (SGO).[10]  HPV DNA testing is not recommended in women younger than 21 years of age for any reason or following any cytology result (and if performed should be ignored). In addition, testing for low-risk HPV types is never appropriate and should not be performed under any circumstances.

Reviews conducted for the US Preventive Services Task Force support these indications. Vesco et al confirmed the low incidence of cervical cancer among women younger than 20 years and underscored the difficulties of detection and the high frequency of false-positives in this age group.[11] Further, incidence of and mortality rates from cervical cancer in women aged 65 years and older who have had a Pap smear within 3 years have decreased since 2000, and available evidence reinforces discontinuation of cervical cancer screening among these women who have had satisfactory screening and are not otherwise at high risk.

In a systematic review, Whitlock et al found evidence supporting the use of liquid-based or conventional cytology for cervical cancer screening, but cautioned that more evidence is needed before adopting HPV-enhanced primary screening for women aged 30 years and older.[12]   



See the list below:

  • Ideally, cervical screening should be scheduled when the patient is not menstruating.

  • Avoid vaginal intercourse, douching, use of tampons, use of medicinal vaginal cream or contraceptive cream for 24-48 hrs prior to cervical screening.

  • Ideally, pre-existing cervicitis should be treated prior to cervical screening.

  • Screening, however, should proceed in the presence of bleeding or cervicitis, as these symptoms may be related to cervical dysplasia or neoplasm, which may be detected with cervical screening.



The following equipment is necessary to perform the Pap test:

  • Examination table with foot supports

  • Examination light

  • Metal or plastic speculum

  • Examination gloves

  • Cervical spatula and cytobrush

  • Liquid-based cytology container or glass slide and fixative



The patient should be supine, in dorsal lithotomy position to correctly perform a pap smear (see the image below). The coccyx of the patient must be at the edge of the examination table to provide adequate visualization of the cervix once the speculum is inserted.

Positioning and procedure. Positioning and procedure.


A metal or plastic speculum is placed in the vagina to examine the cervix. If necessary, lukewarm water may be used to lubricate and warm the speculum for patient comfort. In situations in which a lubricant must be used, only a small amount should be applied to the outer portion of the speculum, with caution to avoid the tip. According to Hologic Inc, maker of ThinPrep pap test, the following lubricants do not contain substances that interfere with the liquid-based Pap tests: Surgilube, Astroglide, and Crystelle.[13]

To ensure an adequate sample is collected, the surface anatomy of the cervix must be fully visualized, including the squamous epithelium of the ectocervix, squamocolumnar junction, and the external os. The transformation zone of the cervix is the region where squamous epithelium replaces glandular epithelium in a process called squamous metaplasia. Because HPV has a predilection for this region,[14] screening must focus on sampling the cells at the transformation zone to adequately detect the presence of dysplasia. Discharge covering the cervix may be removed carefully using a large swab, ensuring that the cervix is minimally traumatized.

To obtain the specimen, a cervical broom or cervical spatula is applied to the surface of the cervix and turned in a single direction to achieve an adequate sample for cytology, making sure to rotate it at least 360 º for the spatula and 5 rotations for the broom. If the spatula is used, a cytobrush is additionally needed and must be inserted into the cervix so that the outermost bristles are still visible at the external os. The brush is then rotated one half turn in a single direction to achieve an adequate cytology sample (see image below).

The specific protocol for specimen transfer varies depending on the test used. For SurePath, after the cervical broom or cervical spatula and cytobrush are removed from the cervix, they are placed specimen side down into the liquid cytology vial, each removable head is snapped off, and the vial is labeled and sent to pathology. For ThinPrep, the spatula and brush are to be swirled vigorously in the vial 10 times to release the specimen and then discarded. Similarly, if the broom is used, it is to be pushed into the bottom of the vial 10 times and then swirled vigorously and discarded. When conventional cytology is to be performed, the specimens are smeared on a glass slide and subsequently sprayed with fixative or placed in 90% alcohol solution.

Although the FDA-approved protocol for the cervical broom does not require use of the cytobrush, some practitioners use the cytobrush following the broom in an attempt to improve the likelihood of obtaining an endocervical component in the sample. Small studies show no significant difference in acquiring endocervical cells between the broom and spatula plus cytobrush;[15, 16] however, other studies have shown the spatula/cytobrush method to be better at sampling endocervical cells than the broom alone.[17, 18]

In addition, 2 much larger studies found that the broom/cytobrush combination improves sampling of the endocervix compared to the broom alone.[19, 20] Whether these potential sampling differences affect the sensitivity of cervical cytology for detecting moderate or severe dysplasia or cancer is unclear; however, based on the available data, using the cytobrush to obtain endocervical cells in addition to the spatula or the broom is reasonable.

Pap smear. Pap smear.


Complications are extraordinarily rare and include minor bleeding and infection. The patient must be educated on the likelihood of vaginal spotting immediately after a pap smear is performed, as this is considered normal.


Although the Pap smear is one of the best screening tests in medicine and its implementation has decreased the incidence of cervical cancer by over 50%, it does have its limitations. First, the sensitivity of one Pap smear for cervical dysplasia ranges from 30-87%[21] (with the average approximately 58%).[22] In addition, the intraobserver and interobserver reproducibility is poor and ranges from approximately 43-68% at best. Nearly half of all new cervical cancers are found in women who have never had cervical cytology screening prior to diagnosis. Unfortunately, however, false-negative Pap smears are associated with up to 30% of all new cervical cancer diagnoses.

HPV DNA testing has improved sensitivity over cervical cytology but lower specificity. For women age 30 years and older, the sensitivity and specificity of the HPV DNA test for detecting CIN 2 or worse are roughly 95% and 87%, respectively.[23]   For younger women who are more likely to have transient HPV infections, the specificity is much lower.


Interpreting Cytology Results

Results from cervical cytology specimens are reported according to the 2001 Bethesda System Classification, as listed below.[24]

Negative for intraepithelial lesion or malignancy

Epithelial cell abnormality

Squamous cell

  • Atypical squamous cells (ASC) of undetermined significance (ASC-US) or atypical squamous cells that cannot exclude HSIL (ASC-H)

  • Low-grade squamous intraepithelial lesions (LSIL), includes human papillomavirus (HPV), mild dysplasia, and CIN 1

  • High-grade squamous intraepithelial lesions (HSIL), includes moderate to severe dysplasia, carcinoma in situ, CIN 2, and CIN 3

  • Squamous cell carcinoma

Glandular cell

  • Atypical glandular cells (AGC), specify endocervical, endometrial, or not otherwise specified (NOS)

  • Atypical endocervical cells, favor neoplastic, specify endocervical or NOS

  • Endocervical adenocarcinoma in situ (AIS)

  • Adenocarcinoma


See the list below:

  • Endometrial cells in a woman 40 years of age or older


Management of Abnormal Cytology

The following section discusses management of abnormal cytology (2012 ASCCP Consensus Guidelines).[25, 26, 27]

Management of women with ASC-US

HPV testing is preferred; if negative, repeat co-testing in 3 years; if positive, perform colposcopy.

Repeat cytology in 1 year is acceptable; if negative, repeat cytology in 3 years; if ASC or greater, perform colposcopy.

Women aged 21-24 years with ASC-US

  • Repeat cytology in 12 months

  • If repeat cytology is ASC-H, AGC, or HSIL, perform colposcopy; otherwise, repeat cytology in another 12 months

  • If repeat cytology at 24 months is negative, resume routine screening; otherwise, perform colposcopy

  • Alternative is to perform HPV testing; if positive, repeat cytology in 12 and 24 months as above; if negative, resume routine screening.

Women aged 20 years or younger with ASC-US or LSIL

  • HPV infection and minor abnormal cytology results common in adolescents, but invasive cancer is rare

  • Per 2009 ACOG guidelines, Pap tests only recommended beginning at age 21 years, regardless of sexual history; however, Pap tests are still performed in the 20 years or younger age group in some cases because of lack of knowledge of current guidelines

  • Conservative management preferred for this group because of the high likelihood of spontaneous resolution within 2 years of initial infection; abnormal cervical cytology in adolescents, therefore, should be followed according to the recommendations for women ages 21-24 years, as described above

Pregnant women with ASC-US

  • Managed same as nonpregnant women

  • Endocervical curettage (ECC) is contraindicated in pregnant women and should not be collected if colposcopy is performed

  • Deferring colposcopy until at least 6 weeks postpartum is also acceptable

Management of women with ASC-H

Perform colposcopy (regardless of HPV status).

Management of women with LSIL

Women aged 25 years or greater with LSIL

  • Perform colposcopy

  • If HPV co-testing was performed and negative, repeat co-testing in 1 year is preferred

Women aged 21-24 years with LSIL

  • Repeat cytology in 12 and 24 months; follow guidelines for ASC-US

Pregnant women with LSIL

  • Managed same as nonpregnant women

  • ECC is contraindicated in pregnant women and should not be collected if colposcopy is performed

  • Also acceptable to defer colposcopy until at least 6 weeks postpartum

Postmenopausal women with LSIL

  • Acceptable options include reflex HPV testing, repeat Pap at 6 and 12 months, and colposcopy

  • If HPV negative or no CIN on colposcopy, repeat cytology in 12 months; if HPV positive or repeat cytology is ASC or greater, perform colposcopy

  • May return to routine screening if 2 consecutive negative cytology results

Management of women with HSIL

Refer to colposcopy regardless of age.

Immediate loop electrosurgical excision is acceptable, except in patients younger than 25 years or who are pregnant.

Pregnant women with HSIL

  • Managed same as nonpregnant women

  • ECC and immediate loop electrosurgical excision are contraindicated in pregnant women and should not be performed

Management of women with AGC

Women with AGC, including ASC-NOS, AGC-favor neoplasia, and AIS

  • Refer to colposcopy with endocervical sampling

  • If age 35 or greater or with other risk factors for endometrial neoplasia, endometrial sampling should also be performed.

Women with atypical endometrial cells

  • Perform endometrial biopsy and endocervical sampling. If no pathology found, proceed with colposcopy.

Management of women with benign endometrial cells found in cervical cytology

No additional evaluation is required in asymptomatic premenopausal women.

In postmenopausal women, perform endometrial biopsy.

Management of women age 30 years and older who are Pap negative and HPV positive

Repeat cytology and HPV DNA testing in 12 months

  • If cytology negative, HPV negative, repeat co-testing in 3 years

  • If cytology abnormal with any HPV result, perform colposcopy

  • If cytology negative, HPV positive, perform colposcopy

Another option would be to perform HPV 16 and 18 testing

  • If 16 or 18 positive, perform colposcopy

  • If 16 and 18 negative, repeat co-testing in 12 months

  • If cytology negative, HPV negative, repeat co-testing in 3 years

  • If cytology abnormal with any HPV result, perform colposcopy

  • If cytology negative, HPV positive, perform colposcopy


Primary High-Risk HPV Screening

In 2014, the FDA approved one of the HPV DNA tests (cobasHPV test) for primary screening for cervical cancer in women aged 25 years and older. Although not yet fully endorsed by major societies, a panel of experts co-sponsored by the ASCCP and the SGO published recommendations for using primary HPV testing for cervical cancer screening.[10]  In the proposed algorithm, women aged 25 years and older would undergo cervical HPV testing for screening, as opposed to cytology. Women who test negative for high-risk HPV should be re-screened no sooner than every 3 years. For women who test positive for high-risk HPV, HPV genotyping is performed, and those who test positive for HPV 16/18 are referred for coloposcopy. Those who are high-risk HPV positive but 16/18 negative undergo cervical cytology. Those with negative cytology results are rescreened in 1 year, and those with ASC-US cytology or greater are referred for colposcopy.  

The above algorithm appears to be at least as effective as current cytology-based screening guidelines and therefore may be considered as a reasonable alternative. Nonetheless, cytology alone and cotesting, as outlined above, remain the screening modalities recommended by guidelines of major societies.