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Cervical cytological specimen


Medical editor Gunnar B. Kristensen MD
Gynecological Oncologist
Oslo University Hospital

Maj Liv Eide
Bioengineering
St. Olavs Hospital HF

General

Screening of cervical cancer is unique in that it can prevent further development by treating pre-invasive stages. Well-organized screening programs can reduce both the incidence and mortality of cervical cancer. In Norway the incidence of cervical cancer is reduced by 25 % and mortality by 50 % since the national screening program was introduced in 1995. Cervical cancer screening has led to a decrease in incidence of cervical cancer by 60–70 %. A cervical smear is taken every three years on women between 25-69 years in the Norwegian Cevical Cancer Screening Program.

The sensitivity of a cervical smear is relatively low (55–80 %), but the specificity is high. This means that a negative test will not exclude pre-stages of cancer or invasive cancer. Cytology testing should never be the only diagnostic tool in patients with symptoms. With a positive cytological diagnosis, a histological test will show the equivalent or a more serious diagnosis in 80–90 % of cases.

A cervical sample can be taken as a conventional smear or as fluid-based cytology. Fluid-based cytology is now the preferred method. The method allows HPV testing on the same sample and enables HPV testing as a primary screening method in the future.

The quality of the sample depends more on the technique of the practitioner than on the devices used for collection. It is important to read the instructions carefully.  Incorrect specimen collection is a frequent cause of false negative tests.

Indication

  • Screening of healthy women without symptoms as part of the national screening program. This method should not be used as a diagnostic tool for cervical cancer.

Goal

  • To obtain an adequate sample for diagnosing pre-stages of cervical cancer. 

Factors resulting in poor testing:

  • Inflammation
  • Pregnancy
  • Postmenopausal women with atrophic mucosa

Postpone the test:

  • If the patient is menstruating.
  • If the patient has used vaginal cream or a diaphragm in the last 24 hours.

Equipment

  • Examination table
  • Adjustable lamp with adequate illumination
  • Water or water-soluble lubrication
  • At least 3 types of specula: Self-retaining specula are recommended. All specula should be autoclaved at least 15 minutes at 121°C if not diposible. Chemical disinfection is not sufficient.  
  • Gloves
  • Either: cervical brush and spatula 
  • Or: combination brush (for example Cervex-Brush®) 
  • Fluid- based cytology:
    • A specially designed vial containing a preservative for fluid-based cytology (ThinPrep®Pap Test eller BD SurePath™Pap Test is currently used in Norway). Which vial used depends on the method chosen by the cytology laboratory.
  • Conventional smear:
    • Slide glass with frosted end
    • Pencil for labelling the sample
    • Fixation spray, containing alcohol
    • Holder for transport of slides
  • Referrals

Preparation

  • Ask the patient about her general health and possible symptoms of discharge or abnormal bleeding.
  • Explain the procedure to the patient. The examination is rarely painful. Taking the sample takes 5-10 minutes. The result is available after 2-3 weeks. Calm the patient. Remember that many women associate abnormal test results with invasive cancer. Inform the patient when and how the test result will be communicated to the patient. Inform the patient about what the next step will be if the result is positive.
  • The patient should empty her bladder before the test.
  • There should be space/room for the patient to undress.

The referral should include:

As a minimum, The Quality Assurance Manual Cervical Cancer Screening Programme in Norway require information about:

  • Patient’s name and ID number.
  • Appearance of the cervix.
  • First day of last menstruation.
  • Birth control method, if applicable. 
  • It should be stated if the sample is follow-up after previous findings (cytology, HPV and/or histology), preferably with statement on relevant diagnosis.

Implementation

  • Help the patient to be as comfortable as possible on the examination table.
  • Make sure there is adequate lighting.
  • Wash your hands and put gloves on.

Insert the speculum

  • Choose the largest speculum the patient is comfortable with.
  • The speculum should have a comfortable temperature for the patient.
  • Use water or water-soluble lubrication on the speculum, but only if necessary. Avoid spreading lubrication to the tip of the speculum, which can be transferred to the cervix and hence the sample.
  • Insert the speculum in the direction of the patient's tailbone.
  • Do not open the speculum until it is in place.

Localization of the cervix

  • Adjust the light.
  • Open the speculum slowly and carefully. The cervix will then become visible.
  • In overweight patients or women with flaccid vaginal walls, it may help to cut the thumb off of a large, sterile glove and put it over the tip of the speculum to prevent the vaginal wall from falling inward and blocking the cervix.
  • Use a long speculum on tall women.
  • If there is abundant mucus on the mouth of the uterus, wipe it carefully off before sampling
  • When it is difficult to localize the cervix, it is possible to search manually and/or ask the patient to change positions.

Fluid-based cytology

Use either a cervical brush and spatula or a combination brush. How to transfer the sample to the vial with a combination brush, depends on which fluid-based method is used.

Spatula and endocervical brush (ThinPrep®Pap Test)

  • Sampling with the cervical brush may cause bleeding and should be taken after the spatula sampling.
  • Ectocervical cytological sampling: Insert the contoured end of the plastic spatula into the mouth of the cervix and rotate 360 grades. Keep a steady pressure and make good contact with the surface. Rinse the brush as quickly as possible into the vial by rotating vigorously.
  • Endocervical cytological sampling: Insert the endocervical brush into the cervical canal and rotate carefully 180-360 grades around. Rinse the brush as quickly as possible into the vial by rotating vigorously.

Combination brush for ectocervical and endocervical cytologic sampling (ThinPrep®Pap Test or BD SurePath™Pap Test)

  • Perform sampling from the ecto- and endocervix by inserting the brush into the mouth of the cervix until the short bristles make contact with the ectocervix.
  • Use light pressure and slowly rotate the brush in one direction, simultaneously. There are different kinds of brushes. Be aware of that some brushes should be inserted into the endocervical canal and rotated five times (alternative 1) or rotated twice (alternative 2).
  • ThinPrep®Pap Test: Rinse the brush quickly into the solution vial by pushing the brush into the bottom of the vial 10 times. Swirl the brush vigorously before disposal.
  • BD SurePath™Pap Test: Snap off the device handle and drop the head of the device into the fixative solution vial
  • Tighten the cap. 
  • Record the patient’s name, ID number and date of sampling on the vial.
  • Keep the vial in room temperature before sending. Send the vial to the laboratory as soon as possible.
  • This also applies to HPV testing on the same sample. 
  • When using ThinPrep®Pap Test, never let any of the brushes remain in the fixation solution before swirling. This will cause that the sample material will be fixated to the brush and provide cell-poor material.

Conventional cytological smear

  • Label the slide with the patient name and ID number on the frosten end, using a pencil.
  • Never wash/disinfect the cervix before specimen collection, except when there is abundant mucus on the mouth of the uterus. Wipe it carefully off before sampling.
  • Insert the tip of the spatula up to the opening of the cervix with the inner rounded part of the tip close to the surface.
  • Always rotate the spatula more than 360° and simultaneously press firmly but carefully to retrieve surface cells from the entire cervical opening.
  • The amount of pressure is determined by the texture of the spatula and experience and routine of the practitioner. If uncertain, increase pressure.
  • The procedure should be repeated if there is uncertainty whether the entire area has been included or if sparse material is collected.
  • If the tip of the spatula cannot be placed in the opening of the cervix, the back side of the rounded tip can be used to take an equivalent sample, or invert the spatula and use the other side.
  • Always take a sample of the cervix with a cervical brush in addition to the spatula sample. It is important to take a sample from the junction between the endo and ectocervix.
  • The cervical brush is placed in the cervical opening and is inserted almost all the way to the end of the opening so that the lower end of the brush is visible and is rotated 180–360° before it is removed.
  • The cervical brush specimen should always be taken after the spatula specimen since it often causes bleeding.
  • If the brush is inserted all the way to the end, there is risk of mixing with endometrial cells, which may interfere with the result.
  • A cervical brush alone should never be used for specimen collection.
  • The patient may have light bleeding after the specimen is taken.

Specimen preparation

When the spatula specimen has been taken, it can be placed to the side while the brush specimen is collected. When the brush specimen is taken, both specimens can be transferred to the slide. Transfer all material from the spatula and brush to each half of the slide. All the material should be evenly distributed, however, the frosted end should not be covered.

  • Hold the spatula parallel to the surface on the slide and transfer the cells with a vertical sweep to the part of the slide closest to the frosted end.
  • Spread the cells evenly and not too thick.
  • All specimen material from both sides of the spatula should be transferred.
  • The brush should be rolled carefully over the lower third of the slide.
  • Fix the smear immediately with fixation spray containing alcohol. Fixation should be done before the smear dries. Remember that cells dry more quickly when smeared onto a slide.
  • Be extra quick and careful if the specimen contains blood or is from a postmenopausal patient. This material dries especially quickly.
  • If the cytological specimen collection dries before fixation, this will complicate the cytological assessment and increase the risk of false negative tests.

Removing the speculum

  • Do not close the speculum before there is an adequate distance from the cervix and ensure that no mucosa will be pinched, as this will be painful to the patient.

Concluding the procedure

  • Help the patient down from the examination table.
  • Inform the patient of possible bleeding. Light bleeding immediately after specimen collection may occur.
  • Offer the patient a sanitary pad.
  • Make sure the patient understands when the result can be expected.
  • There should agreement on how the patient is handled further depending on the results.
  • Document any clinical findings on the referral, as well as whether the cervix was difficult to inspect and why it was necessary to collect two specimens or use a brush.


Follow-up

  • Normal cytology specimen: The woman returns to three-yearly screening intervals. The physician (doctor) who has taken the sample, informs the patient.
  • Unsuitable specimen: Control within 1-3 months. The physician (doctor), who has taken the sample informs the woman. The Norwegian Cervical Cancer Screening Programme sends a reminder to the women, if no new sample is registered. If there are repeated unsuitable samples, the woman should be referred to a gynecologist.
  • ASC-US and LSIL on aconventional smear without HPV testing: A new cytology specimen and HPV testing (triage) are recommended after minimum 6 and maximum 12 months. The doctor informs the woman. The Norwegian Cervical Cancer Screening sends a letter to the woman if no repeat test is registered.
  • ASC-US and LSIL and a negative HPV test: The woman returns to three-yearly screening intervals. The physician (doctor) who has taken the sample, informs the patient.
  • ASC-US and LSIL and a positive HPV test: A new cytology specimen and a HPV test after 6-12 months. The doctor summons. The Norwegian Cervical Cancer Screening Programme sends a letter to the woman if no repeat test is registered.
  • ASC-H, HSIL, AGUS, AIS/ACIS and cervical cancer: The woman should be referred for colposcopy and biopsy directly. The doctor informs, summons and refers. The Norwegian Cervical Cancer Screening sends a letter to the laboratory and possibly to the doctor who took the test, if no repeat test is registered. 

When cytological samples are taken to diagnose women with suspicious symptoms such as persistent vaginal discharge, abnormal bleeding and pain: It is important to remember that normal cytology does not rule out cancer, especially in women with symptoms after menopause. If symptoms occur before the next screening test during the three-year interval, a new sample is recommended together with colposcopy examination.


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