HPV SPECIMEN COLLECTION AND HANDLING:

A. Specimen collection and processing

Collect cervical specimens in ThinPrep Pap Test vials containing PreservCyt Solution with broom-type or cytobrush/spatula collection devices according to the manufacturer’s instructions listed below:

B. PREPARATION FOR COLLECTION OF THINPREP SPECIMENS:

1.     Preferably a woman should be tested two weeks after the first day of her last menstrual period (LMP) and definitely not when she is menstruating. A woman should not use vaginal contraceptives or douches during the 48 hours before exam, and the woman should abstain from intercourse 24 hours prior to the exam.

2.     Prior to obtaining the sample, write the patient’s first and last name on the PreservCyt Solution vial along with a second identifier such as patient’s date of birth, location or chart number.  Proper patient identification is mandatory.

3.     For proper evaluation of the specimen, the demographic information and patient history section of the requisition form should be completed in its entirety.

C. SAMPLING:

1.     PANCERVICAL-ENDOCERVICAL BRUSH SMEAR TECHNIQUE:

a. Insert a plastic spatula into the cervical canal and rotate 360 degrees about the circumference of the os and ectocervix while maintaining firm contact with the epithelial surface. 

b. Rinse the spatula as quickly as possible into the PreservCyt Solution vial by swirling the spatula vigorously in the vial 10 times.  Discard the spatula.

c. Insert the endocervical brush device into the cervix until only the bottom-most fibers are exposed.  Slowly rotate ¼ or ½ turn in one direction.  Do not over-rotate.

d. Rinse the brush as quickly as possible in the PreservCyt Solution by rotating the device in the solution 10 times while pushing against the PreservCyt vial wall.  Swirl the brush vigorously to further release material.  Discard the brush.

e. Tighten the cap so that the torque line on the cap passes the torque line on the vial.

        2.   BROOM-LIKE BRUSH TECHNIQUE:

a. Another collection instrument is a “broom-like” brush that simultaneously samples the endocervix and ectocervix.  Insert the central bristles of the broom into the endocervical canal deep enough to allow the shorter bristles to fully contact the ectocervix.  Push gently, and rotate the broom in a clockwise direction 5 times.

b. Rinse the broom as quickly as possible into the PreservCyt Solution vial by pushing the broom into the bottom of the vial 10 times, forcing the bristles apart.  Finally, swirl the broom vigorously to further release material.  Discard the collection device.   

c. Tighten the cap so that the torque line on the cap passes the torque line on the vial. 

Procedure notes:

1. The best sampling results are obtained when two types of samples are taken (ectocervix and endocervix).  The pancervical-endocervical brush is the preferred method of sampling.  Collection of samples with a cotton swab is not recommended.

2. Water may be used to lubricate the speculum; however, lubricants should never be used.  It is critical to obtain a specimen that is not obscured by blood, mucus, or inflammatory exudate.  Excess mucus or other discharge should be gently removed with a ring forceps holding a folded gauze pad.  Inflammatory exudate may be removed by placing a dry 2x2 inch piece of gauze over the cervix and peeling it away after it absorbs the exudate.  The cervix should not be cleaned by washing with saline and the sample should always be obtained before the application of acetic acid.

3. Cytological specimens should be considered infectious until fixed with alcohol based fixative.  Observe universal precautions when handling specimens from all patients.

4. For ThinPrep specimens, the collection device must be rinsed in the PreservCyt Solution as quickly as possible.

There must be at least 1.0mLs of PreservCyt Solution remaining for the APTIMA HPV Test to be performed.   Samples with less than 1.0mLs, after the Pap Test has been prepared, may contain insufficient material and could produce a falsely negative APTIMA HPV result.

D.  SUBMISSION:

Transport and storage prior to testing

1. Transport the ThinPrep liquid cytology specimens by mail or courier at 4°C to 30°C.   Place the PreservCyt Solution vial into a biohazard bag.  Zip the bag to ensure proper closure.  Insert the completed request form in the outside pocket of the biohazard bag. 

2.    Specimens should be transferred to an APTIMA Specimen Transfer tube within 21 days of collection. This transfer is performed by the cytology department.

3.    Prior to transfer, ThinPrep liquid cytology specimens should be stored at 4°C to 30°C, with no more than 30 days at temperatures above 8°C.

4.    ThinPrep liquid cytology specimens transferred to an APTIMA Specimen Transfer tube may be stored at 4°C to 30°C for up to 60 days.

5.    If longer storage is needed, the ThinPrep liquid cytology specimen or the ThinPrep liquid cytology specimen diluted into the APTIMA Specimen Transfer tube may be stored at -20°C to -70°C for up to 24 months.

Note: Specimens must be shipped in accordance with applicable local, national and international transportation regulations.

Specimens should be shipped along with a requisition slip to the lab.  Specimens may be shipped without refrigeration to a testing laboratory.   Protect specimens from extreme temperatures.

HPV TESTING OPTIONS

HPV REFLEX TESTING AND ADD-ONS:

When the patient Gyn requisition form is being completed, the clinician may select the option to order reflex HPV testing if the Pap Test result is ASCUS.  An HPV option (High Risk HPV type) must be selected and marked on the Gyn requisition form for the testing to be performed.  If a clinician wishes to order the HPV test as an add-on, it may be added by calling Pathology Consultants, P.C. at 701-222-2480 (1-800-659-0395) within 21 days of sample collection.

HPV TESTING WITH THINPREP PAP (DNA WITH PAP):

A clinician may order HPV testing with the ThinPrep regardless of the Pap Test result, referred to as cotesting.  In 2003, the FDA approved the use of the HPV DNA testing in women 30 and older for primary cervical cancer screening.  Expanded testing or cotesting (consisting of both Pap and HPV DNA tests) provides more targeted screening by identifying women who have an increased risk of cervical cancer.  Women with HPV in their cervical cells require more diligent follow-up as long as the virus persists—even when their Pap Test result is negative.  Due to low sensitivity on any single test, the Pap requires frequent retesting.  The combination of a Pap and a HPV test is more sensitive than a Pap alone and identifies women who possibly have long-term persistent infection or disease missed by the Pap.  Women who test negative on both the HPV and Pap Test have virtually no risk of cervical cancer over the next few years and may safely widen their screening interval.  The frequency of testing can be individualized based on a women’s screening history, age at first intercourse, HPV status (if known), age and perception of risk.

HPV Testing with a Conventional Pap:

A sample may be submitted for HPV testing when doing a Conventional Pap by collecting and submitting a ThinPrep sample in addition to the Conventional Pap smear.

HPV TESTING ONLY:

HPV Testing Only may be requested without a Pap Test if you want to know the patient’s HPV status.  You must procure a cervical/vaginal ThinPrep sample.  For proper evaluation of the specimen, the demographic information and patient history section of the GYN requisition form should be completed in its entirety.

REFLEX TO HPV 16, 18/45 IF HPV SCREEN POSITIVE

Testing for HPV 16, 18/45 may be requested by checking the appropriate box that indicates to perform HPV 16, 18/45 if HPV screen is positive.   The Aptima HPV 16, 18/45 genotype assay can differentiate HPV 16 from HPV 18 /45, but does not differentiate between HPV 18 and HPV 45. In women infected with HPV types 16 and/or 18/45, the cumulative risk of developing cervical disease is 10-fold higher compared to the risk for disease development due to other high-risk types.

Example of GYN Cytology Request:

WOMEN 20 YEARS AND YOUNGER: □ THINPREP WITH REFLEX HPV TESTING IF ASC-US    □ PERFORM HPV 16, 18/45 IF HPV SCREEN POSITIVE □ COTESTING THINPREP AND HPV TESTING    □ PERFORM HPV 16, 18/45 IF HPV SCREEN POSITIVE □ THINPREP ONLY	WOMEN 30 YEARS AND OLDER: □ COTESTING THINPREP AND HPV TESTING (PREFERRED PER ASCCP GUIDELINES)     □ PERFORM HPV 16, 18/45 IF HPV SCREEN POSITIVE □ THINPREP WITH REFLEX HIGH-RISK HPV TESTING IF ASC-US (ACCEPTABLE PER       ASCCP GUIDELINES)     □ PERFORM HPV 16, 18/45 IF HPV SCREEN POSITIVE □ THINPREP ONLY
WOMEN 21-29 YEARS: □ THINPREP WITH REFLEX HPV TESTING IF ASC-US         (RECOMMENDED PER ASCCP GUIDELINES)    □ PERFORM HPV 16, 18/45 IF HPV SCREEN POSITIVE □ COTESTING THINPREP AND HPV TESTING    □ PERFORM HPV 16, 18/45 IF HPV SCREEN POSITIVE □ THINPREP ONLY	OTHER TESTS: □ HPV TESTING WITHOUT THINPREP    □ PERFORM HPV 16, 18/45 IF HPV SCREEN POSITIVE □ CHLAMYDIA TRACHOMATIS / NEISSERIA GONORRHEA TESTING       (PERFORMED ON THINPREP VIAL) □ CONVENTIONAL PAP NUMBER OF SLIDES 1 2 3 4     □ CONVENTIONAL PAP WITH HPV TESTING (REQUIRES THINPREP VIAL)        □ PERFORM HPV 16, 18/45 IF HPV SCREEN POSITIVE