Quick Reference

SurePath Thin-Layer Pap Preps

1. Perform routine GYN exam. Insert Broom-type device (for example Cervex-Brush™ into the cervical os or vaginal cuff
2. Rotate the Broom/Brush 5 times in a clock-wise direction or counter clock-wise direction NOTE: Either direction may be used, but direction should be consistent. If begin in a clock-wise direction continue all 5 times in same direction, DO NOT alter or vary the direction of the Broom during sampling. NOTE:Vaginal specimens may be collected by using the Broom device in a sweeping motion to obtain appropriate cellular material
3. REMOVE the Broom tip from the handle by gently popping tip off handle and drop tip into the SurePath™ preservative fluid (TIP MUST BE SUBMITTED IN VIAL)
4. Cap vial
5. Send to Cytopathology Laboratory.

Thin-Pap Preps

Broom-like collection device collection:

Do not recommend using lubricant as it interferes with the processing of the specimen and may result in an inadequate slide.

1. Label ThinPrep® PreservCyt vial with patient name and MRN.
2. Perform routine GYN exam. Insert collection device into the cervical os or vaginal cuff.
3. Insert central bristles into the endocervical canal deep enough to allow the shorter bristles to contact the ectocervix.
4. Push gently and rotate CLOCKWISE five times.
5. Rinse the broom immediately in the PreservCyt vial by pushing the broom into the bottom of the vial 10 times, forcing bristles apart.
6. Swirl brush vigorously in solution.
7. Discard brush. DO NOT submit brush in vial.
8. Tighten cap on vial until torque line on cap passes torque line on vial.
9. Transport laboratory for processing.

Endocervical Brush/Spatula Collection

Do not recommend using lubricant as it interferes with the processing of the specimen and may result in an inadequate slide.

1. Label ThinPrep® PreservCyt vial with patient name and MRN.
2. Obtain a sample from the ectocervix using a plastic spatula.
3. Rinse the spatula into the PreservCyt vial by swirling vigorously in the vial 10 times. Discard the spatula.
4. Obtain a sample from the endocervix using an endocervical brush. Insert brush in to the cervix until only the bottom fibers are exposed. Slowly rotate 1/4 to 1/2 turn in one direction. DO NOT OVER ROTATE.
5. Rinse the brush quickly in the PreservCyt vial by rotating the device in the solution 10 times while pushing against the vial wall.
6. Swirl vigorously to further release material.
7. Discard brush. DO NOT submit in vial.
8. Tighten cap on vial until torque line on cap passes torque line on vial.
9. Transport to laboratory for processing.

Conventional Pap Smears

1. Write the patient name and medical record number on frosted end of slide with pencil.
2. Sampling site material (cervical/endocervical) is spread thinly on slide and immediately preserved with a spray fixative.
3. Material for hormonal assessment must be taken from the lateral vaginal wall and smear on pre-labeled separate slide.
4. Ectocervical material is usually obtained by spatula and endocervical material by brushing technique.
5. Fill out requisition form completely.
6. Smear diagnosis report will be according to the Bethesda system.

Collection Method for Sputum Cytology

(Physician office, clinic, or patient floor collection)

1. Saccomanno fixative (98%, 50% ethyl alcohol, 2% polyethylene glycol, tinted green) which is available in the Cytology Laboratory must be added to the cytology specimen container before the collection begins. Usually 20-30 cc fixative (or enough to thoroughly surround specimen) is sufficient. As much as 50 cc of fixative may be used for copious specimens. Do not overfill container.
2. Immerse sputum specimen into fixative container.
3. Send labeled specimen to cytology laboratory with appropriate, complete requisition form.

Collection Method for Cytological Examination of Pleural, Pericardial, and Ascitic Fluids

1. Fluid is collected by the usual methods for thoracentesis or paracentesis.
2. When possible, 100 mL or more of fluid should be collected.
3. An anticoagulant should be added to the container to prevent clotting.
4. Cell blocks are routinely performed on all pleural, pericardial, and ascitic fluids if specimen amount and contents permit.
5. Promptly deliver specimen to the Cytopathology Laboratory with the appropriate request slip.
6. After 5:00 p.m. weekdays and on weekends, the specimen must be placed in the refrigerator in the Cytopathology Laboratory.

Procedure for GI Tract Brushing for Cytological Examination

1. Label slides on frosted end in pencil with patient name and medical record number.
2. Prepare thin smears from GI tract brushing material and immediately spray with alcohol fixative.
3. Complete cytology request form with appropriate information, e.g., patient name, medical record number, physician, source of specimen, and any appropriate clinical history.

FNA Procedure

1. Label an appropriate number of slides with patient name and medical record number with pencil on frosted end.
2. Aspirated material is spread thinly on slides. Allow half the slides to air dry. Fix the other half with aerosol fixative or immerse in 95% ETOH.
3. Any remaining aspirate may be placed in a 50 mL leak-proof plastic tube that contains Saccomanno fixative or RPMI (if cell block is desired).
4. If a lymphoma is suspected, some remaining aspirated material should be placed in a 15 mL plastic centrifuge tube containing RPMI, which may be used for flow cytometry.
5. Fill out requisition completely.
6. Please refer to NCCLS FNAB Technique Guideline (follows) for more in-depth discussion from patient preparation through aspiration.

Procedure for Bronchial Brushings and Washings for Cytological Examination

Bronchial Brushings

1. Prepare thin smear from bronchial brushing and immediately place slide in Coplin jar containing either 95% ethyl alcohol, Saccomanno fixative (98%, 50% ethyl alcohol, 2% polyethylene glycol; tinted green), or spray fixative.
2. Identify Coplin jar with label containing patient's name and medical record number. Identify slides with patient name and medical record number on frosted end.
3. Complete cytology request slip specifying location (side). Send request slip with each container to Cytopathology Laboratory.

Bronchial Washings

1. Place one (1) ounce of Saccomanno fixative (98%, 50% ethyl alcohol, 2% polyethylene glycol, tinted green) in a plastic sputum container.
2. Immerse the bronchial washings or secretions into fixative container.
3. Individual containers should be used for right and left bronchial washings or secretions.
4. Identify each container with patient's name and hospital number.
5. Identify each container as to the left or right side.
6. Complete cytology request slip for each container.
7. Bring specimen request form and container to Cytopathology Laboratory.

Bronchial-Aveolar Lavages (BAL)

1. Specimens for routine cytology, iron stain, cell count, and differential, and/or Hemosiderin-laden macrophages, follow steps 1-7 for Bronchial Washings, making sure to identify specimen on container and requisition as BAL.
2. Specimens for lipid-laden macrophages(Oil Red O) must NOT be put in fixative (this would destroy the lipid). Submit lavage specimen FRESH in sputum container, making sure to identify specimen as a BAL on container and requisition, and indicate request to test for lipid-laden macrophages on requisition. Proceed with steps 3-7 under Bronchial Washings.

Use of Hairspray for Fixation of Cytologic Smears

1. Patient's name and medical record number must be written on frosted end of slide in pencil (not pen or china marker pencil).
2. Smear is obtained by the physician in the usual manner.
3. Immediately spray entire slide with a thin film of fixative by holding can approximately six inches from smear. This must be done before smear begins to dry.
4. The protective film will dry on the slide in about five minutes.
5. Place smears in slide holders provided by the Cytopathology Laboratory.
6. Transport smears in slide holders with usual request forms to Cytopathology Laboratory.

CAUTION: Do not store fixative spray can in heated areas above 120°F.

Collection Method of Cytological Examination of Urines and Bladder Washings

The Cytology Lab requires prompt fixation of urine specimens which are submitted for cytologic examination. This includes urine specimens which are to be examined for tumor cells or for inclusion bodies. Fixation can be accomplished by adding an equal volume of Saccomanno fixative* to the specimen immediately at the time of collection. The specimen may be submitted in an appropriately labeled paraffin collection cup or 150 mL plastic collection cup. Failure to fix the specimen promptly results in the degeneration of cells which may render the specimen useless.

Unlike routine types of urine specimens, the first morning urine is not ideal for cytologic examination because of degenerative changes which take place in cells during storage of the urine in the bladder for a prolonged time. For the same reason, 24-hour urine specimens are not appropriate for cytologic examination.

* Saccomanno Fixative = 95% , 50% ethyl alcohol; 2% polyethylene glycol

Tzanck Smear Procedure

The Tzanck smear has traditionally been used as a means of identifying Herpes infection. To best accomplish this, we require Tzanck slides to be prepared and submitted as follows:

1. With wooden spatula, gauze, etc., remove dry or crusted surface from vesicle to be sampled, revealing more viable skin surface beneath.
2. Scrape vesicle vigorously with wooden spatula and promptly smear all visible material onto the clear portion of a glass slide (which at the frosted end has been labeled with the patient's name and medical record number).
3. Allow slide to air dry completely before enclosing in a slide folder.
4. Complete a Cytology requisition form, filling in all pertinent information. Indicate under "Specimen Source" that the slide is air dried and that the specimen is a "Tzanck Smear for Herpes." Also, identify the site on the body from which the smear was made.
5. Rubber band requisition around slide folder containing specimen and transport to Cytopathology Lab.
6. Notify Cytology personnel when leaving specimen that the specimen is a Tzanck smear to facilitate prompt processing.
7. The slides will be stained with Diff-Quik stain.

*If you have any questions, please call Client Services at 1 (800) 695-6491.

Adopted MKurtinecz, 12/04
Reviewed MKurtinecz, TSp 12/04
Reviewed MKurtinecz, TSp 1/06
Reviewed MKurtinecz,TSp 5/07
Reviewed: MKurtinecz, TSP 7/08
Reviewed: MKurtinecz, TSp 8/09
Supersedes: CytoCollFeb2001 Manual