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Cytology for technicians (Proceedings)

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Aug 01, 2009

In the last ten years, exfoliative cytology has been gaining popularity as a diagnostic aid in veterinary practice. Effective use of diagnostic cytology depends upon proper specimen preparation; however, there are far more descriptions of slide interpretation than there are of slide preparation. In this manuscript we therefore emphasize successful techniques for collection of cytologic specimens from a variety of sources. A discussion of smear preparation and staining techniques is also included.

In vivo collection techniques

Cytologic preparations may be made from lesions present in the living animal (in vivo collections) or from masses or lesions which have been excised (in vitro collections). In vivo collections offer several advantages over surgical removal (e.g., significantly lower cost, no general anesthetic risk, etc.) and is recommended whenever feasible. Several different in vivo collection techniques will be described: the majority are variations of fine needle aspiration, aspiration through catheters, or swabbing of superficial lesions.

Fine needle aspiration of superficial masses and lesions, lymph nodes, and internal masses

Fine needle aspiration can be used to collect cytologic specimens from virtually any lesion internal or superficial which can be identified and stabilized during the collection procedure. The only equipment required is a small syringe (5-10cc) and a 1.5-inch fine gauge (20-25) needle. The site to be aspirated is clipped, washed and prepared for minor surgery. Sterility of the needle is maintained, but surgical gloves and surgical drapes are not necessary. General anesthesia or tranquilization is required only in fractious animals.

The needle is attached to the syringe and introduced into the lesion to be aspirated. As negative pressure is applied to the syringe, the needle is redirected through the mass several times. Moving the needle through different areas of the lesion before the negative pressure is removed assures that the entire lesion area is sampled. Negative pressure is then released and after release of the negative pressure the needle is withdrawn from the lesion.

If the sample has been properly collected (assuming the lesion is solid rather than fluid-filled), the majority of the sample will be contained in the hub of the needle. The needle is separated from the syringe, and the syringe is filled with air and re-attached to the needle. The material in the needle is forced onto a series of microscope slides and finished preparations are made by placing a clear slide across the aspirated material, allowing the material to spread between the two slides, and then gently drawing the top slide across the surface of the bottom slide. The finished "squash preparations" are allowed to air dry and are then stained with a standard hematologic stain (Wright's, Wright's-Giemsa, Diff-Quick, etc.).

Thoracocentesis and abdominal paracentesis

Thoracocentesis and abdominal paracentesis are modifications of basic fine needle aspiration technique. For abdominal paracentesis an eighteen-gauge 1.5-inch needle and 10-20cc syringes are used; similar equipment is used for thoracocentesis and a three-way value is added.

Thoracocentesis may be done on either the left or right side, generally in the 68th rib space. Local anesthesia is recommended with this procedure so that the animal does not move when the thoracic cavity is entered. Movement pre-disposes to lung laceration. The three-way value is employed so that all of the fluid in the chest may be removed in order to provide therapeutic relief of dyspnea. At least two tubes of fluid are saved: one sterile tube for microbiologic evaluation and an EDTA tube for cytologic evaluation. The EDTA sample is used for the evaluation of physical parameters (total cell counts, total protein, specific gravity) and cytologic preparations. Both direct smears and sediment smears may be prepared. Direct smears are made by taking a drop of fluid; placing it on a microscope slide and making a push smear as is done with peripheral blood. Sediment smears are made by centrifuging the sample to obtain a sediment, decanting the supernatant, and preparing a push smear from a drop of sediment. The prepared smears are allowed to air dry and are then stained with standard hematologic stains.

Abdominal paracentesis is performed with the animal standing. The needle is introduced just anterior and to the right of the umbilicus. Neither local nor general anesthesia is generally required. A three-way value is not needed for abdominal paracentesis; there is no therapeutic benefit to removing all of the fluid from the abdominal cavity. The fluid collected by abdominal paracentesis is handled in the same manner as fluid obtained by thoracocentesis.

A key to successful fluid collection from the body cavities is careful positioning of the bevel of the needle as the cavity is entered and aspiration begun. The bevel of the needle should always be directed outward towards the body wall. This prevents plugging of the needle opening by either omental fat or lung. If initial aspiration yields a dry tap, negative pressure is released, the syringe is rotated a full 360° and aspiration is again attempted.