Wet mount

The diagnosis of scabies is made by obtaining skin scrapings and detecting the mite in a wet mount. Topical therapy is 5% permethrin. 

Diagnosis is usually performed with a wet mount or Papanicolaou smear. 

Microscopic investigation for the presence of blas-tospores or pseudohyphae saline wet mount has a sensitivity of 40% to 50%, while a potassium hydroxide (KOH) preparation has a sensitivity of 50% to 70%.7... 

The three principal microscopic examinations performed on stool specimens are direct wet mount, wet mount after concentration, and permanent stain. Although each examination can contribute to diagnosis, the yield of some methods is small with certain kinds of specimens. As a minimum, formed specimens should be examined by a concentration procedure. Soft specimens should be examined by concentration and permanent stain, and, if submitted fresh, by direct wet mount. Loose and watery specimens should be examined by wet mount and permanent stain. If specimens are received in fixative and the consistency is not known, concentration and permanent stain should be performed. Other examinations may be helpful. Special procedures which may assist in the diagnosis of specific parasites are noted below in discussions of the parasites. 

Type of specimen Direct wet mount Method Concen- tration Permanent stain... 

The direct wet mount made from unconcentrated fresh feces is most useful for the detection of the motile trophozoites of intestinal protozoa and the motile larvae of Strongyloides spp. It is also useful for the detection of protozoan cysts and helminth eggs. For fixed feces, the direct wet mount may allow the detection of parasites which do not concentrate well. This method is also useful for the examination of specific portions of feces, such as flecks of blood or mucus. 

Direct wet mounts are prepared by placing a small drop of 0.85% saline toward one end of a glass slide (2 by 3 in. [ca. 5 by 7.5 cm]) and a small drop of appropriate iodine solution (see below) toward the other end. With an applicator stick, a small portion of specimen (1 to 2 mg) is thoroughly mixed in each diluent, and a no. 1 cover slip (22 mm) is added. The density of fecal material should be such that newspaper print can be read with difficulty through the smear. The material should not overflow the edges of the cover slip. Grit or debris may prevent the cover slip from seating and may be... 

For the examination of wet mounts, the light of the microscope must be properly adjusted. To achieve optimal resolution, the condenser should be centered and focused for Kohler illumination (racked up). To achieve contrast of the objects in the field, light intensity is diminished with the iris diaphragm of the condenser rather than by lowering the condenser. 

A solution of buffered methylene blue (pH 3.6) may be used as a vital stain for the examination of fresh specimens for protozoa. The wet mount is prepared as described above, with buffered methylene blue substituted as diluent and 5 to 10 min allowed for the dye to become incorporated in the organisms before examination. Organisms become overstained in 20 to 30 min. 

Prepare wet mounts as described above, and examine them. 

A funnel with a clamped rubber tube on the stem is placed in a ring stand. A circular mesh screen is placed across the funnel approximately one-third from the top, a portion of coarse fabric such as muslin is placed on the screen, and feces is added. Tap water at 37°C is added so that the water just touches the feces. Let the specimen stand 1 h, remove 2 ml of fluid from the stem, and centrifuge the sample at 300 x g for 3 min. Prepare a wet mount of sediment, and examine it for larvae. 

Permanent stains of fecal smears are most needed for the detection and identification of protozoan trophozoites, but they are also used for the identification of cysts. Wet mounts of fresh feces, even with stains such as methylene blue, are not as sensitive for trophozoites and therefore do not substitute for permanent stains. It is sometimes difficult to identify cysts which are detected in wet mounts thus, for each specimen, regardless of consistency, it may be worthwhile to fix a portion in PVA fixative or to prepare two fecal films fixed in Schaudinn fixative so that permanent stains can be performed if needed. Permanent stains also provide a permanent record and are easily referred to consultants if there are questions on identification. 

The last material aspirated is most likely to contain amebae. Material may be examined microscopically in wet mounts and permanent stains, and in addition, it can be cultured for amebae if bacteria are also added to the culture as described below. Abscess material is often thick and difficult to examine. It may be treated with streptokinase and streptodonase enzymes to liquefy the specimen. 

The sediment may be used for microscopic examinations for amebae (wet mounts and permanent stains) and for the culture of amebae. 

At 24 and 48 h, remove 1 drop of liquid from the lowest point of the overlay, and prepare a wet mount. 

Fluids such as tissue aspirates, cyst fluid, bronchial washings, cerebrospinal fluid, pleural fluid, and peritoneal fluid can be examined directly, or they can be centrifuged and the sediment examined by wet mounts or stains (or both), depending on the parasite suspected, as described above for abscesses or tissue. 

Spread several drops of sediment on a glass slide (1 by 3 in.), and examine two such uncovered wet mounts for motile microfilariae. Allow the wet mounts to dry before they are fixed and stained. 

Prepare four or five similar wet mounts and examine them as described above. To. each slide, immediately add 2 drops of 1% acetic acid solution and mix it well (microfilariae will be killed and straightened). Allow the slide to air dry. 

Vaginal material is best submitted as liquid in a tube, although swabs submitted in a small amount of saline may be used. A drop of the material is covered with a cover slip and examined with reduced light. To culture, 1 or 2 drops of urine sediment or vaginal exudate are inoculated into tubes of warmed, modified Diamond medium. If vaginal swabs are submitted, the swab is immersed in the medium and pressed against the side of the tube to express material. Tubes are incubated at 35°C, and drops of culture are examined by wet mount at 48 and 72 h for motile trophozoites. 

Fluoro-Jade staining was performed as described in Schmued and Hopkins (2000) with modifications (Butler et al. 2002). The sections were wet mounted onto microscope glass slides and air-dried for 30 min at 37°C in an oven. Then, the sections were pretreated for 5 min in absolute alcohol, followed by 3 min in 70% ethanol, 3 min in 50% ethanol, and 5 min in distilled water. The slides were then immersed in a solution of 0.05% KMn04 for 30 min at room temperature, and stained for 30 min in a solution of 0.001% Fluoro-Jade B (Chemicon, Temecula, CA, USA) in 0.1% acetic acid. Finally, the slides were then rinsed in distilled water, dried, cleared in xylene, and coverslipped. 

The simplest and most reliable means of diagnosis is a wet-mount examination of the vaginal discharge. Trichomoniasis is confirmed if characteristic pear-shaped, flagellating organisms are observed. Newer diagnostic tests such as monoclonal antibody or DNA probe techniques, as well as polymerase chain reaction tests are highly sensitive and specific. 

A motility test is usefrd because B. anthracis is a nomnotile bacterium. Two motihty tests available are the wet mount and motihty medium variety. In a wet-mount preparation, organisms with Brownian movement or no movement will support the presence of B. anthracis. The presence of B. anthracis in a motdity medium preparation would be a single line of growth along the original inoculum stab (CDC, ASM, APHL, 2002). 

Prepare wet mounts using drops of the test fluids and observe under high power. 

Direct examination of tissue or body fluids believed to be infected can provide simple, rapid information to the clinician. Microscopic examination of wet-mount specimen preparations can provide valuable information regarding potential pathogens. Applications of this procedure with or without staining preparations include direct examination of sputum, bronchial aspirates, scrapings of mucosal lesions, and urinary sediment. The Gram stain is one of the... 

In males, demonstration of trichomonads in urethral specimens or urine sediment by wet mount is difficult, and diagnosis depends largely on culture. Specimens from males should be taken prior to first voiding because the small number of trichomonads in males may be reduced by micturition. " ... 

Water from an ice bath was passed through the jacket to dissipate heat generated from the sonifer. Lysis was checked visually by monitoring the number of whole cells seen in a wet mount under an optical microscope at 1000X magnification. 

---