Transcript Evaluation of Wet Mount and KOH Preparations

Evaluation of Wet Mount
and KOH Preparations
Phase 1 Pharmacokinetic Trial of Two Intravaginal
Rings (IVRs) Containing Different Dose Strengths of
Vicriviroc (MK-4176) and MK-2048
MTN-028 Study Specific Training
Wet Mount for Clue Cells and
Trichomonas
• Remove the swab from the tube with saline and dot a
liberal amount onto a glass slide
• Place a coverslip over the specimen
• Scan the slide on 100X and 400X
–
–
–
–
Positive for clue cells: >20%
Negative for clue cells: <20%
Positive for Trichomonas: any motile trichomonads seen
Positive for yeast: any budding yeast and/or pseudohyphae
seen
KOH preparation for Yeast
• Place a coverslip over the KOH preparation
after smelling for amine odor
• Scan the slide at 100X and 400X
– Positive for yeast: any budding yeast and/or
pseudohyphae
– Negative for yeast: no yeast cells seen on slide
Gram stain of vaginal flora
Normal flora
BV
Wet Mount Evaluation
• Minimum of 5 fields should be evaluated.
• Ask yourself: What is your first impression?
• Assess the epithelial cells and background
bacteria present.
Shape: Symmetrical rods or pleomorphic
coccobacillary?
Numbers: Fewer or many?
Normal flora: No clue cells
1000X
400X
BV: clue cells and WBC
1000X
400X
BV: Clue cells
1000X
400X
Wet Mount Evaluation: Clue Cells
• The following tips should be utilized for determining clue
cells:
1. Count the number of distinguishable epithelial cells in your field of
view.
2. To determine if any of the epithelial cells are clue cells, it is
important to study ONLY THE BORDERS OF THE CELL.
Note: Normal variation in cell membranes can result in a “grainy”
appearance of the cell and can mimic bacterial adhesion.
3. To determine the percentage of clue cells in your field:
a. Count the number of clue cells and divide that number by the
total number of distinguishable epithelial cells.
Normal Cells
400X
Clue Cells
400X
Normal Cells
400X
Clue Cells
400X
Clue Cells
400X
Normal Cells
400X
Normal Cells and yeast
400X
Wet Mount Evaluation: Yeast
• Determining Yeast in Wet Mounts or KOH preps
• In order for yeast/pseudohyphae not to be mistaken
for amorphous material, nuclei or artifacts there
must be “budding”.
“Bud”
Pseudohyphae with “buds”
Budding yeast
Pseudohyphae
400X
Budding yeast
400X
Pseudohyphae and
budding yeast
Budding yeast
KOH 400X
KOH 400X
Pseudohyphae and
budding yeast
KOH 400X
Pseudohyphae,
budding yeast and
amorphous material
KOH 400X
Pseudohyphae,
budding yeast and
amorphous material
KOH 400X
Budding yeast
KOH 400X
Amorphous material
KOH 400X
Amorphous material
KOH 400X
Amorphous material
KOH 400X
Amorphous material
KOH 400X
Pseudohyphae,
budding yeast and
amorphous material
KOH 400X
Pseudohyphae and
budding yeast
KOH 400X
Wet Mounts: Other Common
Morphotypes
Trichomonas vaginalis
Sperm
Neutrophils
Red blood cells
Epithelial Count = 8
Clue Cells = 3
(38%)
BV Morphotype
Dominated
Flora
Clue Cell Negative
Epithelial Count = 4
Mixed Flora
Epithelial Count = 6
Clue Cell Negative
Epithelial Count = 11
Clue Cell = 3
(27%)
Budding yeast
Artifact
“Bud”
Pseudohyphae
Clue cell
RBCs
artifact
WBC covered with
bacteria
Microstructures of leaves called stellate hairs found in a vaginal KOH prep
Know Your Microscope
Adjusting the Condenser for Contrast of Cells
Move the condenser
up and down
Adjustments for Optimal Illumination
Centers the
condenser
Adjust condenser
aperture diaphragm
Adjust light source diaphragm
Care and Cleaning of Microscope
• Cover the scope when not in use
• Use water or mild cleaning solutions for the body of
the scope
• Clean the lenses with optics cleaning paper, Kimwipes,
or a cotton cloth (do not use facial tissue)
• Use an optics cleaning solution to remove oily or
greasy dirt (fingerprints, immersion oil)
• Check the alignment of the condenser with “Kohler
illumination”