Saline Wet Mount Microscopy: Introduction, Principle, Preparation, Result

Saline Wet Mount Microscopy: Introduction, Principle, Preparation, Result Interpretation, Limitations and Related Videos

Introduction of Saline Wet Mount Microscopy

Saline wet mount microscopy of the stool or stool wet mount microscopy is the simplest and basic method for the study of feces and is applicable in every medical laboratory even in a small setup. Saline wet mount microscopy uses for the following purposes-

To observe live trophozoites ( e.g. Entamoeba histolytica/dispar, Girdia lablia, Trichomonas, etc.) and larvae of parasites ( e.g. Strongyloides stercoralis) are motile except inactive forms.
To find out eggs, cysts, and oocysts of different parasites ( Helminths, protozoa, and coccidian parasites).
To determine the presence of leukocytes and erythrocytes in a fecal smear.
It also gives clues about the motility of bacteria (Shigella non-motile causing bacillary dysentery whereas Vibrio cholerae shows darting motility which is a causative agent of cholera).
It also remarks on the presence of fungal elements ( yeast cells, hyphae, or fungal spores).
It also visualizes the presence of non-parasitic structures like Charcot Leyden crystal, muscle fibers, fat globules, starch cells, vegetable fibers, hair, etc.
The growth of yeasts and bacteria on culture media may be differentiated by this simple technique.
The shape of bacteria, cocci, or bacilli can be differentiated from the colony of the culture plate by saline wet mount microscopy.

Principle of the Saline Wet Mount Microscopy

Saline wet mount preparation for stool uses analyzing a stool specimen in coprology (study of feces). It utilizes a physiological saline solution (0.85% NaCl ) as an isotonic media to maintain the cellular structure of the various organisms as well as our cells that are found in stool and other stools too.

Requirements for Saline Wet Mount Microscopy

Physiological saline ( 0.85% NaCl)
Specimen: stool
Sterile bamboo sticks or a low cone on the end of a wooden applicator stick
Clean and grease-free slides and
Cove slips(22- by 22-mm)
Microscope
Gloves

Saline wet mount of stool Preparation

First, wear the groves.
Take a clean and grease-free slide.
Add one drop of physiological saline and then add a stool equivalent to a match stick head (2 mg) with the help of a stick.
Mix it properly and apply a coverslip over a uniform suspension without creating bubbles.
Note: If a fresh stool specimen is received and if blood and mucus are present, the specimen should be examined as a direct mount making sure to sample the bloody areas.
Examine the entire 22- by 22-mm coverslip systematically with the low power objective (10X ) and low light intensity.
If any suspicious objects encounter, examine them with the high dry objective (40X).

Result Interpretation of Feces Saline Wet Mount Microscopy

Presence of active trophozoite/s: Motile retractile bodies
Cyst, oocyst, egg, inactive trophozite/s, larvae: Retractile bodies and finally focus at high dry power field

Keynotes on Saline Wet Mount Microscopy

There is little difference between normal and physiological saline. Physiological saline is 0.85% NaCl whereas normal saline is 0.9% NaCl.
Gram’s iodine is not applicable for staining parasitic organisms and for this D’Antoni’s iodine uses.
Oil immersion examination is also preferred in parasitology for the permanent stained smear of parasites.
Intestinal protozoa can not conform on the basis of a wet mount alone and thus permanent stained smears require to confirm the specific identification of suspected organisms.

Limitations of Saline Wet Mount Microscopy

Due to the lack of stain, it is difficult to get morphological details.
Inappropriate preparation of the smear may hide parasites.
Improper adjustment of the microscope in relation to the objective may create problems.

Saline Wet Mount Microscopy Related Videos-

Heavy load of parasites in stool|| Stool microscopic examination|| Trichomonas hominis in a saline wet mount of feces as shown below-

Trichuris trichura or whipworm under saline wet mount at 40X objective under the microscope –
Features: Barrel shape
Mucus thread at each pole as shown in the video

Egg of Taenia species
They are spherical, brown in color (bile stained), measure 30-40 µm in diameter, and are surrounded by embryophore which is brown, thick-walled, and radially striated. Inside embryophore, the hexacanth embryo (oncosphere) presents three pairs of hooklets. They do not float in a saturated solution of common salt (brine solution) and are viable for 8 weeks.

Roundworm or Ascaris lumbicoides egg under the microscope in saline preparation
egg-infertile
Finding of eggs
In stool: Direct microscopic examination of a saline emulsion of the stool
Concentration methods may be used.
Note: The fertilized egg floats in a salt solution.
Unfertilized eggs do not float.

Hymenolypsis (now called Vampirolepsis) egg in a saline wet mount of stool: showing hooklets egg of Hymenolepis liberated in feces by the gradual disintegration of terminal segments

Spherical or oval in shape, 30-45 µm
Two distinct membranes
Outer membranes are thin and colorless
The inner embryophore encloses an oncosphere with 3 pairs of hooklets.
Space between two membranes –filled with yolk granules and polar filaments emanating from little knobs at either end of embryophore.

Egg of hookworm ( Ancylostoma duodenale or Necator americanus) in saline preparation of stool under the microscope
Egg features-
Shape: oval or elliptical with flattened poles( one pole more often flattened than the other), size: 65 X 40 um, color: colorless ( no bile stain), dark brown as stained with iodine. Shell: very thin transparent hyaline shell membrane, appears as a black line and contains: segmented ovum with 4 blastomeres, has a clear space between eggshell and segmented ovum. Float in saturated NaCl. Type: A( fresh stool): 4,8, 16 grey granular cell clear blastomeres. Type: B(a few hours old): a uniform mass of many grey granular cells. Type: C( 12-48 hr): the whole egg is filled with larva, embryonated.

Enterobious vermicularis ( common manes pinworm or threadworm or seatworm) – eggs and some are with larva in a saline wet mount of feces –
Shape: oval, planoconvex.

Size : 50-60μm x 20-30μm.
Surrounded by double-layered eggshell
Embryonated when passed fresh; contains a tadpole larva inside.

Entamoeba coli Cyst with 8 nuclei in iodine wet mount as shown below-

Giardia lamblia cysts in iodine wet mount as shown below ( look at center)-

Entamoeba histolytica (Amoeba) trophozoites and cyst in LPCB preparation as shown below-

Blastocystis hominis cyst in Sargeaunt stained slide under the microscope as shown below-

Oocyst of Cyclospora cayetanensis ( coccidian parasite) in a saline wet mount of stool under the microscope as shown below-

A very simple Saline wet mount technique helps you to identify yeast cells of candida from bacteria-

Bacteria and Fungus in saline wet mount Microscopy at Various magnifications 400X, 800X and 1600X

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