Stool Examination: Wet Mount

10% KOH, Saline Wet Mount And

Lugol Iodine Preparation

Stool Examnimation
Intestinal parasites are commonly predominant as the leading reason of
diarrhea specifically between  children and  frequently  linked with  illness  and 
death  in  developing countries.  Parasites of diarrheal background are
prevalent, infecting a significant percentage of the human population in 3 rdworld
nations. Most commonly in the subcontinent region. Faeces are the most
repeated samples, are typically studied in the laboratory setup by microscopic
examination, that is basically contains of Direct  wet-mount preparation,
concentration, and permanent staining.Microscopically examination of wet-
mounts is the utmost broadly used technique for the identification of ova/eggs
and cysts of parasites from stool samples.  The precise analysis  of 
the situation  is  organized by  evaluation  of protozoan  cyst  or trophozoite;
helminthes ova/eggs or less commonly adult worms or larvae in the stool
specimens and smears by microscopically examined.

Direct Wet Mount KOH Preparation:

Direct wet-mount  preparation of  stool specimen is  widely used  for the
parasitological analysis in laboratories and also  for  the diagnosis  of 
intestinal parasites by microscopically method. KOH – Potassium
Hydroxide, is  extensively  used in  the direct wet-mount  preparation for
different specimens for examples fungal elements and fungi, hair stuffs, skin
scales, and nail scraping or other clinical samples with a small drops of 10%
Potassium Hydroxide – KOH on  a glass  slide. The  Potassium Hydroxide 
digests the  pertinacious/resolute cellular  debris and bleaches  pigments, 
loosens their the  sclerotic  constituents but don’t damaging their the
clinical constituents. So they give excellent view under microscopic examination.

Wet Mount Saline Method:

As the name suggested,

 It is made by the help of Normal saline by mixing a little amount

around 2 milligram of stool specimens.
  Take a small drop of normal saline on a glass slide and cover with a
cover slip. Normal saline is basically an Isotonic Solution which help
to maintain the osmotic pressures inside the cells and doesn’t not lyse
or destroy their structure.
 Under the microscope examine the smear, initially at low power 4X and
then focus on at 10X objectives.
 The Saline wet-mount procedure is widely used to help for the
detection and identification of parasitic cysts and trophozoite and ova
and larvae of helminthes parasites.
 It is predominantly valuable for identification of live and motile
trophozoite of parasites such as Entamoeba histolytica, Balantidium
coli, G.lamblia and helminthes larva such S.stercolaris andova’s
of ancylostoma hook worm, trichuris trichuira, Enterobius
vermicularis etc.

Lugol Iodine Staining Method:

Lugol Iodine is the same method as the wet Saline preparation but there is only
difference is despite the use of normal saline, we use lugol iodine in this
method. Lugol iodine help to stains the some cyst’s nuclei of protozoa such
as E.histolystica and easily seen under microscope. A basic disadvantage with
this technique isthe capability of lugol iodine to kill the motile parasites
trophozoite.
Iodine and Lugol Stain

Procedure for Wet Mount:

  Fully mix the stool specimen to make a  homogeneous or
heterogeneous fusion dependent on its integral material.
 Take an applicator stick, take the stool specimen and make a good
smear on a clean dry glass slide. Remove all gross fibers, debris and
particles.
 Add 1 to 2 drops of Lugol iodine or saline according to the method of
choice, and do it before the sample get dry. It is suitable to make 2
smear slides and it has extra advantage to recover parasites.
 Cover the sample with the help of cover slip. (Note: Avoid any air
bubbling by draw one side of the cover slip marginally into the
suspension and lowering it nearly to the slide before allowing it to fall).
 The quantity must be sufficient thick, a newspaper can easily be read
through the glass slide.
  If preferred, the cover slip might be sealed with the help of Paraffin
oil or Petroleum jelly. Sealing the cover slip preserves parasites from
moving.

 Interpretation, Examination and Results:

  Examine the sample initially at 4X and then focus at 10X by lower the
light of microscope. Start to view from one edge of the specimen and
observe consecutive bands. Low power at 10X examination comprises
full area of 22/22 millimeter cover slip (both in saline and iodine
preparation).

Also Read  Haemagglutionation Assay – HA

Parasites Seen Under Microscope

 If ova/egg, trophozoite or larva is seen, move the objectives lens at
40X and with the help of aperture increase the power from low to high.
This will help, object further visible, clear and more detailed with a
good resolution. High dry power examination must comprise at least
1/3rd of the cover slip part (both saline and iodine preparation).