Concentration

Technique
CM/Para
 Provide the ability to detect small numbers
of parasite
Purpose: aggregate parasite present into
small volume and remove debris.

Concentration Fresh or preserved stool

Detects protozoan cyst, oocyst, helminth
Technique eggs and larvae.
Two types of concentration methods:
a. Sedimentation
b. Flotation
Sedimentation Method Floatation Method

Chemicals used have a lower sp gr than Chemicals used have a higher sp gr than
the parasites. the parasites.

Parasites are concentrated and settle to Parasites are less dense than the solution
the bottom of the tube and float to the surface

Sediment after centrifugation is examined Material from the surface film is examined
microscopically microscopically
 Widely used sedimentation technique
Principle: Specific gravity based
Formalin-Ethyl Ethyl acetate is added to a saline-washed
formalin-fixed sample and then centrifuged.
Acetate Parasite settle, debris floats.
Sedimentation Advantage: Good recovery of most parasites
Procedure and easy to perform
Disadvantage: Contains more fecal debris
and is more challenging.
Formalin-Ethyl Acetate Sedimentation
Concentration
1.Mix the specimen well.
2.Strain 5ml of the fecal suspension (more or less depending
on its consistency) through wetted cheesecloth-type gauze
placed over a disposable paper funnel into a 15 ml conical
centrifuge tube.
3.Add 0.85% saline or 10% formalin through the debris on the
gauze to bring the volume in the centrifuge tube to 15 ml. Distilled water may
be used; however, Blastocystis hominis may be deformed or destroyed.
4.Centrifuge at 500 × g for 10 minutes.
5.Decant supernatant. Add 10 ml of 10% formalin to the sediment and mix
thoroughly with wooden applicator sticks.
6.Add 4 ml of ethyl acetate, stopper the tube, and shake vigorously in an
inverted position for 30 seconds. Carefully remove the stopper.
7.Centrifuge at 500 × g for 10 minutes.
8.Free the plug of debris from the top of the tube by ringing the sides with an
applicator stick. Decant the top layers of supernatant.
9.Use a cotton-tipped applicator to remove debris from sides of the centrifuge
tube.
10.Add several drops of 10% formalin to resuspend the concentrated specimen.
Proceed with applicable testing.
 Principle: Specific gravity based
Zinc sulfate, SG 1.18 to 1.20
Zinc Zinc sulfate is added to the sample and
centrifuged
Sulfate Parasites floats to the surface, debris sinks
to the bottom of the tube.
Flotation Advantage: More debris is removed.
Technique Disadvantage: Some parasites will be
missed.
 Zinc Sulfate Flotation
Technique
1.For fresh, unpreserved stool, dilute the specimen
1:1 in physiological saline.
2.Mix the specimen well.
3.Strain 5 mL of the fecal suspension (more or less
depending on its consistency) through wetted
cheesecloth-type gauze placed over a disposable
paper funnel into a 15 mL conical centrifuge tube.
(Conical paper cups with the tips cut off are
sufficient). Remaining suspension can be stirred
and pressed gently with an applicator stick
through the gauze.
4.Add 0.85% saline or 10% formalin through the
debris on the gauze to bring the volume in the
centrifuge tube to 15 mL. Distilled or tap water
may be used; however this may deform or
destroy Blastocystis species.
5.Centrifuge at 500 × g for 10 minutes.
6.Decant supernatant. Add 10 mL of flotation
solution to the sediment and mix thoroughly with
wooden applicator sticks.
Zinc Sulfate Flotation
Technique
7. Centrifuge at 500 × g for 5 minutes, allow
the centrifuge to stop without using the brake.
8. Carefully remove the tube from the
centrifuge without disturbing the pellet, place
in a rack.
9. Avoiding disturbing the surface, add
sufficient flotation solution to completely fill the
15 mL tube, ensuring a slightly convex
meniscus is formed.
10. Place a 15 mm x 15 mm coverslip over the
meniscus.
11. Set a timer for 10 minutes.
12. After 10 minutes flotation, carefully remove
the cover slip, ensuring that a drop of solution
remains hanging from the coverlsip, and place
this on a microscope slide.
13. Perform microscopy for parasites within 15
minutes of preparing the microscope slide.
Thank you