Microbiology ( Faecal Occult Blood Test )

Fecal Occult Blood Test (FOBT) and Ova, Cyst and Parasite test are two of the most common tests done in the Parasitology bench.

FOBT is done to determine the presence of hidden (occult) blood in the stool. This may be caused by undetected gastrointestinal bleeding, which could be an indicator for colorectal cancers. The test is based on immunochromatography. It uses antibodies against haemoglobin, a main component of blood. . The presence of haemoglobin will cause it to bind to the antibody, and cause a colour change in the test strip.




The stool needs to be sampled at 6 places to allow for homogenous sampling of the stool.


After the sampling vial has been inoculated with the stool, a test strip is then inserted into the test solution in the vial. The result should be visible within 5 minutes. Any result obtained after the 5 minutes window period will be considered void, as there may be a chance of false positives occurring.




The control line must be working in order to obtain a validated test result. The control line ensures that all the procedures have been done correctly. If the control line does not show, it means that the test is equivocal and cannot be used. Thus, a retest needs to be done.


In ova, cyst and parasite testing, there are two main procedures – the fixed smear and the wet mount. Fixation of the stool sample helps to preserve the state of the parasitic organisms in the stool. The wet mount enables the visualization of motile parasitic organisms in the stool.

Fixed Smear
1. Mix 1 part stool to 3 parts of Zn Poly-Vinyl Alcohol (PVA) in the provided PARA-PAK tube.
2. Let stand for 30 minutes.
3. After 30 minutes, pour some of the Zn-PVA-fixed material onto a paper towel. Let most of the excess PVA be absorbed.
4. Once most of the excess PVA has been absorbed, spread some of the fixed stool material evenly onto a glass slide. The edges of the slide must be smeared with some of the fixed material to adhere better onto the slide.
5. Once dry, heat-fix for 3 hours on a slider warmer at about 35oC.
6. After which, perform a trichrome stain.

Wet Mount
1. 10 drops of surfactant is added to the remaining Zn-PVA fixed sample and shaken vigorously. The surfactant helps to break down the larger stool particles.
2. The Macro-Con conical filter tube is then attached to the PARA-PAK tube and inverted to allow filtration of the sample.
3. Once the sample has been filtered, the empty PARA-PAK tube and filter is discarded, leaving the MACRO-CON tube full of fixed sample solution.
4. Add buffered formaldehyde (formalin) (4.4 % w/w of solution) up until the indicator line on the tube.
5. Add 5 ml of clearene.
6. Spin at 2000 rpm for 10 minutes in the centrifuge.
7. Discard the supernatant and clean the sides of the tube with a clean cotton swab to remove excess clearene and formalin.
8. Resuspend residue in equal amounts of normal saline.
9. Place a drop of the resuspended solution onto a glass slide and mount with a coverslip.

View both the fixed smear and the wet mount under a microsope.


MUNA
0703791D

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On 2 August 2009.