Activity No.

18
Knott’s Concentration Technique

Objectives:

After discussing the exercise, the student should be able to:

1. state the procedure of Knotts’s Concentration method
2. explain how to recover microfilariae

Discussion:

There are cases wherein the density of microfilariae is low. In this case, blood is mixed
with formalin and then centrifuged. The sediment is spread like a thin blood film and is stained.

Materials:
2 mL blood
10 mL of 2% Formalin solution (2mL formaldehyde solution in 98% NSS)
15 mL centrifuge tube
Pipette
Slides

Procedure:

1. Mix the blood with the 2% formalin in the centrifuge tube)

2. Centrifuge at moderate speed for 2 minutes
3. Decant the supernatant
4. Pipette the sediment on the slide and examine under the microscope. The sediment
maybe stained with 1:10,000 aqueous solution of either methylene blue or azure blue or
Giemsa stain and examine under the microscope.
 Activity No.18
Knott’s Concentration Technique

Name: _____________________________ Score: ____________________

Yr. /Sec. ____________________________ Lab Instructor: ______________

Guide Questions:
1. What are the identifying characteristics of microfilariae?

2. What is the ideal time of collecting blood sample for detection of microfilariae?

3. What are the conditions where microfilariae are low in density in blood?

Instruction:
Illustrate the procedure of Knott’s concentration technique
 Activity No. 14
Harada Mori Stool Culture Method

Objectives:
After discussing the exercise the student should be able to:
1. explain how to perform the Harada Mori or Test tube stool culture method.
2. identify the filariform larvae of hookworm.
Discussion:
Harada Mori is a culture method for stool found positive with Hookworm egg. This
technique uses filter paper strip wherein stool is smeared and is placed in a test tube with boiled
distilled or heat sterilized water. Hookworm egg will developed into rhabditiform larvae then into
filariform larvae. This will moved downwards against the upward capillary movement of water.
Since the movement is downward, the filariform will be recovered from water at the bottom of
the tube. For Strongyloides stercoralis larvae it may move upwards and accumulate at the upper
end of the filter paper strip.
Materials/Reagents:
Test tubes and racks applicator stick
Distilled or heat sterilized water aluminum foil 1”x1” size
Feces positive for hookworm ova beaker with hot water
Filter paper strips slides and cover slips
Forceps pipette with rubber bulb
Procedure:
1. Put about 7 mL distilled water or heat sterilized water in the test tube.
2. Using an applicator stick, smear thinly on the filter paper strip about a half gram of feces
leaving about 5cm space at each end of the filter paper unsmeared.
3. Insert the smeared filter paper strip into the test tube with the 5cm unsmeared portion
touching the water near the bottom of the tube.
4. Cover the tube with aluminum foil to prevent flies and dust from entering the tube.
5. Keep the tube in a dark place at RT for 10 days. After 10 days the larvae will have
developed to the filariform stage. This is now ready for morphological differentiation.
6. Immerse the test tubes in very hot water (but not boiling) for 15 minutes to kill the larvae.
7. Remove the filter paper with a pair of forceps and transfer the contents of the test tube to
a centrifuge tube.
8. Centrifuge for 3-5 minutes. Pipette and throw away the supernate carefully.
9. Transfer the sediment to a slide, put a cover slip and examine under LPO for the
presence of larvae; identify the species, examine under HPO.
 Activity No. 14
Harada Mori Culture Method

Name: _________________________ Score: ____________________

Yr./Sec. ________________________ Date: ______________

Guide Questions:
1. What is the importance of Harada Mori stool culture method?

2. Why do you have to use heat sterilized water?

3. Why do you have to develop the hookworm egg into filarifom?

Instruction:
Draw and label the set-up of Harada Mori