Lab 4: Thick and Thin Blood Film for Malarial Parasite Identification

thick and thin blood smears stained with Giemsa or Wright’s stain

DEFINITIVE DIAGNOSIS and remains as the GOLD STANDARD for malarial parasite identification

ring forms

In malarial infections caused by plasmodium falciparum only the __________ of the parasites may be found in the blood smear

T

T/F: In malarial infections, 10 days after the symptoms begin, gametocytes may be found as well.

6-8 hours

There are no standard recommendations on how often the blood smears should be taken in order to diagnose malaria but obtaining smears every _______ is usually appropriate.

T

T/F:

Microscopic examination of thick and thin blood smears is the easiest and most reliable test for malaria.

T

T/F: The microscopy of thick blood films is the usual diagnostic test for plasmodium falciparum malaria

T

T/F: The Parasite count is usually assessed by the thick films either counting the parasites per microscope field or by counting parasites per hundred white blood cells.

T

T/F: Plasmodium parasites are more concentrated on thick films than on thin films and are easier to see and identify

T

T/F: The identification of the specific type of plasmodium species cannot be done in the thick film

Thick films

most useful for detecting the presence of parasites because they examine a larger sample of blood

Thick films

allow the microscopist to examine a larger number of red cells for the presence of parasites, and low parasitaemias can be more readily identified

Thin films

are preferred to examine the morphology of parasites and determine species

Thin films

used to confirm the malaria parasite species when this cannot be done in the thick film

T

T/F: A well-prepared thin film consists of a single layer of red and white blood cells spread over less than half the slide

T

T/F: Plasmodium specie is an intracellular parasite (We can find them inside the red blood cells)

1. POORLY POSITIONED
2. TOO MUCH BLOOD
3. TOO LITTLE BLOOD
4. GREASY
5. CHIPPED

Factors affecting blood film quality

1. Immerse the thick film in tap water for 2-3 minutes.
2. Fix the thin film with methanol for 2-3 minutes.
3. Dry, then stain with Giemsa for 30 minutes.
4. Rinse with tap water and allow to air dry.
5. Observe under the OIO

Procedure for Staining the Blood Films

Robin’s egg blue

\[Color\]

Plasmodium cytoplasm

Crimson or Violet

\[Color\]

Nuclear chromatin

Yellowish or Salmon pink

\[Color\]

Cytoplasm of infected RBC

Brown or Black

\[Color\]

Malarial pigment

* Fungus
* Airborne pollen and spores
* Dirt and bacteria
* Contaminated water

Contaminants and artefacts

Thick films

* 15 – 20 WBC/THICK FILM FIELD
* 100 GOOD FIELDS (OIO)

Thin films

* Examine until presence and specie of malaria parasites have been identified
* At least 800 FIELDS before declaring slide NEGATIVE

RECORD number of parasite per 200 WBCs

200 WBCs + 100 or more parasites =

CONTINUE counting until 500 WBCs

200 WBCs +

Parasite count for Heavy parasitaemias

\
100 WBCS (or the total number in FIVE oil-immersion fields )

* Clinician needs to know the severity of the infection
* Clinician needs to know how the infection is responding to treatment
* Parasite counts are important in P. falciparum infections, which are always considered potentially dangerous
* District health officers should be aware of the severity of the cases being observed in health facilities in their area
* Determination of the density of infections may be required in cross-sectional and epidemiological investigations or in special studies, such as monitoring the therapeutic efficacy of antimalarial medicines.

Parasite density must be known because:

1-10 PARASITES PER 100 OIO THICK FILM FIELD

\[Plus system\]

\+

\[Plus system\]

\++

1-10 PARASITES PER SINGLE OIO TFF

\[Plus system\]

\+++

MORE THAN 10 PARASITES PER SINGLE OIO TFF

\[Plus system\]

\++++

Glass slide with yellow band

spreader slide

T

T/F: A good field should posses 15-20 white blood cells per thick film filmed fields

T

T/F: Thin films are not routinely examined except when:

* When thick film is too small
* Was lost during staining
* Became autofixed or unexamined for some reason

* one for the white blood cells (leukocytes)
* one for the malarial parasites

Why do we need TWO tally counters?

pH of 7.2

It is the pH that is essential for good staining of the parasites and the white blood cells elements.