Giemsa Stain (for Plasmodium species)

Introduction
Giemsa stain is used to demonstrate the presence of Plasmodium species in thick and thin blood films. A thick film is about 30 times more sensitive than a thin film; detecting about 20 parasites per μl. Thick films are therefore the most suitable method for the rapid detection of the parasite. A thin film is required to confirm the Plasmodium species if this is not clear from the thick film. Thin films are also of value in assessing whether a patient with Plasmodium falciparum malaria is responding to treatment in areas where drug resistance is suspected.


Method
Thin films of blood or marrow
• Fix in methanol for 5min
• Place in a Coplin jar containing Giemsa stain diluted 1:10 for 20min
• Rinse in distilled water
• Drain and dry at room temperature
• Mount in DPX or leave unmounted and use oil immersion

Thick films for malaria parasites
• Air dry the slides
• Place in a Coplin jar containing Giemsa stain diluted 1:50 at pH 7.2 for 1hr
• Wash with distilled water (flushing the stain from the slides and the staining container is necessary to avoid the films being covered with a fine deposit of stain)
• Differentiate in 1:1,500 acetic acid (control by viewing at intervals under a microscope. Sections should have an overall pink colour, with the nuclei blue and eosinophil granules red)
• Rapidly rinse in distilled water
• Air dry


Interpretation

Positive Result

Negative Result

Quality Control Organism
Positive control
Plasmodium species

Negative control
A proven negative smear may be used as the negative control.


Technical Information
Rapid diagnostic tests (RTDs) are available as alternatives for microscopy. These tests detect three main groups of antigens including Histidine-rich protein 2 (HRP2) specific to P. falciparum, plasmodium lactate dehydrogenase (pLDH), and Aldolase. These products are available in the forms of plastic cassettes, cards, dipsticks, and hybrid cassette-dipsticks. Factors such as parasite prevalence, availability of skilled personnel and resources, the capacity for maintaining quality assurance of microscopy and RDT, and the need for quantitative assessment of parasite density need to be considered when selecting microscopy, or an RTD, as an identification method