Parasitological diagnosis demonstration: Giemsa staining and microscopic detection of LD bodies

Once the criteria of VL case definition have been fulfilled and VL is suspected, confirmation is ideally by parasitological diagnosis. This involves microscope examination of biopsy material in order to identify Leishmania amastigotes (LDUs) in the sample. Material may be sampled from the spleen, bone marrow or lymph node and requires a high level of skill to conduct as well as hospital facilities to deal with any adverse events following an invasive procedure.

For the parasitological diagnosis of PKDL a thin skin biopsy, skin snip or slit skin sample is collected, smeared onto a slide and similarly examined under the microscope.

Examining Giemsa-stained aspiration samples

The sample is fixed with 100% methanol and stained with Giemsa dye before being examined with a light microscope, under the x100 objective with oil immersion. Watch the following video for the procedure.

Notes

1. In a busy diagnostic lab, when the turnover of samples is high, it may not be possible to review 10% of slides as this could lead to excessive workload. In such cases, it is best to agree upon a capped number of slides for systematic review.
2. DISCLAIMER: LSHTM and partners collaborating in this video can take no legal responsibility for the use of any information contained. Users implementing the techniques shown do so at their own risk and must take responsibility for local verification/ validation of these techniques before they are used for the diagnosis of patients.