**4. Serological methods**

Serological methods are highly sensitive and non-invasive. They are comparatively more suited for diagnosing VL in endemic regions. These methods are either based on detection of antibodies (produced against parasite by polyclonal activation of B cells) or antigens. Many conventional methods for antibodies detection for instances, gel diffusion, complement fixation test, indirect haemagglutination test, indirect fluorescent antibody detection test (IFAT), and counter current electrophoresis have been evaluated with varying sensitivities and specificities.

Direct agglutination test (DAT) has been found to be 91-100% sensitive and 72-100% specific in various studies elsewhere in the world. [26]

Detection of anti-K39 by immune-chromatographic strip testing is a rapid and non-invasive method of diagnosing Kala azar. It entails a good sensitivity and specificity. In symptomatic patients, anti-K39 strip-test sensitivity is high (90-100%), while specificity might vary by region. [3]

Two urinary antigens of 72-75 and 123 kDa have been reported to be very useful in diagnosis and prognosis of Kala azar with sensitivity of 96% and specificity of 100%. [26]

DNA detection methods: a variety of nucleic acid detection methods targeting both DNA and RNA have been developed. The most suitable target for the DNA based diagnosis is kinetoplast DNA minicircle (K-DNA). The leishmania polymerase chain reaction (PCR) assays using peripheral blood as clinical specimen showed to be a highly efficient non-invasive alternative with sensitivity varying from 80-100%. [27]

The *Leishmania* skin test (LST): LST is a measure of delayed hypersensitivity to leishmanial antigen. The test remains negative through the period of active disease. The change from negative to a positive LST is regarded as a prognostic sign. [28]
