Background: Meliodosis in an infectious disease affecting multi-organ system of man and animals caused by a bacterium, Burkholderia pseudomallei. The disease is potentially fatal and early detection is crucial for institution of life saving antimicrobial therapy. Culture isolation remains as a mainstay for definitive laboratory diagnosis of melioidosis. But it is less sensitive, laborious and timeconsuming task. In this report, an improved dot EIA test (patent pending) with user friendly assay protocol for serodiagnosis of melioidosis is described.Methods: Two optimised concentrations of antigens were dotted onto a nitro-cellulose membrane (Microfiltration System, Ca, USA, 0.45uM) divided into 9 mm × 4 mm. The membranes were blocked for 30 minutes in tris-buffered saline (TBS) wiht 5% nonfat skim milk. The membranes were rinsed and cut according to the divided size into test strips and placed in 48 well flat tissue culture plates (Costar 3548, UK.) The test strips were then incubated with patient serum at 1:100 in TBS for 1 hour at room temperature. After incubation, the test strips were washed with TBS for three times and each washing was done for 5 minutes. Subsequently, the diluted alkaline phosphatase conjugated goat anti-human immunoglobulin IgM, IgG and IgA isotypes were added into the respective wells and incubated for 1 hour at room temperature. After incubation the strips were washed three times in TBS as described above. Chromogenic substrate was used in the colour development reaction. Aftwer after 15 minutes the strips washing with distilled water to stop coloue development. The positive and negative results were determined by visual comparison of the dot intensity for each test with the positive cut off control.Results: The sensitivity and the specificity of the assay was 95% and 92%, respectively for the detection of IgG, IgM and IgA antibody isotypes. However, detection of IgG alone showed sensitivity value of 89.2%. The sensitivity value for the detection of IgM alone and IgA alone were 47.7% and 60% respectively.
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