To detect IgG antibody in the serodiagnosis of visceral leishmaniasis (VL), a recombinant antigen rK39, which is part of a Leishmania chagasi kinesin-related protein, has been used successfully and showed high sensitivity and specificity. We report production of a recombinant protein rKRP42, which is part of an L. donovani kinesin-related protein and a homolog of rK39, and its application in an enzyme-linked immunosorbent assay (ELISA) for the diagnosis of VL. When rKRP42 and rK39 were compared, amino acid sequence analysis showed 89.3% identity and 98.7% homology, with rKRP42 having 39 more amino acids than rK39. The ELISA using rKRP42 showed a sensitivity of 94.6% (70 positive samples among 74 from VL patients) and a specificity of 99.3% (148 negative samples among 149 samples from Japanese controls), whereas the sensitivity of the commercial rK39 dipstick test was 93.2% (69 positive samples among 74 from patients with VL). The rKRP42 is a promising new antigen in developing immunodiagnostic methods for VL.