Dirofilaria immitis also known as heartworm, is a parasitic roundworm which causes canine dirofilariasis. Canine dirofilariasis is evolving as a potential zoonotic disease. Traditionally, diagnosis relied exclusively on the detection of microfilariae in the blood. However, this method proved inadequate since a significant percentage of animals harbour occult infection, in which adult worms are present but there are no circulating microfilariae. As the current diagnostic kits available for detection of this heartworm are expensive and it detects only the fecund female worm (found 6-7 months’ post-infection), pre-patent infection remains undetected. Moreover, in India co-infection of D. repens, D. hongkongenesis and D. immitis are common in dogs. A specific diagnostic method which can detect all stages of D.immitis infection alone is the need of thehour. Two antigens from D. immitis namely DiTCTP (Dirofilaria immitis Translationally Controlled Tumor Protein) and DiT33 (Dirofilaria immitis pepsin inhibitor precursor) could detect adult male and female worms as well as the immature worms. In this work, DiT33 and DiTCTP antigens were produced as recombinant proteins in E. coli. The recombinant DiT33 (43 kDa) and DiTCTP (40 kDa) antigens produced were purified and refolded by gradient urea dialysis. Dot blot which was performed to check the cross reactivity of the antigens with other species of Dirofilaria showed that the recombinant DiT33 and DiTCTP antigens do not cross react with D. repensand D. hongkongenesis infected dog serum but it is reactive against polyclonal sera developed against native antigens of D.immitis. Hence, the recombinant antigens (DiT33 and DiTCTP) can be used in the specific detection of all stages of D. immitis infection in dogs.