OBJECTIVE The study was to evaluate the diagnostic efficacy of nested polymerase chain reaction (nPCR) using primers targeting B1 gene of Toxoplasma gondii (T. gondii) with Witmer Desmonts coefficient (WDC) technique in intraocular fluids of clinically suspected toxoplasma retino choroiditis (TRC) patients. MATERIALS AND METHODS Two hundred and seventy eight specimens from 189 patients (25 TRC patients and 164 controls) consisting of 189 serum samples and 89 intraocular fluids were included in the study. The clinical specimens were categorized into TRC patients (typical TRC lesion-group I & atypical TRC lesion-group II) and controls (voluntary blood donors-group III, patients undergoing uncomplicated cataract surgery-group IV, ocular inflammation of non-toxoplasma origin-group V). Detection of anti T. gondii IgG and IgM antibodies in serum samples and intraocular fluids were performed and WDC was calculated by the standard method. The standardized nPCR was applied on the 89 intraocular fluids. RESULTS Clinical diagnosis of TRC based on fundus examination was considered to be the "gold standard." Anti T. gondii IgG/IgM antibodies were detected in serum by ELISA in 95.6% of 25 clinically suspected TRC patients (gp I and II), 28% of gp III, 40.4% of gp IV, and in 58.3% of gpV. Witmer Desmont's coefficient was positive in 72.7% (16/22) and nPCR in 59.1% (13/22) of TRC patients (gp I and II). Both WDC and nPCR were negative in all the controls. The difference in sensitivity of WDC and nPCR was not statistically significant (p=0.5247). CONCLUSIONS Though both WDC and nPCR were reliable diagnostic techniques for ocular toxoplasmosis, nPCR is more acceptable because of the amount of specimen(s) required, rapidity, cost effectiveness, and direct evidence of T. gondii DNA in the intraocular fluids.