Aim: The study’s aim was to compare the effectiveness of HiCHROMagar and PCRrestriction fragment length polymorphism (PCR-RFLP) in identifying Candida species in order to ascertain the advantages and drawbacks of each technique. Methods: One hundred and twenty five Candida strains were isolated from different clinical specimens from patients admitted to different wards of Era's Lucknow Medical College. The Candida isolates were from urine, Sputum, Blood, vaginal swab, pus and nail. Clinical samples werecultured on Sabouraud-dextrose agar (SDA) with chloramphenicol for 48 h at 37 °C.to obtain candida colonies. Wet mount,Gram staining, germ tube, sugar fermentation and assimilation were performed for all isolates. Candida isolates on SDA were subcultured on HiCrome Candida Differential agar (HiMedia, Mumbai, India) for 24-48 hours; plates were incubated at 37 °C and was re-evaluated by employing digestion of the ITS1– 5.8SrDNA–ITS2 region using Msp I restriction enzyme for RFLP and universal primers internal transcribed spacer 1(ITS1) and ITS4 for PCR amplification.