Potato (Solanum tuberosum L.) ranks as the fourth most important staple food globally, after rice, wheat, and maize. In Pakistan, Potato virus Y (PVY) and Potato leafroll virus (PLRV) are among the most critical pathogens affecting potato crops, causing yield losses of up to 90%. Fifteen advanced potato lines from the Potato Program of the Horticultural Research Institute (HRI) were screened for resistance to these economically significant viruses. The experiment was conducted in the glasshouse of the Crop Diseases Research Institute, NARC, Islamabad, during 2023-24, following a completely randomized design with three replications per clone. For screening, small tubers of fifteen potato clones, including one check variety, were planted in pots within the glasshouse. PVY was mechanically transmitted, while a chip graft inoculation assay was used for screening against PLRV. Symptoms began to appear two weeks after inoculation, and data were recorded four weeks post-inoculation. A double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was performed on all plants to confirm the presence of both viruses serologically. The findings revealed that the line HRI-P1 remained asymptomatic and was categorized as highly resistant to both viruses, based on symptom observation and ELISA testing. Six clones exhibited varying levels of resistance, three showed moderate susceptibility, one was susceptible, and three were highly susceptible to PVY. The study highlights the availability of valuable sources of resistance within the Potato Program. HRI, NARC, which could be utilized in breeding potential future varieties. The disease-resistant genotypes identified will be instrumental in future potato breeding programs.