Cervical cancer is a significant public health concern, disproportionately affecting women in less developed regions due to limited access to screening and vaccination programs. Despite advancements in cervical cancer prevention and treatment, there remains a need for efficient and cost-effective diagnostic tools. This study aimed to develop a multiplex real-time PCR assay to rapidly and accurately identify 15 human papillomavirus (HPV) genotypes.The primary objective was to design a screening method capable of simultaneously detecting HPV types 16 and 18, which account for over 70% of cervical cancers, as well as other clinically relevant high-risk and probable/possible high-risk. To validate the assay's performance, we compared its results with those obtained using the commercially available INNO-LiPA HPV Genotyping Extra II Assay kit (FujireBio, Tokyo, Japan). The developed assay successfully identified 15 HPV genotypes in a single reaction. Analysis of 150 positive and 40 negative clinical samples demonstrated excellent concordance between the two assays. The in-house real-time PCR test exhibited a clinical sensitivity of 98% and a clinical specificity of 100%, indicating its reliability and accuracy for HPV genotyping. The multiplex real-time PCR assay is a cost-effective and efficient tool for HPV screening, detecting multiple genotypes simultaneously. It enhances screening efficiency and accuracy, improving early detection and management of HPV-related diseases.