Abstract Background HPV-caused cancers contribute to a large disease burden, accounting for 5% of all cancers. It is estimated that over 700,000 people develop HPV-related cancers each year with the majority being cervical cancer. The burden of this disease is disproportionately carried by low-income countries with the highest rates of HPV cervical infections primarily found in sub-Saharan Africa, Latin America, Eastern Europe, and South-East Asia. Many regions in low-resource countries lack adequate access to sensitive point-of-care screening tools, preventing timely diagnosis and treatment. Current low-cost methods, such as Pap smears and visual inspection using acetic acid suffer from low sensitivity, specificity, and reproducibility which limit their effectiveness as a screening modality. Many countries use nucleic acid amplification tests (NAATs) to identify key HPV infections which may progress towards cervical cancer. However, the expenditure for NAAT-capable equipment and reagents are cost-prohibitive for rural and underserved communities. To reduce screening barriers, we developed an isothermal HPV molecular test that can detect all 14 cancer-causing HPV strains using a novel amplification methodology combined with G-quadruplex DNAzyme colorimetric detection. Methods We developed a novel isothermal amplification technique that is related to loop-mediated isothermal amplification (LAMP) but uses modified looped primers to enable exponential amplification at 60°C for 1 hour. The modified primers only require two binding sites compared to 4-6 in LAMP. To detect the amplified products, we used G-quadruplex peroxidase-like DNAzyme probes which reduce ABTS to produce a visually interpreted result. Analytical validation evaluated the method’s limit of detection, specificity, and accuracy. As proof of concept in a relevant matrix, clinical cervical brushings collected ThinPrep Pap media were tested with the new method and compared to an FDA-approved reference qPCR test. Results Repeated experiments demonstrate that the lowest detectable HPV DNA concentration was approximately 1000 genomic copies/µL with no amplification with other pathogens. As proof of concept, the comparison to the reference standard demonstrated high accuracy. Conclusions The application of our assay is intended as a near-patient screening tool with further evaluation by a clinician for confirmation. With an accurate and rapid screening tool, we envision low-resource regions across the world will have the tools to rapidly detect and treat cervical cancers in their early stages.