Introduction: The optimal clinical management of women diagnosed with low-grade squamous intraepithelial lesions (LSIL) during cervical cancer screening remains unclear. In this prospective cohort study, we compared the clinical performance of two human papillomavirus (HPV) mRNA tests for triage of women with LSIL in Denmark. Material and methods: In a nationwide pathology register, we identified women aged 23-65 years with LSIL during 2008-2012. We included women tested for HPV mRNA with the PreTect HPV Proofer test for five high-risk HPV types (n = 2176) or the Aptima assay for 14 high-risk HPV types (n = 426). Subsequent histological diagnoses of cervical intraepithelial neoplasia grades 2, 3 or cancer (CIN2+) were identified in the register. We calculated the sensitivity and specificity for CIN2+ at 18 and 36 months of follow up, and the cumulative incidence of CIN2+ among women testing positive and negative, overall and by age (23-29, 30-39, 40-65 years). Results: The proportion of women with a positive mRNA test at baseline was higher in women tested with Aptima (66.7%) than in women tested with Proofer (42.8%). After 18 months, Aptima had higher sensitivity for CIN2+ than Proofer (98% [95% CI 94% to 100%] vs 85% [95% CI 82% to 88%]), whereas Proofer showed higher specificity than Aptima (67% [95% CI 64% to 70%]) vs (40% [95% CI 33% to 46%]). Aptima had particularly low specificity in women aged <40 years (23-29: 19% [95% CI 5% to 36%]; 30-39: 10% [95% CI 0% to 33%]). The 36-month cumulative incidence of CIN2+ was higher in Proofer positive (54.3% [95% CI 50.9% to 57.8%]) than in Aptima positive women (37.6% [95% CI 31.2% to 44.8%]). In women with a negative mRNA test, the 36-month cumulative incidences of CIN2+ were 13.1% (95% CI 10.8% to 15.8%) and 5.9% (95% CI 1.7% to 19.0%) for Proofer and Aptima, respectively. Conclusions: In women with LSIL, Aptima had high sensitivity for CIN2+, but low specificity, especially in women aged <40 years. The Proofer test may be useful to limit immediate colposcopy referrals in younger women with LSIL, but given its low sensitivity and negative predictive value, Proofer negative women must be followed with repeat cytology.