biostudies-literatureUnknown16Ding SRibonuclease H (RNase H), a critical functional protein in replication and genome stability, is emerging as a crucial therapeutic target for various diseases, including immune disorders. We present a transcription-based hybrid probe, referred to as Hybprobe, and a CRISPR/Cas12a signal amplifier for the rapid, sensitive, and low-cost detection of RNase H activity. In this method, the RNA strand of the Hybprobe is specifically cleaved by RNase H, releasing a single-stranded DNA activator that facilitates recognition and cleavage by the Cas12a/crRNA complex, triggering signal amplification via Cas12a's trans-cleavage activity. The proposed method demonstrates ultra-high sensitivity, capable of detecting RNase H as low as 9.02 × 10^-10 U/μL, making it approximately 1,000 times more sFrontiers in pharmacology1589150https://www.ebi.ac.uk/biostudies/studies/S-EPMC12021801biostudies-literatureUltrasensitive RNase H activity detection using the transcription-based hybrid probe and CRISPR/cas12a signal amplifier.PMC12021801Li XTang ZDing SRen KYang MShi JWei YfalseUltrasensitive RNase H activity detection using the transcription-based hybrid probe and CRISPR/cas12a signal amplifier.Ribonuclease H (RNase H), a critical functional protein in replication and genome stability, is emerging as a crucial therapeutic target for various diseases, including immune disorders. We present a transcription-based hybrid probe, referred to as Hybprobe, and a CRISPR/Cas12a signal amplifier for the rapid, sensitive, and low-cost detection of RNase H activity. In this method, the RNA strand of the Hybprobe is specifically cleaved by RNase H, releasing a single-stranded DNA activator that facilitates recognition and cleavage by the Cas12a/crRNA complex, triggering signal amplification via Cas12a's trans-cleavage activity. The proposed method demonstrates ultra-high sensitivity, capable of detecting RNase H as low as 9.02 × 10^-10 U/μL, making it approximately 1,000 times more s2025-01-01T00:00:00Z20252025-07-09T03:04:52.561Z2025-07-09T03:04:52.561ZS-EPMC120218014029043610.3389/fphar.2025.1589150