Csm6-DNAzyme Tandem Assay for One-Pot and Sensitive Analysis of Lead Pollution and Bioaccumulation in Mice

Lead contamination in the environment tends to enter the food chain and further into the human body, causing serious health issues. Herein, we proposed a Csm6-DNAzyme tandem assay (termed cDNAzyme) using CRISPR/Cas III-A Csm6 and GR-5 DNAzyme, enabling one-pot and sensitive detection of lead contamination. We found that Pb²⁺-activated GR-5 DNAzyme produced cleaved substrates that can serve as the activator of Csm6, and the Csm6-DNAzyme tandem improved the sensitivity for detecting Pb²⁺ by 6.1 times compared to the original GR-5 DNAzyme. Due to the high specificity of DNAzyme, the cDNAzyme assay can discriminate Pb²⁺ from other bivalent and trivalent interfering ions and allowed precise detection of Pb²⁺ in water and food samples. Particularly, the assay can achieve one-step, mix-and-read detection of Pb²⁺ at room temperature. We used the cDNAzyme assay to investigate the accumulation of lead in mice, and found that lead accumulated at higher levels in the colon and kidney compared to the liver, and most of the lead was excreted. The cDNAzyme assay is promising to serve as analytical tools for lead-associated environmental and biosafety issues.