Molecular logic gates, as biomolecule-based computational systems, are highly suitable for multitarget detection due to their programmability and modularity. However, existing systems are primarily limited to nucleic acid detection and have not been widely applied to disease-related sensing, particularly for disease antigens. CD33 and CD123 are critical biomarkers for acute myeloid leukemia (AML), yet conventional detection methods rely on expensive equipment and complex procedures, limiting their accessibility and practicality. This study designs a DNA logic gate system integrating nucleic acid aptamers, catalytic hairpin assembly (CHA), and CRISPR-Cas12a, pioneering its use for AML antigen detection. The system comprises three modules: input recognition, signal amplification, and signal transduction. Nucleic acid aptamers specifically identify CD33 and CD123, while CHA enables efficient signal amplification and CRISPR-Cas12a generates a fluorescent output via trans-cleavage activity. The system operates stably at room temperature and implements multiple logic gate models, including YES, OR, AND, NOR, and INHIBIT, enabling the simultaneous detection of CD33 and CD123. Experimental results are visually distinguishable under blue light, and the system requires only standard fluorescence detection instruments. In serum samples, it exhibits excellent selectivity and stability, with a detection limit of 0.5 ng/mL. This study pioneers the application of logic gate technology for disease antigen detection, addressing a critical gap in AML biomarker sensing. Our study indicates that this logic detection platform, characterized by its simplicity in operation, high sensitivity, and versatility in logic functions, holds promise as a potent sensing system for the intelligent multiplex target detection of disease antigens, environmental pollutants, and heavy metals.

The oriental fruit fly Bactrocera dorsalis (Hendel) is considered as a quarantine pest in many countries and regions. Challenges remain in distinguishing this species with morphological similarities, especially in relevant development stages. In recent years, CRISPR/Cas12b genetic diagnostics has seen rapid advancements and offers an efficient tool for the identification of pathogens, viruses, and other genetic targets. Here we developed a new and rapid visual detection assay of B. dorsalis using recombinase polymerase amplification (RPA) and the CRISPR/Cas12b system. The system can detect different developmental stages of B. dorsalis within 30–35 min at 43 ℃ and the results are easily observed by the naked eye based on the color change in the tube during the reaction. The specificity and high sensitivity of this method was demonstrated, allowing for detection from 3.2 pg µL^− 1 of DNA. With crude DNA, this diagnostic system successfully identified B. dorsalis by holding the reaction tubes in the hand. Our study demonstrates that RPA-CRISPR/Cas12b visualization system is effective to detect B. dorsalis rapidly and accurately. This approach can be applied for monitoring and identification of other pests in border and relevant locations, preventing biological invasions and ensuring pest control.

The spotted wing drosophila, Drosophila suzukii (Matsumura), is currently distributed in the main soft-skinned fruits production areas in China and 59 other countries, presenting a significant threat to importing nations. Optimal phytosanitary treatments, including fumigation, irradiation, and cold treatment, have been developed to prevent the international movement of this invasive fly. To determine the most cold-tolerant stage and facilitate the development of the technical schedules requested by the Technical Panel on Phytosanitary Treatment (TPPT), cold treatments of D. suzukii immature stages in ‘Red Globe’ grapes were conducted. Dose–mortality data at 0 °C and 2 °C from repeated trials were subjected to analysis of covariance, linear regression, and probit analysis. Results identified 3 d old pupae as the most cold-tolerant stage, followed by 1 d old pupae, 4 d old larvae, and 6 h old eggs with similar tolerance. The 2 d old larvae were the most sensitive stage. In subsequent confirmatory tests, 3 d old pupae were subjected to cold treatment at 0 °C for 9 and 10 days, and at 2 °C for 10 and 12 days, based on the probit estimation of the probit-9 value. No adult emergence occurred in the confirmatory tests except for one deformed adult from a 2 °C 10 d treatment. Therefore, the recommended treatment schedule requires fruit temperatures below 0.00 °C (or 1.62 °C) for no less than 10 (or 12) continuous days, with treatment efficacy not less than 99.9960% (or 99.9955%) at a 95% confidence level, respectively. These schedules are intended for submission to TPPT for the development of phytosanitary treatment standards.