A proximity induced strand displacement amplification and CRISPR/Cas12a-based SERS assay for ultrasensitive detection of dengue virus.

Dengue virus (DENV) is a mosquito-borne single-stranded RNA virus that causes dengue fever and can progress to severe disease, posing a significant public-health threat. Rapid and highly sensitive detection is crucial for the early diagnosis of DENV, which, however, suffers from difficult discrimination from similar symptoms of other virus infection, such as Chikungunya or Zika virus, as well as insufficient specificity and time-consuming procedures. Here, we present a surface-enhanced Raman scattering (SERS) biosensor that integrates strand displacement amplification (SDA) with CRISPR/Cas12a for ultrasensitive detection of DENV. In the absence of DENV, SDA and Cas12a stay inactive and the SERS probes retain an aggregation state due to the crosslinking of single-stranded Linker, exhibiting strong SERS signals. In the presence of target sequences, SDA generates abundant single-stranded Trigger-strands, activating Cas12a through crRNA recognition, cleaving the Linker required to enable the crosslinking and leaving the SERS nanoprobes in a monodispersed state, demonstrating weak SERS signals. This method shows exceptional sensitivity for Dengue virus type 1 (DENV-1), with a limit of detection as low as 4.68 fM. It also provides excellent specificity by accurately distinguishing DENV-1 from other pathogens, while maintaining reliable performance. The modular design of this strategy endows the system capability to be adapted for different targets, making it not only a promising tool for the highly sensitive diagnosis of DENV-1, but also a versatile platform for the precise recognition of other pathogens.