MicroRNAs (miRNAs) have been widely investigated as potential biomarkers for early clinical diagnosis of cancer. Developing an miRNA detection platform with high specificity, sensitivity, and exploitability is always necessary. Electrochemiluminescence (ECL) is an electrogenerated chemiluminescence technology that greatly decreases background noise and improves detection sensitivity. The development of a paper‐based ECL biosensor further makes ECL suitable for point‐of‐care detection. Recently, clustered regularly interspaced short palindromic repeats (CRISPR)/Cas13a as high‐fidelity, efficient, and programmable CRISPR RNA (crRNA) guided RNase has brought a next‐generation biosensing technology. However, existing CRISPR/Cas13a based detection often faces a trade‐off between sensitivity and specificity. In this research, a CRISPR/Cas13a powered portable ECL chip (PECL‐CRISPR) is constructed. Wherein target miRNA activates Cas13a to cleave a well‐designed preprimer, and triggers the subsequent exponential amplification and ECL detection. Under optimized conditions, a limit‐of‐detection of 1 × 10 −15 m for miR‐17 is achieved. Through rationally designing the crRNA, the platform can provide single nucleotide resolution to dramatically distinguish miRNA target from its highly homologous family members. Moreover, the introduction of “light‐switch” molecule [Ru(phen) 2 dppz] 2+ allows the platform to avoid tedious electrode modification and washing processes, thereby simplifying the experimental procedure and lower testing cost. Analysis results of miRNA from tumor cells also demonstrate the PECL‐CRISPR platform holds a promising potential for molecular diagnosis.

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The ability to detect low numbers of microbial cells in food and clinical samples is highly valuable but remains a challenge. Here we present a detection system (called ‘APC-Cas’) that can detect very low numbers of a bacterial pathogen without isolation, using a three-stage amplification to generate powerful fluorescence signals. APC-Cas involves a combination of nucleic acid-based allosteric probes and CRISPR-Cas13a components. It can selectively and sensitively quantify Salmonella Enteritidis cells (from 1 to 105 CFU) in various types of samples such as milk, showing similar or higher sensitivity and accuracy compared with conventional real-time PCR. Furthermore, APC-Cas can identify low numbers of S. Enteritidis cells in mouse serum, distinguishing mice with early- and late-stage infection from uninfected mice. Our method may have potential clinical applications for early diagnosis of pathogens.

MicroRNAs (miRNAs) are short noncoding RNAs that post-transcriptionally regulate gene expression. It has been proved that the aberrant expression of miRNAs is related to disease and miRNAs can serve as potential biomarkers for early tumor diagnosis. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas13a is a recently discovered CRISPR-RNA (crRNA) guided RNA manipulation tool. The recognition of target RNA can morphologically activate the robust nonspecific trans ribonuclease activity of Cas13a. This unique property makes Cas13a ideal for nucleic acid detection. Herein, we first exploited CRISPR/LbuCas13a to directly detect miRNAs with high specificity and simplicity. A limit of detection (LOD) as low as 4.5 amol was achieved by this one-step assay within 30 min, and the dynamic range spanned 4 orders of magnitude from 10 amol to 100 fmol. More importantly, single nucleotide variation, even at the end of target miRNA, can be discriminated by rationally programmed crRNA. In addition, the practical application ability of this Cas13a/crRNA-based signal amplification strategy was demonstrated by miRNA quantification in complex biological samples (total small RNA). With excellent reliability, sensitivity, and simple to implement features, this method promises a great potential for early diagnosis of miRNA-related disease. Moreover, the systematic analysis of the crRNA design could provide guidance to further develop Cas13a-based molecular diagnoses.

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