A reagent-based label free electrochemiluminescence biosensor for ultrasensitive quantification of low-abundant chloramphenicol

Today, great attentions have been attracted on chloramphenicol (CAP) to its nonnegligible biotoxicity to human. It is most important and urgent to quantify the low-abundant chloramphenicol by generating simple and efficient analysis approach for the detection of chloramphenicol. Herein, we proposed a novel reagent-based label free electrochemiluminescence (ECL) biosensor based on AuNPs@CoFe-PBA nanomaterials as highly efficient catalysts for luminol/hydrogen peroxide (H2O2) system and target indirectly induced activation of the CRISPR/Cas 12a amplification system for simple and efficient quantification of the low-abundant chloramphenicol. Typically, AuNPs@CoFe-PBA nanomaterials as a novel catalyst for luminol/H2O2 system were enclosed in the X-type DNA hydrogel to greatly inhibit their catalytic performance due to the blocking effect of DNA hydrogel. In presence of target CAP, it induced DNA structure transformation of the aptamer of chloramphenicol to generate a toehold to present catalyzed hairpin assembly (CHA) amplification with the help of two self-designed hairpin structure DNA to perform the hybridizer of the two hairpins, which could activate the CRISPR/Cas 12a system to carry out the DNA cleavage. Its activity breaks the structure of DNA hydrogels and releases AuNPs@CoFe-PBA nanomaterials, which enhances the ECL signals of the luminol/H2O2 systems for simple and sensitive analysis of chloramphenicol. Most importantly, all of the target induced amplifications were performed in the detection solution and the ECL reactions were generated on the bare electrode without any modification to construct a reagent-based label-free analysis system simply with high efficiency for quantification of the low-abundant chloramphenicol. The results showed that there was a good linear relationship between ECL intensity and CAP concentration in range from 10 pg/mL to 1 ug/mL with a detection limit of 7.75 pg/mL, providing great potentials to quantify the low-abundant chloramphenicol and some other biomarkers for early disease diagnosis, food safety testing and environmental analysis.