Zhina Wu,Yingtong Wang,Yaqin Zhang,Jingzheng Yi,Yutong Li,Jiasi Wang,Min Hu,Di Wang
Periodontitis is one of the most prevalent oral diseases, and the diagnosis and monitoring of periodontitis rely on clinical and imaging examinations. Matrix metalloproteinase-8 (MMP-8) in saliva is a reliable indicator of periodontitis. The traditional detection method is enzyme-linked immunosorbent assay (ELISA), which requires professional expertise and is not sensitive enough to low-abundance protein detection. In this study, we developed a nanoimmunoassay combined with clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR associated protein (Cas)12a for ultrasensitive detection of MMP-8 in saliva. Antibody-DNA-gold nanoparticles (AuNPs) and trans-cleavage activity of CRISPR/Cas12a were employed for signal amplification. With this protocol, the limit of detection (LoD) for MMP-8 was reduced to the fg/mL, indicating 100-fold higher sensitivity than that of the commercial ELISA kit and its amenability for ultrasensitive detection of trace proteins. The diagnostic sensitivity of the assay was 100 %, and the specificity was 83.3 % based on the testing of 23 clinical saliva samples. The sensitivity and specificity were considerably higher than those for ELISA. Thus, this immuno-CRISPR/Cas12a assay holds promise for both diagnosing and monitoring periodontitis in clinical settings.
Jingqiu Xu,Mingxing Li,Wenting Wang,Zhifang Wu,Chaoyang Wang,Xiaoting Jin,Ling Zhang,Wenxiang Jiang,Baiping Fu
Objectives The study investigated the effects of novel prime-&-rinse mode using MDP (10-methacryloyloxydecyl dihydrogenphosphate) and matrix metalloproteinase (MMPs) inhibitors on dentin microtensile bond strengths (MTBS) of self-etch adhesive, resin-dentin interface degradations, and activity of recombinant human (rh) MMP-8, -9. Materials and methods Eight experimental primers were prepared using 5% and 15% of MDP ethanol-aqueous (1:1) solution in combination with/without MMPs inhibitors (1%benzalkonium chloride (BAC), 1000 μm/mL polyvinylphosphonic acid (PVPA) and 15%proanthocyanidin (PA)). Ninety human mid-coronal dentin surfaces were applied with the experimental primers, water-sprayed and gently air-dried (prime-&-rinse mode), or not (control, self-etch mode). The specimens were bonded with self-etch adhesive (Clearfil S3 Bond) and composite resin (Clearfil Majesty). The resin-bonded specimens were prepared into multiple micro-beams for MTBS tests after 24 h and 1 yr of water storage. The resin-dentin interfaces were analyzed with SEM/TEM. The inhibitory effects of eight primers on rhMMP-8, 9 were determined. The data were analyzed using two-way ANOVA and LSD multiple comparisons tests. Results Compared with control, all the primers used in prime-&-rinse mode could significantly improve long-term dentin MTBS ( P < 0.05), while 5%MDP-BAC, 15%MDP and 15%MDP+MMPs inhibitors could significantly increase the short-term dentin MTBS ( P < 0.05). The SEM/TEM findings revealed that the resin-dentin interfaces were stable over time when the prime-&-rinse mode used. Eight primers possessed the high inhibitory ratio of rh MMP-8, 9. Conclusions The novel prime-&-rinse mode using 5%MDP-BAC, 15%MDP and 15%MDP+MMPs inhibitors could significantly increase the short- and long-term dentin MTBS of self-etch adhesive. This might be a supplement to contemporary dentin bonding strategies.
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