Reductive prodrug and AIE copolymer nanoparticle for monitoring and chemotherapy.

Polymeric micelle systems for drug delivery, monitor and chemotherapy have gained significant attention, and reductive polymeric micelle systems have become particularly attractive due to their controlled release behavior without additional assistance. However, there are challenges in accurately controlling drug and probe release from the nanoparticles and determining the loading content of drug and probe. To address these issues, we have developed a reduction-responsive Pt(IV) prodrug-based polymeric delivery system that can be dynamically monitored using aggregation-induced emission luminogens (AIE) based bioprobes. These polymeric micelle can self-assemble into nanoparticles and release both bio-active Pt(II) drug and bio-probe upon reduction activation. TPE molecules released in the inner endo/lysosomal microenvironment aggregate and fluoresce upon irradiation, thus allowing real-time tracking of drug biodistribution without additional contrast agents. Advantages of this system include position-specific chemical bond cleavage, control of platinum content, and monitoring of drug reduction and biodistribution.

A rapid and sensitive CRISPR-Cas12a for the detection of Fusobacterium nucleatum.

Fusobacterium nucleatum (Fn), as a conditional pathogen, can cause a range of oral and gastrointestinal diseases. However, existing clinical detection methods require expensive equipment and complex procedures, which are inconvenient for large-scale screening in epidemiological research. The purpose of this study was to establish a reliable, rapid, and inexpensive detection method based on CRISPR/Cas12a technology for the detection of Fn. Specific recombinase polymerase amplification (RPA) primer sequences and crRNA sequences were designed based on the nusG gene of Fn. Subsequently, a fluorescence assay and a lateral flow immunoassay were established using the RPA and CRISPR-Cas12a system (RPA-CRISPR-Cas12a). Sensitivity validation revealed a limit of detection of 5 copies/µL. This method could distinguish Fn from other pathogens with excellent specificity. Furthermore, the RPA-CRISPR-Cas12a assay was highly consistent with the classical quantitative real-time PCR method when testing periodontal pocket samples. This makes it a promising method for the detection of Fn and has the potential to play an increasingly important role in infectious disease testing.IMPORTANCEFusobacterium nucleatum (Fn) naturally exists in the microbial communities of the oral and gastrointestinal tracts of healthy individuals and can cause inflammatory diseases in the oral and gastrointestinal tracts. Recent studies have shown that Fn is closely associated with the occurrence and development of gastrointestinal cancer. Therefore, the detection of Fn is very important. Unlike the existing clinical detection methods, this study established a fluorescence-based assay and lateral flow immunoassay based on the RPA and CRISPR-Cas12a system (RPA-CRISPR-Cas12a), which is fast, reliable, and inexpensive and can complete the detection within 30-40 minutes. This makes it a promising method for the detection of Fn and has the potential to play an increasingly important role in infectious disease testing.

Rapid screening for Chlamydia trachomatis infection by detecting α-mannosidase activity in urogenital tract specimens.

BACKGROUND: Chlamydia trachomatis may cause multiple different urogenital tract disorders, but current non-culture assays for rapid screening of C. trachomatis typically use immunochromatography-based methods. We established another new rapid non-culture method for detection of C. trachomatis based on the measurement of α-mannosidase enzymatic activity in urogenital tract specimens. METHOD: To evaluate the performance of this method, α-mannosidase activities of C. trachomatis serotype D strain 、 and 29 standard strains related to clinical urogenital pathogens were investigated. Furthermore, 553 urogenital tract specimens were used for clinical assays via cell culture method and ligase chain reaction method (LCR), adopting an expanded gold standard. RESULTS: Only C. trachomatis was positive for α-mannosidase activity among different types of microbes tested in the research. When prostate fluid specimens, which have some interfering activity, were excluded, the sensitivity and specificity of the enzymatic method were 91.8% (78/85) and 98.3% (409/416), respectively. There were no significant differences (P > 0.05). CONCLUSIONS: These results showed that α-mannosidase activity could be utilised as a screening marker of C. trachomatis infection.

Detection of Trichinella spiralis circulating antigens in serum of experimentally infected mice by an IgY-mAb sandwich ELISA.

In this study, a sandwich enzyme-linked immunosorbent assay (ELISA) based on IgY (egg yolk immunoglobulin) and monoclonal antibody (mAb) against excretory-secretory (ES) antigens of Trichinella spiralis muscle larvae was developed for detection of circulating antigens (CAg) in serum from mice infected with T. spiralis. The IgY-mAb sandwich ELISA involved the use of chicken antibody IgY as a capture antibody and mouse mAb 35B9 as a detecting antibody. This method was able to detect as little as 1 ng/mL of ES antigens added to normal mouse serum. A group of 15 mice was orally inoculated with 500 T. spiralis muscle larvae per animal and the serum samples were daily taken during 1-49 days post-infection (dpi). The level of CAg was detectable as early as 3 dpi in the sera from infected mice, increased gradually, and reached two peaks with detection rate of 40% at 13 dpi and 100% at 24 dpi, respectively. The anti-Trichinella antibodies was first detected in 33.3% of the infected mice at 3 week post-infection (wpi), and reached a peak positive rate of 100% at 5 wpi. Moreover, the infected mice were treated with abendazole at 5 weeks post-infection, and the serum levels of CAg in treated group began to increase rapidly at 2 days post-treatment (dpt) and reached a peak with detection rate of 100% (10/10) at 8 dpt, and then decreased gradually. By 42 dpt, the CAg levels decreased to the undetected level, but the anti- Trichinella antibodies were still detected in 100% of the infected mice. The novel assay appears to be sensitive for detection of antigens of T. spiralis and should be valuable to the early diagnosis and evaluation of the efficacy of chemotherapy in trichinellosis.