Development of a novel homogeneous immunoassay using mutant beta-glucuronidase

Βeta-glucuronidase (GUS) catalyzes breakdown of complex carbohydrates, whose activity can be detected quantitatively and sensitively by using fluorogenic and chromogenic substrates. GUS is a tetramer composed of four identical subunits, and assembly of all these subunits is necessary to attain its activity. Based on a previous study, a set of interface mutations (M516K, Y517E) is known to effectively inhibit the assembly and makes it inactive [1]. Usually, the affinity between the two variable region domains (VH and VL) of an antibody recognizing a small molecule is relatively low. However, in the presence of antigen, this affinity becomes higher so that they bind each other more tightly [2]. This gives the idea that a fusion protein system comprising VH and VL of an antibody as the detector each tethered to a mutant GUS subunit (GUSm) as the reporter can be used as a biosensor for small molecules. In this study, we aimed at detecting 4-hydroxy-3-nitrophenylacetyl (NP) and bone Gla protein (BGP) as targets of this novel immunosensor (Fig. 1). Please click Additional Files below to see the full abstract

Application and Safety Evaluation of Integrated TCM-WM in the Treatment of COVID-19

Introduction: With the emergence of more infectious mutant strains and the appearance of new crown sequelae, early precise treatment is particularly important. The traditional Chinese medicine and Western medicine (TCM-WM) treatment plan exhibits unique superiority in the COVID-19 treatment, but the efficacy and safety have not been fully elucidated. Methods: A analysis of the clinical characteristics of 7 COVID-19 patients diagnosed in our hospital, as well as the results of precise individual intervention treatment with TCM-WM, including laboratory examination, and CT changes of imaging. Results: On admission, laboratory results showed that 7 patients' overall Lymphocyte count (LYM#), Percent Lymphocytes (LYM %), and Alanine aminotransferase (ALT) were reduced, but Neutrophil percentage (NEU%) were increased. CT imaging showed that most patients had multiple patchy shadows and ground-glass shadows in both lungs. After TCM-WM treatment, the above pathological states were significantly improved. All patients were followed up and had a good prognosis. Conclusions: Accurate therapy with TCM-WM can quickly and effectively alleviate and treat patients with COVID-19, without severe Long Covid

Characterization of stress degradation products of curcumin and its two derivatives by UPLC–DAD–MS/MS

AbstractCurcumin and its two derivatives of PB-3 and DY-1 were subjected to the forced degradation studies under the conditions of hydrolysis (acidic and alkaline), oxidation, photolysis, and thermal stress recommended by ICH Q1A (R2) by means of UPLC–DAD–MS/MS. Three analytes and their degradation products were separated on a column of ACQUITY UPLC®BEH C18 (100mm×2.1mm, 1.7μm) with an in-line filter prior to the column using acetonitrile and 10mM ammonium acetate buffer (pH adjusted to 3.5) as a mobile phase. Both curcumin and DY-1 showed extensive degradation under alkaline condition and gave rise to two degradation products for curcumin and three for DY-1, respectively, while no degradation product was observed under other tested conditions. PB-3 was found to be unstable in acid and alkaline conditions, two degradation products in acidic hydrolytic condition and one in alkaline condition were obtained, while it was stable in photolytic, oxidative and thermal stress conditions. The degradation products of three analytes were characterized as follows by analyzing the mass fragmentation patterns of curcumin and mass analysis of the degradation products: ferulic acid and vanillin for curcumin, 3-methoxyl-4-[3-(1-tetrahydropyrrolyl)propoxyl] benzoic aldehyde, 2,2-di-(1-phenylmethyl)-3-oxo-5-[3-methoxyl-4-(3-tetrahydropyrolylpropoxyl)] pent-4-enoic acid, and tetrapyrolyl propoxyl methoxyl phenyl prop-2-enoic acid for PB-3, 3-oxo-5-[4-hydroxyl-5-methoxyl-3-(1-morpholinylmethyl)] pent-4-enoic aldehyde, 3-oxo-5-(4-hydroxyl-5-methoxyl-3-(1-morpholinylmethyl)) pent-4-enoic acid and 2-hydroxyl-3-methoxyl-5-(7-(3-methoxyl-4-hydroxylphenyl)-3,5-dioxo-4,4-dimethyl hept-1,6-dienyl) benzoic acid for DY-1, respectively. The degradation pathways of curcumin and its derivatives were presented in addition

Fluorescence quenching-based immunological probe for ticagrelor monitoring

Introduction: Ticagrelor is extensively utilized for the treatment of acute coronary syndromes (ACS), but its platelet aggregation inhibitory effects can potentially result in tissue bleeding, posing a serious risk to patients’ lives.Methods: In this study, we developed highly sensitive full length anti-ticagrelor Quenchbodies (Q-bodies) for fast monitoring of ticagrelor both in solution and serum for the first time. Ticagrelor coupled with N- hydroxysuccinimide (Ticagrelor-NHS) ester was also designed and synthesized for interaction and biological activity detection.Results: Both ATTO-labeled MEDI2452 (2452A) Q-body and TAMRA-labeled IgG 152 (152T) Q-body demonstrated efficient detection of ticagrelor and its active metabolite (TAM). The 2452A Q-body exhibited a broader detection range, while the 152T Q-body displayed a lower limit of detection (LOD). Under physiological conditions (Ticagrelor:TAM, 3:1), the concentration of ticagrelor was further measured, yielding LOD values of 4.65 pg/mL and 2.75 pg/mL for the two Q-bodies, with half-maximal effect concentrations of 8.15 ng/mL and 3.0 ng/mL, respectively.Discussion: Compared with traditional liquid chromatography-mass spectrometry (LC-MS) methods, anti-ticagrelor Q-bodies have higher sensitivity and detection speed. It enabled the completion of analysis within 3 min, facilitating rapid preoperative detection of blood drug concentration in ACS to determine the feasibility of surgery and mitigate the risk of intraoperative and postoperative hemorrhage. The swift detection of ticagrelor holds promise for enhancing individualized drug administration, preventing adverse reactions, and providing preoperative guidance

Vehicle Cardinality Estimation in VANETs by Using RFID Tag Estimator

Nowadays, many vehicles equipped with RFID-enabled chipsets traverse the Electronic Toll Collection (ETC) systems. Here, we present a scheme to estimate the vehicle cardinality with high accuracy and efficiency. A unique RFID tag is attached to a vehicle, so we can identify vehicles through RFID tags. With RFID signal, the location of vehicles can be detected remotely. Our scheme makes vehicle cardinality estimation based on the location distance between the first vehicle and second vehicle. Specifically, it derives the relationship between the distance and number of vehicles. Then, it deduces the optimal parameter settings used in the estimation model under certain requirement. According to the actual estimated traffic flow, we put forward a mechanism to improve the estimation efficiency. Conducting extensive experiments, the presented scheme is proven to be outstanding in two aspects. One is the deviation rate of our model is 50 % of FNEB algorithm, which is the classical scheme. The other is our efficiency is 1.5 times higher than that of FNEB algorithm.Second International Conference, IOV 2015Chengdu, ChinaDecember 19-21, 201