| 1. |
- Meng, Qinglai, et al.
(författare)
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Rapid personalized AMR diagnostics using two-dimensional antibiotic resistance profiling strategy employing a thermometric NDM-1 biosensor
- 2021
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Ingår i: Biosensors and Bioelectronics. - : Elsevier BV. - 0956-5663. ; 193
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Tidskriftsartikel (refereegranskat)abstract
- Antimicrobial resistance (AMR) threatens global public health and modern surgical medicine. Expression of β-lactamase genes is the major mechanism by which pathogens become antibiotic resistant. Pathogens expressing extended spectrum β-lactamases (ESBL) and carbapenemases (CP) are especially difficult to treat and are associated with increased hospitalization and mortality rates. Despite considerable effort, identification of ESBLs and CPs in a clinically relevant timeframe remains challenging. In this study, a two-dimensional AMR profiling assay strategy was developed employing panels of antibiotics (penicillins, cephamycins, oximino-cephalosporins and carbapenems) and β-lactamases inhibitors (avibactam and EDTA). The assay required the development of a novel biosensor that employed New Delhi metallo-β-lactamase-1 (NDM-1) as the sensing element. Functionally probing β-lactamase activity using substrates and inhibitors combinatorically increased the informational content that enabled the development of assays capable of simultaneous, differential identification of multiple β-lactamases expressed in a single bacterial isolate. More specifically, the assay enabled the simultaneous identification of ESBL and CP in mock samples, as well as in an engineered construct which co-expressed these β-lactamases. The NDM-1 biosensor assay was 16 times and 8 times more sensitive than the ESBL Nordmann/Dortet/Poirel (NDP) and Carba Nordmann/Poirel (NP) assays, respectively. In a retrospective study, NDM-1 biosensor assays were able to differentially identify ESBLs, metallo-CPs and serine-CPs β-lactamases in 23 clinical isolates with 100% accuracy. An assay algorithm was developed which accelerated data analytics reducing turnaround to <1 h. The assay strategy integrated with AI-based data analytics has the potential to provide physicians with a comprehensive readout of patient AMR status.
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| 2. |
- Adlerberth, Josefin, et al.
(författare)
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Thermometric analysis of blood metabolites in ICU patients
- 2020
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Ingår i: Journal of Thermal Analysis and Calorimetry. - : Springer Science and Business Media LLC. - 1388-6150 .- 1588-2926. ; 140:2, s. 763-771
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Tidskriftsartikel (refereegranskat)abstract
- Real-time monitoring of patient’s blood metabolites, such as glucose and lactate, could potentially improve surgery and recovery outcomes for patients in surgical and intensive care units. Our enzyme thermometric biosensor which is based on flow injected calorimetric determination of immobilized enzyme reaction is capable of performing continuous, fast, and quantitative analysis of metabolites using whole blood. A key technical advantage the assay affords is the ability to use unpretreated whole blood. In this article, the enzyme thermometric biosensor was used, for the first time, to determine glucose and lactate concentrations in the blood of ICU patients. The linear detection range for glucose was 0.5–30 mM and 0.25–12 mM for lactate, using a 20 μL sample volume. A maximum sampling rate of 15 measurements per hour was achieved using venous blood samples, which corresponds to a 4-min measurement interval. In order to validate the accuracy of the results, a comparative analysis between the thermometric biosensor and the clinically applied instrument (LifeScan’s OneTouch®) which is based on disposable dry chemical reaction was performed using samples from 33 patients. The results showed a good correlation between the two methods for both glucose (r = 0.843, p < 0.0001) and lactate (r = 0.78, p = 0.0105). The ability to monitor metabolite levels and trends on a clinically relevant timescale of 5 min is critical for intensive monitoring of ICP and operative patients.
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| 3. |
- Liu, Shichao, et al.
(författare)
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一种β-内酰胺类抗生素的酶热检测方法
- 2019
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Ingår i: Journal of Shanxi University (Natural Science Edition). ; :2021-02
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Tidskriftsartikel (refereegranskat)abstract
- The penicillinase thermistor biosensor（Penicillinase sensor） was developed for the rapid monitoring of blood penem antibiotics concentration and rapid identification of extraneous penicillinase in milk on site.However, the wide application of the penicillinase thermistor biosensor was limited due to its intrinsic poor activity to hydrolyze cephem and carbapenem antibiotics.The recently identified carbapenemase New Delhi metallo-beta-lactamase 1（NDM-1） is able to hydrolyze all commercially available β-lactam antibiotics in high efficacy.We coupled the NDM-1 and the enzymatic thermistor biosensor to develop a NDM-1 thermistor biosensor（NDM-1 sensor） by the installment of the enzymatic thermistor with an enzyme column loaded with NDM-1 conjugated CPG beads.The NDM-1 sensor shows high response activity to Piperacillin（PIP）,Ceftriaxone（CTRX）, and Meropenem（MEM）, and the response activity of the NDM-1 sensor to these three β-lactam antibiotics are all Zn2+ dependent.Moreover, the response activity of the NDM-1 sensor to Penicillin G（P）, PIP, Cefazolin（CZO）, CTRX, Cefepime（FEP） and MEM all linearly correlated with antibiotic concentration from 31.25 to 1 000 mg/L.Within pH from 6.0 to 8.0, the optimal response activity of the NDM-1 sensor to P,PIP, CZO, CTRX and FEP are found at pH 6.5, while the optimal response activity of the NDM-1 sensor to MEM is found at pH8.0.These data indicate that the featured activity of NDM-1 was well maintained after conjugation on CPG beads, and NDM-1 sensor is capable to quantitate three classes of β-lactam antibiotics including penem, cephem and carbapenem within a wide concentration range.
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| 4. |
- Meng, Qinglai, et al.
(författare)
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Development of a tRNA-Synthetase Microarray for Protein Analysis
- 2004
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Ingår i: Sensors and Materials. - 0914-4935. ; 16:8, s. 401-412
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Tidskriftsartikel (refereegranskat)abstract
- Proteins are composed of 20 different amino acids. In the translation process, each of these 20 amino acids is specifically recognized by their cognate aminoacyl-tRNA synthetase. The fidelity of this recognition system is essential if translation is to function properly. The development of an in vitro system based on this recognition scheme would make a powerful analytical tool with which to analyse translation, as well as providing an additional biomimetic scheme for protein analysis. Aminoacyl-tRNA synthetases microarrays could be applied to protein fingerprinting and sequence analysis. The fabrication of aminoacyl-tRNA synthetase arrays requires the use of advanced protein arraying technology that has only recently become available. In order to demonstrate the feasibility of this scheme, glutamyl-tRNA synthetase (GluRS) was immobilized on the streptavidinbased XNA on GoldTM biochip platform. The streptavidin layer provides a simple, efficient immobilization scheme that reduces nonspecific binding and improves the biocompatibility of the surface. Here, we demonstrate that biotinylated GluRS can be successfully immobilized on XNA on GoldTM. The immobilization efficiency was determined by double labelling GluRS with biotin and the fluorescent label Cy5. The CCD fluorescent microscopy images revealed that the GluRS was efficiently immobilized and evenly distributed over the surface. Control experiments indicate a very low degree of nonspecific binding which is essential if detection of these multicomponent, low-affinity interactions is to be realized. Furthermore, we show that immobilization does not significantly reduce the function of the enzyme. In addition to the specific aims of this study, this technology would provide valuable insights into the biomechanics of translation as well as being a tool for studying tRNA modifications and subclasses. Moreover, the implications for developing coupled transcription and translation systems should not be overlooked. Protein analysis schemes based on this approach would provide an urgently needed compliment to traditional methods. Finally, these arrays might also be useful tools in our efforts to understand the regulatory functions that small RNAs, i.e., iRNA, have been shown to play.
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| 5. |
- Meng, Qinglai, et al.
(författare)
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Rapid Detection of Multiple Classes of β-Lactam Antibiotics in Blood Using an NDM-1 Biosensing Assay
- 2021
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Ingår i: Antibiotics. - : MDPI AG. - 2079-6382. ; 10:9
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Tidskriftsartikel (refereegranskat)abstract
- Currently, assays for rapid therapeutic drug monitoring (TDM) of β-lactam antibiotics in blood, which might be of benefit in optimizing doses for treatment of critically ill patients, remain challenging. Previously, we developed an assay for determining the penicillin-class antibiotics in blood using a thermometric penicillinase biosensor. The assay eliminates sample pretreatment, which makes it possible to perform semicontinuous penicillin determinations in blood. However, penicillinase has a narrow substrate specificity, which makes it unsuitable for detecting other classes of β-lactam antibiotics, such as cephalosporins and carbapenems. In order to assay these classes of clinically useful antibiotics, a novel biosensor was developed using New Delhi metallo-β-lactamase-1 (NDM-1) as the biological recognition layer. NDM-1 has a broad specificity range and is capable of hydrolyzing all classes of β-lactam antibiotics in high efficacy with the exception of monobactams. In this study, we demonstrated that the NDM-1 biosensor was able to quantify multiple classes of β-lactam antibiotics in blood plasma at concentrations ranging from 6.25 mg/L or 12.5 mg/L to 200 mg/L, which covered the therapeutic concentration windows of the tested antibiotics used to treat critically ill patients. The detection of ceftazidime and meropenem was not affected by the presence of the β-lactamase inhibitors avibactam and vaborbactam, respectively. Furthermore, both free and protein-bound β-lactams present in the antibiotic-spiked plasma samples were detected by the NDM-1 biosensor. These results indicated that the NDM-1 biosensor is a promising technique for rapid TDM of total β-lactam antibiotics present in the blood of critically ill patients.
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| 6. |
- Meng, Qinglai, et al.
(författare)
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The NDM-1 biosensor rapidly and accurately detected antibiotic plasma concentrations in Cefuroxime-treated patients
- 2024
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Ingår i: International Journal of Antimicrobial Agents. - 1872-7913. ; 64:2
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Tidskriftsartikel (refereegranskat)abstract
- Therapeutic drug monitoring (TDM) of β-lactam antibiotics in critically ill patients may benefit dose optimization, thus improving therapeutic outcomes. However, rapidly and accurately detecting these antibiotics in blood remains a challenge. Our research group recently developed a thermometric biosensor called the New Delhi metallo-β-lactamase-1 (NDM-1) biosensor, which detected multiple classes of β-lactam antibiotics in spiked plasma samples. This study assesses the NDM-1 biosensor's effectiveness in detecting plasma concentrations of β-lactam antibiotic in treated patients. Seven patients receiving Cefuroxime were studied. Plasma samples collected pre- and post-antibiotic treatment were analyzed using the NDM-1 biosensor and compared with liquid chromatography coupled with ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The biosensor detected plasma samples without dilution, and a brief pre-treatment using a PVDF filter significantly lowered matrix effects, reducing the running time to 5-8 minutes per sample. The assay's linear range for Cefuroxime (6.25 to 200 mg/L) covered target concentrations during the trough phase of pharmacokinetics in critically ill patients. The pharmacokinetic properties of Cefuroxime in treated patients determined by the NDM-1 biosensor and the UPLC-MS/MS were comparable, and the Cefuroxime plasma concentrations measured by the two methods showed a statistically good consistency. These data demonstrate that the NDM-1 biosensor assay is a fast, sensitive, and accurate method for detecting Cefuroximeplasma concentration in treated patients and highlights the NDM-1 biosensor as a promising tool for on-site TDM of β-lactam antibiotics in critically ill patients.
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| 7. |
- Song, Chenchen, et al.
(författare)
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Activity fingerprinting of AMR β-lactamase towards a fast and accurate diagnosis
- 2023
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Ingår i: Frontiers in cellular and infection microbiology. - 2235-2988. ; 13
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Tidskriftsartikel (refereegranskat)abstract
- Antibiotic resistance has become a serious threat to global public health and economic development. Rapid and accurate identification of a patient status for antimicrobial resistance (AMR) are urgently needed in clinical diagnosis. Here we describe the development of an assay method for activity fingerprinting of AMR β-lactamases using panels of 7 β-lactam antibiotics in 35 min. New Deli Metallo β-lactamase-1 (NDM-1) and penicillinase were demonstrated as two different classes of β-lactamases. The panel consisted of three classes of antibiotics, including: penicillins (penicillin G, piperacillin), cephalosporins (cefepime, ceftriaxone, cefazolin) and carbapenems (meropenem and imipenem). The assay employed a scheme combines the catalytic reaction of AMR β-lactamases on antibiotic substrates with a flow-injected thermometric biosensor that allows the direct detection of the heat generated from the enzymatic catalysis, and eliminates the need for custom substrates and multiple detection schemes. In order to differentiate classes of β-lactamases, characterization of the enzyme activity under different catalytic condition, such as, buffer composition, ion strength and pH were investigated. This assay could provide a tool for fast diagnosis of patient AMR status which makes possible for the future accurate treatment with selected antibiotics.
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| 8. |
- Wang, Xi, et al.
(författare)
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氨曲南竞争性抑制NDM-1对β-内酰胺类抗生素的水解
- 2019
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Ingår i: Journal of Shanxi University (Natural Science Edition). ; :2021-01
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Tidskriftsartikel (refereegranskat)abstract
- We investigated the effect of Aztreonam on hydrolysis of β-lactam antibiotics by MBL using New Delhi Metallo-β-lactamase-1(NDM-1) as a model. The results showed that Aztreonam significantly inhibited hydrolysis of Nitrocefin and Meropenem by soluble NDM-1, but also inhibited hydrolysis of Penicillin G by CPG beads immobilized NDM-1(NDM-1 beads). Moreover, in spite of extensive washing for multiple times, the activity to hydrolyze Penicillin G by the Aztreoman pre-treated NDM-1 beads was just partially recovered. These data suggest that Aztreonam can covalently and stably bind on NDM-1, thus efficiently inhibiting hydrolysis of other kinds of β-lactam antibiotics by NDM-1 in a competitive way.
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| 9. |
- Xie, Bin
(creator_code:cre_t)
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氨酰基转运核糖核酸合成酶识别型蛋白芯片的制备方法
- 2004
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Patent (övrigt vetenskapligt/konstnärligt)abstract
- A method of manufacturing a protein chip for identifying aminoacyl-tRNA synthetases, suitable for detecting basic amino acids in proteins. It is characterized in, immoblizing aminoacyl-tRNA synthetases corresponding to 20 basic amino acids in arrays onto the surface of the chip, then fixing 20 kinds of tRNAs labelled by fluorescence onto the corresponding synthetases, then obtaining a chip of aminoacyl-tRNA synthetases. The chip could detect amino acids in proteins. By scanning fluorescence, amino acids in proteins will be determined based on fluorescence of 20 wells.
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| 10. |
- Yao, Xiangyu, et al.
(författare)
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A highly sensitive bead-based flow cytometric competitive binding assay to detect SARS-CoV-2 neutralizing antibody activity
- 2022
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Ingår i: Frontiers in Immunology. - : Frontiers Media SA. - 1664-3224. ; 13
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Tidskriftsartikel (refereegranskat)abstract
- Accurate detection of SARS-CoV-2 neutralizing antibody (nAb) is critical for assessing the immunity levels after virus infection or vaccination. As fast, cost-effective alternatives to viral infection-based assays, competitive binding (CB) assays were developed to quantitate nAb by monitoring the ability of sera to inhibit the binding of viral spike (S) protein to the angiotensin converting enzyme 2 (ACE2) receptor. Herein, we established a bead-based flow cytometric CB assay and tested the detection performance of six combination models, i.e. immobilized ACE2 and soluble Fc-tagged S1 subunit of S protein (iACE2/S1-Fc), immobilized ACE2 and soluble Fc-tagged receptor binding domain (RBD) of S protein (iACE2/RBD-Fc), immobilized S1 and soluble Fc-tagged ACE2 (iS1/ACE2-Fc), immobilized S1 and soluble His-tagged ACE2 (iS1/ACE2-His), immobilized RBD and soluble Fc-tagged ACE2 (iRBD/ACE2-Fc), and immobilized RBD and soluble His-tagged ACE2 (iRBD/ACE2-His). Using SARS-CoV-2 monoclonal antibodies and sera of convalescent COVID-19 patients and vaccinated subjects, the combination models iACE2/RBD-Fc, iACE2/S1-Fc and iS1/ACE2-His were identified to be able to specifically detect SARS-CoV-2 nAb, among which iACE2/RBD-Fc model showed the highest sensitivity, superior to a commercial SARS-CoV-2 surrogate virus neutralization test (sVNT) ELISA kit. Further studies demonstrated that the sensitivity and specificity of CB assays were affected by the tag of ACE2, type of spike and method of measuring binding rate between ACE2 and spike. Moreover, the iACE2/RBD-Fc model showed good performance in detecting kinetic development of nAb against both the prototype SARS-CoV-2 strain and an omicron variant of SARS-CoV-2 in people immunized by an inactivated SARS-CoV-2 vaccine, and the results of iACE2/RBD-Fc model are correlated well with those of live virus-based and pseudovirus-based neutralization tests, demonstrating the potential to be developed into a highly sensitive, specific, versatile and high-throughput method for detecting SARS-CoV-2 nAb in clinical practice.
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