Objective:To evaluate the cytotoxicity and hepatoprotective potentials of extracts,fractions or isolated compound from the leaves of Feronia limonia(F.limonia).Methods:Qualitative phytochemical analysis of extracts,fr...Objective:To evaluate the cytotoxicity and hepatoprotective potentials of extracts,fractions or isolated compound from the leaves of Feronia limonia(F.limonia).Methods:Qualitative phytochemical analysis of extracts,fractions or compound was performed by means of thin layer chromatography and spectroscopic assays.The%purity of compound was measured by analytical HPLC.Extracts,fractions or compound have been individually evaluated for their cytotoxicity effects(10,20,100,250,500,750 and 1 000 μg/mL).Based on the inhibitory concentration(IC_(50)) obtained from the cell viability assay,graded concentrations of extracts,fractions or isolated compound were assessed(10,20,50,100,200 μg/mL) for its hepatoprotective potential against CCl_4-induced hepatotoxicity by monitoring activity levels of serum glutamatic pyruvatic transaminase(SGPT) and serum glutamic oxaloacetic transaminase(SGOT).Results:Results indicated that the methanol extract of F.limonia was non-toxic and hepatoprotective in nature as compared with the petroleum ether extract.The acetone fraction of methanolic extract also showed similar properties but the subsequent two fractions were cytotoxic.However,the pure compound isolated from the penultimate fraction of methanolic extract was non-toxic and hepatoprotective in nature.Biochemical investigations(SCOT,SCPT) further corroborated these cytological observations.Conclusions:It can be concluded from this study that F.limonia methanol extract,some fractions and pure isolated compound herein exhibit hepatoprotective activity.However,cytotoxicity recorded in the penultimate fraction and investigation of structural details of pure compound warrants further study.展开更多
Recombinant human growth hormone(rhGH)has been widely used for the treatment of disorders associated with GH deficiency and multiple clinical indications[1].Accurate determination of biological activity is essential i...Recombinant human growth hormone(rhGH)has been widely used for the treatment of disorders associated with GH deficiency and multiple clinical indications[1].Accurate determination of biological activity is essential in the development,registration,and quality control of rhGH pharmaceutical products[2].However,the existing in vivo bioassay procedure based on somatropin-induced weight gain in rats is complicated,and the use of a rat cell line-based approach(Nb2-11 bioassay),which measures the production of adenosine triphosphate(ATP)as a direct indicator of cell growth,has a low mechanism of action(MOA)relevance.Therefore,novel rhGH bioassays are still needed.To this end,we developed a reporter gene assay(RGA)based on the GH/insulin-like growth factor-1(IGF-1)axis.展开更多
Enzyme-linked immunosorbent assay (ELISA) is the most widely used method for measuring a single cytokine. Recent developments in cytokine quantification such as multiple arrays measure multiple cytokines simultaneousl...Enzyme-linked immunosorbent assay (ELISA) is the most widely used method for measuring a single cytokine. Recent developments in cytokine quantification such as multiple arrays measure multiple cytokines simultaneously. Although good correlations between ELISA and multiplex methods have been observed, side by side comparisons are limited. In the present study we hypothesized that ELISA and Luminex techniques are comparable in detecting cytokines in culture medium when pulmonary artery smooth muscle cells (PASMC) are exposed to stress. Primary human PASMC were cultured in modular chambers and exposed to 21% FiO2 and peak inspiratory and positive end expiratory pressure of 24 and 8 cmH2O respectively, and 95% FiO2. At 24 hours, culture medium was collected and assayed for interleukin-6 (IL-6) and IL-8 by quantitative ELISA and by Human Cytokine 25-Plex Panel using a Luminex 200 analyzer. A comparative analysis of agreement between our ELISA and Luminex data was detailed for control and stress conditions using the Bland-Altman plot analysis. Each assay resulted in comparable increased (p < 0.001) levels of IL-6 and IL-8 as compared to control in response to oxidative and biophysical stress. The Bland-Altman analysis demonstrated that 95% of the differences between ELISA and Luminex values were within ±1.96 SD from the mean difference indicated by the 95% limits of agreement for the measurements of IL-6 and IL-8. There was no systematic bias as a function of inflammation level. We conclude that in this cell culture model, ELISA and Luminex are comparable in detecting the levels of IL-6 and IL-8 in the culture medium. If measurements of multiple cytokines are demanded and the amount of sample is limited, Luminex multi-analyte profiling technology is accurate and sensitive.展开更多
Several nondestructive assay (NDA) methods to quantify special nuclear materials use calibration curves that are linear in the predictor, either directly or as an intermediate step. The linear response model is also o...Several nondestructive assay (NDA) methods to quantify special nuclear materials use calibration curves that are linear in the predictor, either directly or as an intermediate step. The linear response model is also often used to illustrate the fundamentals of calibration, and is the usual detector behavior assumed when evaluating detection limits. It is therefore important for the NDA community to have a common understanding of how to implement a linear calibration according to the common method of least squares and how to assess uncertainty in inferred nuclear quantities during the prediction stage following calibration. Therefore, this paper illustrates regression, residual diagnostics, effect of estimation errors in estimated variances used for weighted least squares, and variance propagation in a form suitable for implementation. Before the calibration can be used, a transformation of axes is required;this step, along with variance propagation is not currently explained in available NDA standard guidelines. The role of systematic and random uncertainty is illustrated and expands on that given previously for the chosen practical NDA example. A listing of open-source software is provided in the Appendix.展开更多
Anti-bungarotoxin anti-serum,which has the internal image of nicotinicacetylcholine receptor,was used as a tool to measure anti-idiotypic antibodies toantibodies to Iigand of nicotinic acctylcholine receptor in scra f...Anti-bungarotoxin anti-serum,which has the internal image of nicotinicacetylcholine receptor,was used as a tool to measure anti-idiotypic antibodies toantibodies to Iigand of nicotinic acctylcholine receptor in scra from 81 patients withmyasthenia gravis.Enzyme-linked immunosorbcnt assay was adopted.Thc positive ratewas 46.9%(38/81).The specific cross inhibitory test with nicotinic acetylcholinereceptor was positive.Anti-idiotype antibodies to antibodies to ligand of nicotinicacetylcholine receptor in sera of different types of myasthenia gravis patients classified ac-cording to modified Osserman’s standard and myasthenia gravis patients with or withoutthymoma were comparcd in this study and the role of anti-idiotype antibodies toantibodies to Iigand of nicotinic acctylcholinc receptor in the immunity of myasthcniagravis and the possibility of thcrapeutic use of anti-idiotype antibodies arc discussed.展开更多
African swine fever(ASF),caused by the African swine fever virus(ASFV),has brought enormous economic loss and represents a major threat to the global pig industry(Vergne et al.2017).ASFVs are divided into 24 genotypes...African swine fever(ASF),caused by the African swine fever virus(ASFV),has brought enormous economic loss and represents a major threat to the global pig industry(Vergne et al.2017).ASFVs are divided into 24 genotypes based on their B646L gene,with only genotypes Ⅰ and Ⅱ circulating globally(Dixon et al.2019).展开更多
Background: The platelet inhibitory response of clopidogrel is substantially variable among patients, and numerous studies have shown that post-percutaneous intervention, patients with high on-treatment platelet react...Background: The platelet inhibitory response of clopidogrel is substantially variable among patients, and numerous studies have shown that post-percutaneous intervention, patients with high on-treatment platelet reactivity have an increase in risk of major adverse cardiovascular events. No published studies to date have utilized platelet function monitoring assays prior to coronary artery bypass graft (CABG) surgery, but determination of patients’ antiplatelet effects prior to surgery may decrease time to surgery and length of hospital stay. The purpose of the study was to evaluate the clinical outcomes of non-elective CABG patients analyzed by the VerifyNow P2Y12 platelet-function monitoring assay prior to surgery compared to a similar set of patients not analyzed by the VerifyNow P2Y12 assay. Methods: This was a retrospective, single center, cohort study. The primary endpoints of this study were time to surgery and length of hospital stay. Results: From March 2013 to July 2013, 60 patient charts were reviewed and included in this study. 49 patients were analyzed by the VerfiyNow P2Y12 assay, and 16 of these patients underwent non-elective CABG surgery. Eleven patients underwent non-elective CABG surgery and were not analyzed by the VerifyNow P2Y12 assay. There was no difference between groups regarding time to surgery (p = 0.75) or length of stay (p = 0.42). Based on the assay’s P2Y12 reaction unit results, 69% of VerifyNow P2Y12 patients went to surgery sooner than the institution’s recommendations which generated more bleeding events, half of which were considered major bleeds. Conclusions: Utilization of the VerifyNow P2Y12 assay prior to non-elective CABG surgery does not shorten time to surgery or overall length of hospital stay. However, insufficient P2Y12 reaction units prior to surgery may lead to more bleeding events, thus the application of platelet function monitoring assays prior to procedures may be beneficial as a bleeding risk-assessment tool.展开更多
BACKGROUND Current diagnosis of hepatitis C virus(HCV)infection requires two sequential steps:testing for anti-HCV followed by HCV RNA PCR to confirm viremia.We have developed a highly sensitive and specific HCV-antig...BACKGROUND Current diagnosis of hepatitis C virus(HCV)infection requires two sequential steps:testing for anti-HCV followed by HCV RNA PCR to confirm viremia.We have developed a highly sensitive and specific HCV-antigens enzyme immunoassay(HCV-Ags EIA)for one-step diagnosis of viremic HCV infection.AIM To assess the clinical application of the HCV-Ags EIA in one-step diagnosis of viremic HCV infection in human immunodeficiency virus(HIV)-coinfected individuals.METHODS The study blindly tested HCV-Ags EIA for its performance in one-step diagnosing viremic HCV infection in 147 sera:10 without HCV or HIV infection;54 with viremic HCV monoinfection;38 with viremic HCV/HIV coinfection;and 45 with viremic HCV and non-viremic HIV coinfection.RESULTS Upon decoding,it was 100%accordance of HCV-Ags EIA to HCV infection status by HCV RNA PCR test.In five sera with HCV infection,HCV RNA was as low as 50-59 IU/mL,and four out of five tested positive for HCV-Ags EIA.Likewise,it was also 100%accordance of HCV-Ags EIA to HCV infection status by HCV RNA PCR in 83 sera with HCV and HIV coinfection,regardless if HIV infection was active or not.CONCLUSION The modified HCV-Ags EIA has a lower detection limit equivalent to serum HCV RNA levels of approximately 100 IU/mL.It is highly sensitive and specific in the setting of HIV coinfection,regardless of HIV infection status and CD4 count.These data support the clinical application of the HCV-Ags EIA in one-step diagnosis of HCV infection in HIV-infected individuals.展开更多
The aims of this study were to evaluate two methods, qPCR and a chemiluminescent assay (ColiLight II), for rapid detection of E. coli in water, and to examine the survival and persistence of clinical E. coli in drinki...The aims of this study were to evaluate two methods, qPCR and a chemiluminescent assay (ColiLight II), for rapid detection of E. coli in water, and to examine the survival and persistence of clinical E. coli in drinking water and biofilm using qPCR and ColiLight II. qPCR and ColiLight II were compared with a cultivation-based method (MPN), and survival and persistence of four clinical E. coli strains in water and biofilms on stainless steel (SS) and polyethylene (PE) surfaces were studied in a flow-through reactor with non-disinfected drinking water using ColiLight II, qPCR, ATP bioluminescence, and MPN. ColiLight II and qPCR correlated well with MPN. In drinking water, some clinical E. coli strains showed prolonged survival in drinking water flow-through systems, and persisted 3 - 3.4 times longer than the theoretical washout due to incorporation into biofilms. Strain specific attributes can significantly affect detection and persistence of E. coli in drinking water matrices.展开更多
Objective:To compare limiting dilution assay and real-time PCR methods in Leishmania tropica parasite load measurement in vaccinated mice.Methods:BALB/c mice were vaccinated by Leishmania tropica soluble Leishmania an...Objective:To compare limiting dilution assay and real-time PCR methods in Leishmania tropica parasite load measurement in vaccinated mice.Methods:BALB/c mice were vaccinated by Leishmania tropica soluble Leishmania antigen or recombinant Leishmania tropica stressinducible protein-1 with/without adjuvant.After three vaccinations,mice were challenged by Leishmania tropica promastigotes.Two months after challenge,the draining lymph nodes of mice footpad were removed and parasite load was assayed by limiting dilution assay and real-time PCR methods.Limiting dilution assay was done by diluting tissue samples to extinction in a biphasic medium.For real-time PCR,DNA of the lymph nodes was extracted,equal dilutions of each sample were prepared and hot-start real-time PCR was done using appropriate primers.The data of the two methods were compared by appropriate statistical methods.Results:Both methods were able to measure different levels of parasite load in vaccinated/unvaccinated mice.In addition,wherever parasite load of a group was estimated high(or low)by one method,the estimated parasite load by another method was the same,although statistically significant differences were found between some groups.Conclusions:Both methods lead to approximately similar results in terms of differentiating parasite load of the experimental groups.However,due to the lower errors and faster process,the real-time PCR method is preferred.展开更多
Background:Colistin(polymyxin E)is a kind of peptide antibiotic which has been approved in animal production for the purposes of disease prevention,treatment,and growth promotion.However,the wide use of colistin in an...Background:Colistin(polymyxin E)is a kind of peptide antibiotic which has been approved in animal production for the purposes of disease prevention,treatment,and growth promotion.However,the wide use of colistin in animal feed may accelerate the spread of colistin-resistance gene MCR-1 from animal production to human beings,and its residue in animal-origin food may also pose serious health hazards to humans.Thus,it is necessary to develop corresponding analytical methods to monitor the addition of colistin in animal feed and the colistin residue in animal-origin food.Results:A one-step enzyme-linked immunosorbent assay(ELISA)and a lateral flow immunochromatographic assay(LFIA)for colistin were developed based on a newly developed monoclonal antibody.The ELISA showed a 50%inhibition value(IC50)of 9.7 ng/m L with assay time less than 60 min,while the LFIA had a strip reader-based detection limit of 0.87 ng/m L in phosphate buffer with assay time less than 15 min.For reducing the non-specific adsorption of colistin onto sample vial,the components of sample extraction solution were optimized and proved to greatly improve the assay accuracy.The spiked recovery experiment showed that the recoveries of colistin from feed,milk and meat samples were in the range of 77.83%to 113.38%with coefficient of variations less than 13%by ELISA analysis and less than 18%by LFIA analysis,respectively.Furthermore,actual sample analysis indicated that the two immunoassays can produce results consistent with instrumental analysis.Conclusions:The developed assays can be used for rapid qualitative or quantitative detection of colistin in animal feed and food.展开更多
Monocot high expression vector pBARUNP1, harboring rabbit defensin(NP 1) gene and selective bar gene for resistance to the herbicide Basta, were constructed and then transferred into immature embryos of wheat (“Bobw...Monocot high expression vector pBARUNP1, harboring rabbit defensin(NP 1) gene and selective bar gene for resistance to the herbicide Basta, were constructed and then transferred into immature embryos of wheat (“Bobwhite” and “Zhong 60634”)via particle bombardment. Southern and RNA dot blots showed the stable integration and transcription of foreign NP 1 gene in the wheat genome. Furthermore, in vitro microbicidal activity assay indicated the proper translation of defensin. Crude protein extraction of transgenic plants exhibited to some extent cytotoxic to several pathogens including G. saubinetii, B. subtilis, E.coli, and A. tumefaciens.展开更多
In vitro chemosensitivity testing of short term primary glioma cultures derived from brain biopsies is still in the research phase and has not yet found a place in clinical use. The main reasons for this slow progress...In vitro chemosensitivity testing of short term primary glioma cultures derived from brain biopsies is still in the research phase and has not yet found a place in clinical use. The main reasons for this slow progression are the small amounts of tissue available and the lack of a suitably sensitive assay capable of use in the clinical setting. This study examines whether the MTS and ATP cell survival assays, which determine cytotoxicity via colorimetric and luminescence analysis respectively, could potentially fulfill this role. Primary glioma cultures were tested for chemosensitivity using the MTS and ATP assays and were found to be generally sensitive to cisplatin and paclitaxel but relatively resistant to carmustine and etoposide. For both assays, LD50 values lay in the range 2 - 130 μg/ml but in the vast majority of cases, those obtained by the ATP assay were markedly lower those obtained by the MTS assay. Moreover, at cell numbers less than 2000 in the cases of paclitaxel and carmustine and less than 4500 in the case of cisplatin, these drugs were generally indicated as ineffective against the glioma cultures tested by the MTS assay but effective against these cultures by the ATP assay. These data clearly demonstrate that the ATP assay is more sensitive when estimating small cell numbers generated by primary glioma cultures from brain biopsies and more reliably detects higher kill rates by anticancer drugs. This study also supports the feasibility of using the ATP assay for chemosensitivity testing in a clinical setting.展开更多
AIM: To optimize the experimental protocols for a simple, sensitive and accurate bleeding assay.METHODS: Bleeding assay was performed in mice by tail tip amputation, immersing the tail in saline at 37 ℃, continuously...AIM: To optimize the experimental protocols for a simple, sensitive and accurate bleeding assay.METHODS: Bleeding assay was performed in mice by tail tip amputation, immersing the tail in saline at 37 ℃, continuously monitoring bleeding patterns and measuring bleeding volume from changes in the body weight. Sensitivity and extent of variation of bleeding time and bleeding volume were compared in mice treated with the P2 Y receptor inhibitor prasugrel at various doses or in mice deficient of Fc Rγ, a signaling protein of the glycoprotein VI receptor.RESULTS: We described details of the bleeding assay with the aim of standardizing this commonly used assay. The bleeding assay detailed here was simple to operate and permitted continuous monitoring of bleedingpattern and detection of re-bleeding. We also reported a simple and accurate way of quantifying bleeding volume from changes in the body weight, which correlated well with chemical assay of hemoglobin levels(r2 = 0.990, P < 0.0001). We determined by tail bleeding assay the dose-effect relation of the anti-platelet drug prasugrel from 0.015 to 5 mg/kg. Our results showed that the correlation of bleeding time and volume was unsatisfactory and that compared with the bleeding time, bleeding volume was more sensitive in detecting a partial inhibition of platelet's haemostatic activity(P < 0.01). Similarly, in mice with genetic disruption of Fc Rγ as a signaling molecule of P-selectin glycoprotein ligand-1 leading to platelet dysfunction, both increased bleeding volume and repeated bleeding pattern defined the phenotype of the knockout mice better than that of a prolonged bleeding time.CONCLUSION: Determination of bleeding pattern and bleeding volume, in addition to bleeding time, improved the sensitivity and accuracy of this assay, particularly when platelet function is partially inhibited.展开更多
A nanoparticle-based potentiometric immunoassay was designed for the sensitive detection of squamous cell carcinoma antigen(SCCA;cervical carcinoma marker) on a portable pH meter coupling enzyme-labeled hybridization ...A nanoparticle-based potentiometric immunoassay was designed for the sensitive detection of squamous cell carcinoma antigen(SCCA;cervical carcinoma marker) on a portable pH meter coupling enzyme-labeled hybridization chain reaction(HCR) with two alternating hairpin DNA probes for the signal amplification. Initially, a sandwich-type immunoreaction was carried out between anti-SCCA capture antibody-conjugated magnetic bead and detection antibody/initiator strand-coated gold nanoparticle(AuNP). Then, the HCR reaction was readily executed between two glucose oxidase(GOx)-labeled hairpins through the initiator strand to form numerous GOx concatamers on the AuNP via the long nicked double-helix. The concatenated GOx oxidized glucose into gluconic acid and hydrogen peroxide, thus resulting in the pH change of the detection solution on a handheld pH meter. Several labeling protocols including GOx-antibody, GOx-AuNP-antibody and GOx-HCR-AuNP-antibody were investigated for detection of target SCCA, and improved analytical features were obtained with the immune-HCR assay. Under optimum conditions, the immune-HCR assay exhibited good pH responses for the determination of SCCA at a concentration as low as 5.7 pg/mL. Additionally, the immune-HCR assay had good precision and reproducibility, high specificity, and acceptable accuracy for analyzing human serum specimens.展开更多
Among shrimp viral pathogens, white spot syndrome virus(WSSV) and yellow head virus(YHV) are the most lethal agents, causing serious problems for both the whiteleg shrimp, Penaeus(Litopenaeus) vannamei, and the black ...Among shrimp viral pathogens, white spot syndrome virus(WSSV) and yellow head virus(YHV) are the most lethal agents, causing serious problems for both the whiteleg shrimp, Penaeus(Litopenaeus) vannamei, and the black tiger shrimp, Penaeus(Penaeus) monodon. Another important virus that infects P. vannamei is infectious myonecrosis virus(IMNV), which induces the white discoloration of affected muscle. In the cases of taura syndrome virus and Penaeus stylirostris densovirus(Pst DNV; formerly known as infectious hypodermal and hematopoietic necrosis virus), their impacts were greatly diminished after the introduction of tolerant stocks of P. vannamei. Less important viruses are Penaeus monodon densovirus(Pm DNV; formerly called hepatopancreatic parvovirus), and Penaeus monodon nucleopolyhedrovirus(Pemo NPV; previously called monodon baculovirus). For freshwater prawn, Macrobrachium rosenbergii nodavirus and extra small virus are considered important viral pathogens. Monoclonal antibodies(MAbs) specific to the shrimp viruses described above have been generated and used as an alternative tool in various immunoassays such as enzyme-linked immunosorbent assay, dot blotting, Western blotting and immunohistochemistry. Some of these MAbs were further developed into immunochromatographic strip tests for the detection of WSSV, YHV, IMNV and Pemo NPV and into a dual strip test for the simultaneous detection of WSSV/YHV. The strip test has the advantages of speed, as the result can be obtained within 15 min, and simplicity, as laboratory equipment and specialized skills are not required. Therefore, strip tests can be used by shrimp farmers for the pond-side monitoring of viral infection.展开更多
Background:Organ transplantation recipients encounter significant risks from acute or chronic infections that threaten graft survival.BK virus(BKV)and JC virus(JCV)are 2 prominent opportunistic infection viruses,and t...Background:Organ transplantation recipients encounter significant risks from acute or chronic infections that threaten graft survival.BK virus(BKV)and JC virus(JCV)are 2 prominent opportunistic infection viruses,and they may cause polyomavirus-associated nephropathy and graft kidney loss in patients who are in an immunosuppressed state after kidney transplantation.Hence,timely detection and sustained monitoring of the viral load are indispensable.However,the current diagnostic methods remain limited,and the development of new molecular detection technology is extremely urgent.Methods:The sequences and concentrations of clustered regularly interspaced short palindromic repeats(CRISPR)RNA(crRNA),the concentration of Cas13a,and the primers for recombinase polymerase amplification(RPA)were optimized for BKV and JCV detection.Next,a novel microfluidic dual-droplet chip was designed and fabricated,and it was integrated with CRISPR(ddCRISPR)to simultaneously qualitatively detect BKV and JCV.Subsequently,the ddCRISPR assay was verified using clinical samples.Then,a lateral flow strip combined with CRISPR(LFCRISPR)was developed for the detection of BKV and JCV in resource-limited settings.Results:A one-pot RPA-CRISPR reaction system was established and optimized for BKV and JCV detection.ddCRISPR can simultaneously and rapidly detect BKV and JCV with high sensitivity(10 copies/ml for BKV and 1 copy/ml for JCV),and provide absolute quantification,which is suitable for viral load detection and conducive to personalized and precise treatment for organ transplant recipients.LFCRISPR simplified the operational process through a simple visual readout,facilitating virus screening after organ transplantation.Conclusion:These platforms incorporate molecular testing into the transplantation treatment model,thereby reducing costs,prolonging the survival time of the graft,improving the clinical outcomes of postoperative management in kidney transplantation,and enhancing the patients’quality of life.展开更多
AIM: To determine the diagnostic accuracy of a new point-of-care assay detecting anti-deamidated gliadin peptides in celiac disease(CD) patients.METHODS: One-hundred-and-twelve patients(age range: 1.8-79.2 years old) ...AIM: To determine the diagnostic accuracy of a new point-of-care assay detecting anti-deamidated gliadin peptides in celiac disease(CD) patients.METHODS: One-hundred-and-twelve patients(age range: 1.8-79.2 years old) with clinical symptoms suggestive of CD and/or first-degree relatives(FDR) of CD patients(n = 66),and confirmed CD on a gluten-free diet(GFD)(n = 46),were prospectively enrolled in the study at Gastroenterology outpatient clinics for adult patients and from the Gastroenterology Consultation Ward at the Pediatric Department of the University Hospital of Geneva.Written informed consent was obtained from all subjects enrolled.The study received approval from the local ethics committee.The original CD diagnosis had been based on serum-positive IgA anti-tissue transglutaminase enzyme-linked immunosorbent assay(ELISA)(QuantaLite,Inova Diagnostics,San Diego,CA,United States) and on biopsy results.Serum samples from all study participants were tested by the new CD lateral flow immunochromatographic assay(CD-LFIA) device,Simtomax Blood Drop(Augurix SA,BioArk,Monthey,Switzerland) to detect immunoglobulin(Ig)A and IgG antibodies against deamidated gliadin peptides.The diagnostic performance was evaluated using receiver operating characteristic curves with 95%CIs.A cut-off of 2 on the Rann colorimetric scale was used to calculate the device's sensitivity and specificity.RESULTS: CD-LFIA was highly accurate in detecting untreated celiac patients.In the group of patients with CD symptoms and/or FDR,eight new cases of CD were detected by ELISA and biopsy.All of these new cases were also correctly identified by CD-LFIA.The test yielded four false positive and four false negative results.The false positive results were all within the groups with clinical symptoms suggestive of CD and/or FDR,whereas the false negative results were all within the GFD group.The test yeld a sensitivity of 78.9%(95%CI: 54.4-93.9) and specificity of 95.7%(95%CI: 89.4-98.8),and the area under the curve reached 0.893(95%CI: 0.798-0.988).The Kappa coefficient,calculated according to the values obtained by two readers from the same device,was of 0.96(SE: 0.06).When the GFD patients were excluded from the analysis,the area under the curve reached 0.989(95%CI: 0.971-1.000) and the Kappa coefficient,calculated according to the values obtained by two readers from the same device,became 0.96(SE: 0.07).Furthermore,using the Rann scale cut-off of 2 without the GFD patients,sensitivity was 100% and specificity was 93.1%(95%CI: 83.3-98.1).CONCLUSION: The new CD-LFIA rapid screening test shows good diagnostic accuracy,sensitivity and specificity,and may rule out CD in patients with CD-related symptoms.展开更多
Potato late blight caused by Phytophthora infestans is one of the most destructive plant diseases that threaten global food security. Early and effective diagnosis of P. infestans is required before disease management...Potato late blight caused by Phytophthora infestans is one of the most destructive plant diseases that threaten global food security. Early and effective diagnosis of P. infestans is required before disease management decisions are made. Here, we developed a quick protocol to detect P. infestans based on a loop-mediated isothermal amplification(LAMP) assay. The P. infestans specific multiple copy DNA sequences(PiSMC), a transposon-like element, provides an ideal target for molecular detection of this pathogen. We designed highly specific and sensitive primers allowing effective LAMP detection of the pathogen at 64℃ in 70 min. In the validation assay, all 15 P. infestans isolates collected from China, Europe and South America could be positively detected, but results of the other 9 Phytophthora species infecting different plants, fungal and bacterial plant pathogens tested were negative. The detection limit of this assay is 1 pg P. infestans DNA. Moreover, the LAMP-PiSMC assay is able to detect P. infestans from infected leaves, tubers and soil. Taken together, this study reports the development of a specific and sensitive LAMP-PiSMC assay for early diagnosis of potato late blight.展开更多
文摘Objective:To evaluate the cytotoxicity and hepatoprotective potentials of extracts,fractions or isolated compound from the leaves of Feronia limonia(F.limonia).Methods:Qualitative phytochemical analysis of extracts,fractions or compound was performed by means of thin layer chromatography and spectroscopic assays.The%purity of compound was measured by analytical HPLC.Extracts,fractions or compound have been individually evaluated for their cytotoxicity effects(10,20,100,250,500,750 and 1 000 μg/mL).Based on the inhibitory concentration(IC_(50)) obtained from the cell viability assay,graded concentrations of extracts,fractions or isolated compound were assessed(10,20,50,100,200 μg/mL) for its hepatoprotective potential against CCl_4-induced hepatotoxicity by monitoring activity levels of serum glutamatic pyruvatic transaminase(SGPT) and serum glutamic oxaloacetic transaminase(SGOT).Results:Results indicated that the methanol extract of F.limonia was non-toxic and hepatoprotective in nature as compared with the petroleum ether extract.The acetone fraction of methanolic extract also showed similar properties but the subsequent two fractions were cytotoxic.However,the pure compound isolated from the penultimate fraction of methanolic extract was non-toxic and hepatoprotective in nature.Biochemical investigations(SCOT,SCPT) further corroborated these cytological observations.Conclusions:It can be concluded from this study that F.limonia methanol extract,some fractions and pure isolated compound herein exhibit hepatoprotective activity.However,cytotoxicity recorded in the penultimate fraction and investigation of structural details of pure compound warrants further study.
基金supported by the first batch of grants from the State Key Laboratory of Drug Regulatory Science,China(Grant No.:2023SKLDRS0108).
文摘Recombinant human growth hormone(rhGH)has been widely used for the treatment of disorders associated with GH deficiency and multiple clinical indications[1].Accurate determination of biological activity is essential in the development,registration,and quality control of rhGH pharmaceutical products[2].However,the existing in vivo bioassay procedure based on somatropin-induced weight gain in rats is complicated,and the use of a rat cell line-based approach(Nb2-11 bioassay),which measures the production of adenosine triphosphate(ATP)as a direct indicator of cell growth,has a low mechanism of action(MOA)relevance.Therefore,novel rhGH bioassays are still needed.To this end,we developed a reporter gene assay(RGA)based on the GH/insulin-like growth factor-1(IGF-1)axis.
文摘Enzyme-linked immunosorbent assay (ELISA) is the most widely used method for measuring a single cytokine. Recent developments in cytokine quantification such as multiple arrays measure multiple cytokines simultaneously. Although good correlations between ELISA and multiplex methods have been observed, side by side comparisons are limited. In the present study we hypothesized that ELISA and Luminex techniques are comparable in detecting cytokines in culture medium when pulmonary artery smooth muscle cells (PASMC) are exposed to stress. Primary human PASMC were cultured in modular chambers and exposed to 21% FiO2 and peak inspiratory and positive end expiratory pressure of 24 and 8 cmH2O respectively, and 95% FiO2. At 24 hours, culture medium was collected and assayed for interleukin-6 (IL-6) and IL-8 by quantitative ELISA and by Human Cytokine 25-Plex Panel using a Luminex 200 analyzer. A comparative analysis of agreement between our ELISA and Luminex data was detailed for control and stress conditions using the Bland-Altman plot analysis. Each assay resulted in comparable increased (p < 0.001) levels of IL-6 and IL-8 as compared to control in response to oxidative and biophysical stress. The Bland-Altman analysis demonstrated that 95% of the differences between ELISA and Luminex values were within ±1.96 SD from the mean difference indicated by the 95% limits of agreement for the measurements of IL-6 and IL-8. There was no systematic bias as a function of inflammation level. We conclude that in this cell culture model, ELISA and Luminex are comparable in detecting the levels of IL-6 and IL-8 in the culture medium. If measurements of multiple cytokines are demanded and the amount of sample is limited, Luminex multi-analyte profiling technology is accurate and sensitive.
文摘Several nondestructive assay (NDA) methods to quantify special nuclear materials use calibration curves that are linear in the predictor, either directly or as an intermediate step. The linear response model is also often used to illustrate the fundamentals of calibration, and is the usual detector behavior assumed when evaluating detection limits. It is therefore important for the NDA community to have a common understanding of how to implement a linear calibration according to the common method of least squares and how to assess uncertainty in inferred nuclear quantities during the prediction stage following calibration. Therefore, this paper illustrates regression, residual diagnostics, effect of estimation errors in estimated variances used for weighted least squares, and variance propagation in a form suitable for implementation. Before the calibration can be used, a transformation of axes is required;this step, along with variance propagation is not currently explained in available NDA standard guidelines. The role of systematic and random uncertainty is illustrated and expands on that given previously for the chosen practical NDA example. A listing of open-source software is provided in the Appendix.
文摘Anti-bungarotoxin anti-serum,which has the internal image of nicotinicacetylcholine receptor,was used as a tool to measure anti-idiotypic antibodies toantibodies to Iigand of nicotinic acctylcholine receptor in scra from 81 patients withmyasthenia gravis.Enzyme-linked immunosorbcnt assay was adopted.Thc positive ratewas 46.9%(38/81).The specific cross inhibitory test with nicotinic acetylcholinereceptor was positive.Anti-idiotype antibodies to antibodies to ligand of nicotinicacetylcholine receptor in sera of different types of myasthenia gravis patients classified ac-cording to modified Osserman’s standard and myasthenia gravis patients with or withoutthymoma were comparcd in this study and the role of anti-idiotype antibodies toantibodies to Iigand of nicotinic acctylcholinc receptor in the immunity of myasthcniagravis and the possibility of thcrapeutic use of anti-idiotype antibodies arc discussed.
基金supported by the National Natural Science Fund for Distinguished Young Scholars of China(31925036 and 32025034)the Shandong Provincial Key R&D Program,China(2021LZGC001)+3 种基金the Biological Breeding-Major Projects,Institute of Zoology,Chinese Academy of Sciences and Institute of Animal Science,Chinese Academy of Agricultural Sciences(2023ZD04074 and 2023ZD0404604)the Special Project of Longhu Laboratory,China(LHLab_ZD20230012)the National Natural Science Foundation of China(32230100,32330099 and 32201257)the National Key Research and Development Program of China(2020YFA0509503,2022YFF0710703,2021YFA0805902 and 2022XAGG0121,2022YFF1002803).
文摘African swine fever(ASF),caused by the African swine fever virus(ASFV),has brought enormous economic loss and represents a major threat to the global pig industry(Vergne et al.2017).ASFVs are divided into 24 genotypes based on their B646L gene,with only genotypes Ⅰ and Ⅱ circulating globally(Dixon et al.2019).
文摘Background: The platelet inhibitory response of clopidogrel is substantially variable among patients, and numerous studies have shown that post-percutaneous intervention, patients with high on-treatment platelet reactivity have an increase in risk of major adverse cardiovascular events. No published studies to date have utilized platelet function monitoring assays prior to coronary artery bypass graft (CABG) surgery, but determination of patients’ antiplatelet effects prior to surgery may decrease time to surgery and length of hospital stay. The purpose of the study was to evaluate the clinical outcomes of non-elective CABG patients analyzed by the VerifyNow P2Y12 platelet-function monitoring assay prior to surgery compared to a similar set of patients not analyzed by the VerifyNow P2Y12 assay. Methods: This was a retrospective, single center, cohort study. The primary endpoints of this study were time to surgery and length of hospital stay. Results: From March 2013 to July 2013, 60 patient charts were reviewed and included in this study. 49 patients were analyzed by the VerfiyNow P2Y12 assay, and 16 of these patients underwent non-elective CABG surgery. Eleven patients underwent non-elective CABG surgery and were not analyzed by the VerifyNow P2Y12 assay. There was no difference between groups regarding time to surgery (p = 0.75) or length of stay (p = 0.42). Based on the assay’s P2Y12 reaction unit results, 69% of VerifyNow P2Y12 patients went to surgery sooner than the institution’s recommendations which generated more bleeding events, half of which were considered major bleeds. Conclusions: Utilization of the VerifyNow P2Y12 assay prior to non-elective CABG surgery does not shorten time to surgery or overall length of hospital stay. However, insufficient P2Y12 reaction units prior to surgery may lead to more bleeding events, thus the application of platelet function monitoring assays prior to procedures may be beneficial as a bleeding risk-assessment tool.
文摘BACKGROUND Current diagnosis of hepatitis C virus(HCV)infection requires two sequential steps:testing for anti-HCV followed by HCV RNA PCR to confirm viremia.We have developed a highly sensitive and specific HCV-antigens enzyme immunoassay(HCV-Ags EIA)for one-step diagnosis of viremic HCV infection.AIM To assess the clinical application of the HCV-Ags EIA in one-step diagnosis of viremic HCV infection in human immunodeficiency virus(HIV)-coinfected individuals.METHODS The study blindly tested HCV-Ags EIA for its performance in one-step diagnosing viremic HCV infection in 147 sera:10 without HCV or HIV infection;54 with viremic HCV monoinfection;38 with viremic HCV/HIV coinfection;and 45 with viremic HCV and non-viremic HIV coinfection.RESULTS Upon decoding,it was 100%accordance of HCV-Ags EIA to HCV infection status by HCV RNA PCR test.In five sera with HCV infection,HCV RNA was as low as 50-59 IU/mL,and four out of five tested positive for HCV-Ags EIA.Likewise,it was also 100%accordance of HCV-Ags EIA to HCV infection status by HCV RNA PCR in 83 sera with HCV and HIV coinfection,regardless if HIV infection was active or not.CONCLUSION The modified HCV-Ags EIA has a lower detection limit equivalent to serum HCV RNA levels of approximately 100 IU/mL.It is highly sensitive and specific in the setting of HIV coinfection,regardless of HIV infection status and CD4 count.These data support the clinical application of the HCV-Ags EIA in one-step diagnosis of HCV infection in HIV-infected individuals.
文摘The aims of this study were to evaluate two methods, qPCR and a chemiluminescent assay (ColiLight II), for rapid detection of E. coli in water, and to examine the survival and persistence of clinical E. coli in drinking water and biofilm using qPCR and ColiLight II. qPCR and ColiLight II were compared with a cultivation-based method (MPN), and survival and persistence of four clinical E. coli strains in water and biofilms on stainless steel (SS) and polyethylene (PE) surfaces were studied in a flow-through reactor with non-disinfected drinking water using ColiLight II, qPCR, ATP bioluminescence, and MPN. ColiLight II and qPCR correlated well with MPN. In drinking water, some clinical E. coli strains showed prolonged survival in drinking water flow-through systems, and persisted 3 - 3.4 times longer than the theoretical washout due to incorporation into biofilms. Strain specific attributes can significantly affect detection and persistence of E. coli in drinking water matrices.
基金supported by Pasteur Institute of Iran(funding No 754)Kermanshah University of Medical Sciences(funding No 980467).
文摘Objective:To compare limiting dilution assay and real-time PCR methods in Leishmania tropica parasite load measurement in vaccinated mice.Methods:BALB/c mice were vaccinated by Leishmania tropica soluble Leishmania antigen or recombinant Leishmania tropica stressinducible protein-1 with/without adjuvant.After three vaccinations,mice were challenged by Leishmania tropica promastigotes.Two months after challenge,the draining lymph nodes of mice footpad were removed and parasite load was assayed by limiting dilution assay and real-time PCR methods.Limiting dilution assay was done by diluting tissue samples to extinction in a biphasic medium.For real-time PCR,DNA of the lymph nodes was extracted,equal dilutions of each sample were prepared and hot-start real-time PCR was done using appropriate primers.The data of the two methods were compared by appropriate statistical methods.Results:Both methods were able to measure different levels of parasite load in vaccinated/unvaccinated mice.In addition,wherever parasite load of a group was estimated high(or low)by one method,the estimated parasite load by another method was the same,although statistically significant differences were found between some groups.Conclusions:Both methods lead to approximately similar results in terms of differentiating parasite load of the experimental groups.However,due to the lower errors and faster process,the real-time PCR method is preferred.
基金financially supported by Beijing Advanced Innovation Center for Food Nutrition and Human HealthBasic Research Program of Science and Technology(2014FY111000).
文摘Background:Colistin(polymyxin E)is a kind of peptide antibiotic which has been approved in animal production for the purposes of disease prevention,treatment,and growth promotion.However,the wide use of colistin in animal feed may accelerate the spread of colistin-resistance gene MCR-1 from animal production to human beings,and its residue in animal-origin food may also pose serious health hazards to humans.Thus,it is necessary to develop corresponding analytical methods to monitor the addition of colistin in animal feed and the colistin residue in animal-origin food.Results:A one-step enzyme-linked immunosorbent assay(ELISA)and a lateral flow immunochromatographic assay(LFIA)for colistin were developed based on a newly developed monoclonal antibody.The ELISA showed a 50%inhibition value(IC50)of 9.7 ng/m L with assay time less than 60 min,while the LFIA had a strip reader-based detection limit of 0.87 ng/m L in phosphate buffer with assay time less than 15 min.For reducing the non-specific adsorption of colistin onto sample vial,the components of sample extraction solution were optimized and proved to greatly improve the assay accuracy.The spiked recovery experiment showed that the recoveries of colistin from feed,milk and meat samples were in the range of 77.83%to 113.38%with coefficient of variations less than 13%by ELISA analysis and less than 18%by LFIA analysis,respectively.Furthermore,actual sample analysis indicated that the two immunoassays can produce results consistent with instrumental analysis.Conclusions:The developed assays can be used for rapid qualitative or quantitative detection of colistin in animal feed and food.
文摘Monocot high expression vector pBARUNP1, harboring rabbit defensin(NP 1) gene and selective bar gene for resistance to the herbicide Basta, were constructed and then transferred into immature embryos of wheat (“Bobwhite” and “Zhong 60634”)via particle bombardment. Southern and RNA dot blots showed the stable integration and transcription of foreign NP 1 gene in the wheat genome. Furthermore, in vitro microbicidal activity assay indicated the proper translation of defensin. Crude protein extraction of transgenic plants exhibited to some extent cytotoxic to several pathogens including G. saubinetii, B. subtilis, E.coli, and A. tumefaciens.
文摘In vitro chemosensitivity testing of short term primary glioma cultures derived from brain biopsies is still in the research phase and has not yet found a place in clinical use. The main reasons for this slow progression are the small amounts of tissue available and the lack of a suitably sensitive assay capable of use in the clinical setting. This study examines whether the MTS and ATP cell survival assays, which determine cytotoxicity via colorimetric and luminescence analysis respectively, could potentially fulfill this role. Primary glioma cultures were tested for chemosensitivity using the MTS and ATP assays and were found to be generally sensitive to cisplatin and paclitaxel but relatively resistant to carmustine and etoposide. For both assays, LD50 values lay in the range 2 - 130 μg/ml but in the vast majority of cases, those obtained by the ATP assay were markedly lower those obtained by the MTS assay. Moreover, at cell numbers less than 2000 in the cases of paclitaxel and carmustine and less than 4500 in the case of cisplatin, these drugs were generally indicated as ineffective against the glioma cultures tested by the MTS assay but effective against these cultures by the ATP assay. These data clearly demonstrate that the ATP assay is more sensitive when estimating small cell numbers generated by primary glioma cultures from brain biopsies and more reliably detects higher kill rates by anticancer drugs. This study also supports the feasibility of using the ATP assay for chemosensitivity testing in a clinical setting.
基金Supported by Project and Fellowship Grants from the National Health and Medical Research Council of Australia
文摘AIM: To optimize the experimental protocols for a simple, sensitive and accurate bleeding assay.METHODS: Bleeding assay was performed in mice by tail tip amputation, immersing the tail in saline at 37 ℃, continuously monitoring bleeding patterns and measuring bleeding volume from changes in the body weight. Sensitivity and extent of variation of bleeding time and bleeding volume were compared in mice treated with the P2 Y receptor inhibitor prasugrel at various doses or in mice deficient of Fc Rγ, a signaling protein of the glycoprotein VI receptor.RESULTS: We described details of the bleeding assay with the aim of standardizing this commonly used assay. The bleeding assay detailed here was simple to operate and permitted continuous monitoring of bleedingpattern and detection of re-bleeding. We also reported a simple and accurate way of quantifying bleeding volume from changes in the body weight, which correlated well with chemical assay of hemoglobin levels(r2 = 0.990, P < 0.0001). We determined by tail bleeding assay the dose-effect relation of the anti-platelet drug prasugrel from 0.015 to 5 mg/kg. Our results showed that the correlation of bleeding time and volume was unsatisfactory and that compared with the bleeding time, bleeding volume was more sensitive in detecting a partial inhibition of platelet's haemostatic activity(P < 0.01). Similarly, in mice with genetic disruption of Fc Rγ as a signaling molecule of P-selectin glycoprotein ligand-1 leading to platelet dysfunction, both increased bleeding volume and repeated bleeding pattern defined the phenotype of the knockout mice better than that of a prolonged bleeding time.CONCLUSION: Determination of bleeding pattern and bleeding volume, in addition to bleeding time, improved the sensitivity and accuracy of this assay, particularly when platelet function is partially inhibited.
基金financial support from the National Natural Science Foundation of China (No. 21675029)the Health-Education Joint Research Project of Fujian Province (No. WKJ2016-2-15)
文摘A nanoparticle-based potentiometric immunoassay was designed for the sensitive detection of squamous cell carcinoma antigen(SCCA;cervical carcinoma marker) on a portable pH meter coupling enzyme-labeled hybridization chain reaction(HCR) with two alternating hairpin DNA probes for the signal amplification. Initially, a sandwich-type immunoreaction was carried out between anti-SCCA capture antibody-conjugated magnetic bead and detection antibody/initiator strand-coated gold nanoparticle(AuNP). Then, the HCR reaction was readily executed between two glucose oxidase(GOx)-labeled hairpins through the initiator strand to form numerous GOx concatamers on the AuNP via the long nicked double-helix. The concatenated GOx oxidized glucose into gluconic acid and hydrogen peroxide, thus resulting in the pH change of the detection solution on a handheld pH meter. Several labeling protocols including GOx-antibody, GOx-AuNP-antibody and GOx-HCR-AuNP-antibody were investigated for detection of target SCCA, and improved analytical features were obtained with the immune-HCR assay. Under optimum conditions, the immune-HCR assay exhibited good pH responses for the determination of SCCA at a concentration as low as 5.7 pg/mL. Additionally, the immune-HCR assay had good precision and reproducibility, high specificity, and acceptable accuracy for analyzing human serum specimens.
基金Srinakharinwirot University, the Thai National Center for Genetic Engineering and Biotechnology, the Thailand Research Fund, Office of Higher Education Commission, Ministry of Education, Thailand for their funding supports to our research works
文摘Among shrimp viral pathogens, white spot syndrome virus(WSSV) and yellow head virus(YHV) are the most lethal agents, causing serious problems for both the whiteleg shrimp, Penaeus(Litopenaeus) vannamei, and the black tiger shrimp, Penaeus(Penaeus) monodon. Another important virus that infects P. vannamei is infectious myonecrosis virus(IMNV), which induces the white discoloration of affected muscle. In the cases of taura syndrome virus and Penaeus stylirostris densovirus(Pst DNV; formerly known as infectious hypodermal and hematopoietic necrosis virus), their impacts were greatly diminished after the introduction of tolerant stocks of P. vannamei. Less important viruses are Penaeus monodon densovirus(Pm DNV; formerly called hepatopancreatic parvovirus), and Penaeus monodon nucleopolyhedrovirus(Pemo NPV; previously called monodon baculovirus). For freshwater prawn, Macrobrachium rosenbergii nodavirus and extra small virus are considered important viral pathogens. Monoclonal antibodies(MAbs) specific to the shrimp viruses described above have been generated and used as an alternative tool in various immunoassays such as enzyme-linked immunosorbent assay, dot blotting, Western blotting and immunohistochemistry. Some of these MAbs were further developed into immunochromatographic strip tests for the detection of WSSV, YHV, IMNV and Pemo NPV and into a dual strip test for the simultaneous detection of WSSV/YHV. The strip test has the advantages of speed, as the result can be obtained within 15 min, and simplicity, as laboratory equipment and specialized skills are not required. Therefore, strip tests can be used by shrimp farmers for the pond-side monitoring of viral infection.
基金supported by the National Key Research and Development Program(2023YFC2306200)the National Natural Science Foundation of China(82472374)+1 种基金the Cross-Research Fund of Biomedical Engineering of Shanghai Jiaotong University(YG2024LC02)the Clinical Excellence project of Shanghai Key Laboratory of Nucleic Acid Chemistry and Nanomedicine(2024ZY008).
文摘Background:Organ transplantation recipients encounter significant risks from acute or chronic infections that threaten graft survival.BK virus(BKV)and JC virus(JCV)are 2 prominent opportunistic infection viruses,and they may cause polyomavirus-associated nephropathy and graft kidney loss in patients who are in an immunosuppressed state after kidney transplantation.Hence,timely detection and sustained monitoring of the viral load are indispensable.However,the current diagnostic methods remain limited,and the development of new molecular detection technology is extremely urgent.Methods:The sequences and concentrations of clustered regularly interspaced short palindromic repeats(CRISPR)RNA(crRNA),the concentration of Cas13a,and the primers for recombinase polymerase amplification(RPA)were optimized for BKV and JCV detection.Next,a novel microfluidic dual-droplet chip was designed and fabricated,and it was integrated with CRISPR(ddCRISPR)to simultaneously qualitatively detect BKV and JCV.Subsequently,the ddCRISPR assay was verified using clinical samples.Then,a lateral flow strip combined with CRISPR(LFCRISPR)was developed for the detection of BKV and JCV in resource-limited settings.Results:A one-pot RPA-CRISPR reaction system was established and optimized for BKV and JCV detection.ddCRISPR can simultaneously and rapidly detect BKV and JCV with high sensitivity(10 copies/ml for BKV and 1 copy/ml for JCV),and provide absolute quantification,which is suitable for viral load detection and conducive to personalized and precise treatment for organ transplant recipients.LFCRISPR simplified the operational process through a simple visual readout,facilitating virus screening after organ transplantation.Conclusion:These platforms incorporate molecular testing into the transplantation treatment model,thereby reducing costs,prolonging the survival time of the graft,improving the clinical outcomes of postoperative management in kidney transplantation,and enhancing the patients’quality of life.
基金Supported by the Swiss Celiac Association,Association Romande de Coeliakie,No.ME 8309 awarded to Sch ppi MG
文摘AIM: To determine the diagnostic accuracy of a new point-of-care assay detecting anti-deamidated gliadin peptides in celiac disease(CD) patients.METHODS: One-hundred-and-twelve patients(age range: 1.8-79.2 years old) with clinical symptoms suggestive of CD and/or first-degree relatives(FDR) of CD patients(n = 66),and confirmed CD on a gluten-free diet(GFD)(n = 46),were prospectively enrolled in the study at Gastroenterology outpatient clinics for adult patients and from the Gastroenterology Consultation Ward at the Pediatric Department of the University Hospital of Geneva.Written informed consent was obtained from all subjects enrolled.The study received approval from the local ethics committee.The original CD diagnosis had been based on serum-positive IgA anti-tissue transglutaminase enzyme-linked immunosorbent assay(ELISA)(QuantaLite,Inova Diagnostics,San Diego,CA,United States) and on biopsy results.Serum samples from all study participants were tested by the new CD lateral flow immunochromatographic assay(CD-LFIA) device,Simtomax Blood Drop(Augurix SA,BioArk,Monthey,Switzerland) to detect immunoglobulin(Ig)A and IgG antibodies against deamidated gliadin peptides.The diagnostic performance was evaluated using receiver operating characteristic curves with 95%CIs.A cut-off of 2 on the Rann colorimetric scale was used to calculate the device's sensitivity and specificity.RESULTS: CD-LFIA was highly accurate in detecting untreated celiac patients.In the group of patients with CD symptoms and/or FDR,eight new cases of CD were detected by ELISA and biopsy.All of these new cases were also correctly identified by CD-LFIA.The test yielded four false positive and four false negative results.The false positive results were all within the groups with clinical symptoms suggestive of CD and/or FDR,whereas the false negative results were all within the GFD group.The test yeld a sensitivity of 78.9%(95%CI: 54.4-93.9) and specificity of 95.7%(95%CI: 89.4-98.8),and the area under the curve reached 0.893(95%CI: 0.798-0.988).The Kappa coefficient,calculated according to the values obtained by two readers from the same device,was of 0.96(SE: 0.06).When the GFD patients were excluded from the analysis,the area under the curve reached 0.989(95%CI: 0.971-1.000) and the Kappa coefficient,calculated according to the values obtained by two readers from the same device,became 0.96(SE: 0.07).Furthermore,using the Rann scale cut-off of 2 without the GFD patients,sensitivity was 100% and specificity was 93.1%(95%CI: 83.3-98.1).CONCLUSION: The new CD-LFIA rapid screening test shows good diagnostic accuracy,sensitivity and specificity,and may rule out CD in patients with CD-related symptoms.
基金funded by the Fok Ying Tung Education Foundation of China(151025)the National Natural Science Foundation of China(31772144)the Development and Reform Commission of Shandong Province,China for funding the project。
文摘Potato late blight caused by Phytophthora infestans is one of the most destructive plant diseases that threaten global food security. Early and effective diagnosis of P. infestans is required before disease management decisions are made. Here, we developed a quick protocol to detect P. infestans based on a loop-mediated isothermal amplification(LAMP) assay. The P. infestans specific multiple copy DNA sequences(PiSMC), a transposon-like element, provides an ideal target for molecular detection of this pathogen. We designed highly specific and sensitive primers allowing effective LAMP detection of the pathogen at 64℃ in 70 min. In the validation assay, all 15 P. infestans isolates collected from China, Europe and South America could be positively detected, but results of the other 9 Phytophthora species infecting different plants, fungal and bacterial plant pathogens tested were negative. The detection limit of this assay is 1 pg P. infestans DNA. Moreover, the LAMP-PiSMC assay is able to detect P. infestans from infected leaves, tubers and soil. Taken together, this study reports the development of a specific and sensitive LAMP-PiSMC assay for early diagnosis of potato late blight.
