BACKGROUND Influenza A and B virus detection is pivotal in epidemiological surveillance and disease management.Rapid and accurate diagnostic techniques are crucial for timely clinical intervention and outbreak prevent...BACKGROUND Influenza A and B virus detection is pivotal in epidemiological surveillance and disease management.Rapid and accurate diagnostic techniques are crucial for timely clinical intervention and outbreak prevention.Quantum dot-encoded microspheres have been widely used in immunodetection.The integration of quantum dot-encoded microspheres with flow cytometry is a well-established technique that enables rapid analysis.Thus,establishing a multiplex detection method for influenza A and B virus antigens based on flow cytometry quantum dot microspheres will help in disease diagnosis.AIM To establish a codetection method of influenza A and B virus antigens based on flow cytometry quantum dot-encoded microsphere technology,which forms the foundation for the assays of multiple respiratory virus biomarkers.METHODS Different quantum dot-encoded microspheres were used to couple the monoclonal antibodies against influenza A and B.The known influenza A and B antigens were detected both separately and simultaneously on a flow cytometer,and the detection conditions were optimized to establish the influenza A and B antigen codetection method,which was utilized for their detection in clinical samples.The results were compared with the fluorescence quantitative polymerase chain reaction(PCR)method to validate the clinical performance of this method.RESULTS The limits of detection of this method were 26.1 and 10.7 pg/mL for influenza A and B antigens,respectively,which both ranged from 15.6 to 250000 pg/mL.In the clinical sample evaluation,the proposed method well correlated with the fluorescent quantitative PCR method,with positive,negative,and overall compliance rates of 57.4%,100%,and 71.6%,respectively.CONCLUSION A multiplex assay for quantitative detection of influenza A and B virus antigens has been established,which is characterized by high sensitivity,good specificity,and a wide detection range and is promising for clinical applications.展开更多
Background Screening for opisthorchiasis,a parasitic worm infection affecting many millions of people in Southeast Asia,has traditionally relied on faecal egg examination such as the formalin-ethyl acetate concentrati...Background Screening for opisthorchiasis,a parasitic worm infection affecting many millions of people in Southeast Asia,has traditionally relied on faecal egg examination such as the formalin-ethyl acetate concentration technique(FECT)and Kato-Katz method.Although the urinary enzyme-linked immunosorbent assay(ELISA)has been used more recently,we developed a urinary antigen-based rapid diagnostic test(RDT)to simplify diagnosis and as a point-of-care testing(POCT)and field applications for surveillance and control of opisthorchiasis.Methods A urinaryOpisthorchis viverrini(OV)-RDT was developed using immunochromatographic methodology with a specific monoclonal antibody against OV.The diagnostic performance of the urinary OV-RDT was compared to that of quantitative faecal FECT and urinary antigen ELISA(n=493).Cross-reactivities of urinary OV-RDT with other helminthiases coexisted withO.viverrini were determined(n=96).A field trial in the application of urinary OV-RDT was compared with urinary antigen ELISA at baseline screening and assessment of drug treatment outcomes in opisthorchiasis(n=1629).The McNemar chi-square,Kruskal-Wallis and Cohen’s kappa coefficient(κ-value)tests were used for statistical analyses.Results Urinary OV-RDT had sensitivity of 94.2%and specificity of 93.2%,compared to faecal FECT.Urinary OV-RDT had high diagnostic agreement(Kappa=0.842-0.874,P<0.001)and quantitative correlation with urinary antigen ELISA(Kruskal-Wallis tests=316.2,P<0.0001)and faecal FECT(Kruskal-Wallis tests=362.3,P<0.0001).The positive rates by OV-RDT,ELISA and FECT were 48.9%,52.5%and 49.3%,respectively.Cross-reactions of urinary OV-RDT with other helminthiases were few(2%).Field trials of urinary OV-RDT yielded comparable prevalence ofO.viverrini between urinary OV-RDT(53.2%)and urinary antigen ELISA(54.0%).OV screening showed high diagnostic agreement(kappa>0.8,P<0.0001)between urinary OV-RDT and urinary antigen ELISA.The cure rates of opisthorchiasis at 1 month post-praziquantel treatment determined by urinary OV-RDT(86.6%)and urinary antigen ELISA(80.5%)were similar(P>0.05).Conclusions The urinary OV-RDT test has high potential as a new tool for screening and evaluating treatment outcomes in opisthorchiasis.The ease of sample collection and simplicity of urinary OV-RDT may facilitate mass screening,control and elimination of opisthorchiasis,thereby contributing to a reduction in the disease burden in Southeast Asia.展开更多
To develop a national reference panel for severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)antigen detection kit and establish a quality standard.The cultures of SARS-CoV-2 and other pathogens were collected...To develop a national reference panel for severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)antigen detection kit and establish a quality standard.The cultures of SARS-CoV-2 and other pathogens were collected to establish a national reference panel for SARS-CoV-2 antigen detection.The stability and homogeneity of the reference panel were evaluated.Based on World Health Organization(WHO)guidance and nucleic acid quantitative results,a quality standard reference panel was established.Currently,three generations of SARS-CoV-2 antigen national reference materials with batch numbers 370095–202001,370095–202202,and 370095–202203 have been successfully established.These national reference panels comprised 8 positive samples,20 negative samples,1 repetitive sample,and 1 lower detection limit sample.The stability and homogeneity of the reference panel meet the requirements.The quality standards are as follows:the positive and negative coincidence rates are 8/8 and 20/20,respectively.The 10 test results of the medium and low-concentration repetitive reference materials should be positive,and the color rendering should be uniform(or the coefficient of variance should not be higher than 20.0%).The lower detection limit should be at least 5×105 U/mL(equivalent to copies/mL),and higher concentrations above the lower detection limit must be positive.A national reference panel for the SARS-CoV-2 antigen detection kit has been established.As the standard of SARS-CoV-2 antigen reagents,the reference panel has played a crucial role in the pre-marketing quality evaluation and post-marketing quality supervision in China.展开更多
At present,with the development of technology,the detection of cryptococcal antigen(CRAG)plays an increasingly important role in the diagnosis of cryptococcosis.However,the three major CRAG detection technologies,late...At present,with the development of technology,the detection of cryptococcal antigen(CRAG)plays an increasingly important role in the diagnosis of cryptococcosis.However,the three major CRAG detection technologies,latex agglutination test(LA),lateral flow assay(LFA)and Enzyme-linked Immunosorbent Assay,have certain limitations.Although these techniques do not often lead to false-positive results,once this result occurs in a particular group of patients(such as human immunodeficiency virus patients),it might lead to severe consequences.展开更多
Many factors have been identified as having the ability to affect the sensitivity of rapid antigen detection(RAD)tests for severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).This study aimed to identify the i...Many factors have been identified as having the ability to affect the sensitivity of rapid antigen detection(RAD)tests for severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).This study aimed to identify the impact of sample processing on the sensitivity of the RAD tests.We explored the effect of different inactivation methods,viral transport media(VTM)solutions,and sample preservation on the sensitivity of four RAD kits based on two SARS-CoV-2 strains.Compared with non-inactivation,heat inactivation significantly impacted the sensitivity of most RAD kits;however,β-propiolactone inactivation only had a minor effect.Some of the VTM solutions(VTM2,MANTACC)had a significant influence on the sensitivity of the RAD kits,especially for low viral-loads samples.The detection value of RAD kits was slightly decreased,while most of them were still in the detection range with the extension of preservation time and the increase of freeze–thaw cycles.Our results showed that selecting the appropriate inactivation methods and VTM solutions is necessary during reagent development,performance evaluation,and clinical application。展开更多
Antigen detection provides particularly valuable information for medical diagnoses;however,the current detection methods are less sensitive and accurate than nucleic acid analysis.The combination of CRISPR/Cas12a and ...Antigen detection provides particularly valuable information for medical diagnoses;however,the current detection methods are less sensitive and accurate than nucleic acid analysis.The combination of CRISPR/Cas12a and aptamers provides a new detection paradigm,but sensitive sensing and stable amplification in antigen detection remain challenging.Here,we present a PCR-free multiple trigger dsDNA tandem-based signal amplification strategy and a de novo designed dual aptamer synergistic sensing strategy.Integration of these two strategies endowed the CRISPR/Cas12a and aptamer-based method with ultra-sensitive,fast,and stable antigen detection.In a demonstration of this method,the limit of detection was at the single virus level(0.17 fM,approximately two copies/μL)in SARS-CoV-2 antigen nucleocapsid protein analysis of saliva or serum samples.The entire procedure required only 20 min.Given our system’s simplicity and modular setup,we believe that it could be adapted reasonably easily for general applications in CRISPR/Cas12a-aptamer-based detection.展开更多
Middle East respiratory syndrome coronavirus(MERS-CoV)infection in humans has a high mortality of>30%.Dromedaries are the reservoir of MERS-CoV and the main source of human infections.However,MERS-CoV infections in...Middle East respiratory syndrome coronavirus(MERS-CoV)infection in humans has a high mortality of>30%.Dromedaries are the reservoir of MERS-CoV and the main source of human infections.However,MERS-CoV infections in dromedaries are usually subclinical.Rapid diagnosis of MERS-CoV infection in these animals is important in preventing camel-to-human transmission of the virus.The possible cross-reactivity of a previously reported rapid nucleocapsid protein-based antigen detection assay for MERS-CoV was examined with different CoVs,including Tylonycteris bat CoV HKU4,dromedary camel CoV UAE-HKU23,human CoV-229E,human CoV-OC43,severe acute respiratory syndrome CoV-2 and rabbit CoV HKU14,where none of them showed false-positive results.The assay was further validated using quantitative real-time reverse transcription-polymerase chain reaction-confirmed MERS-CoV-positive and MERS-CoV-negative dromedary nasal samples collected in Dubai,the United Arab Emirates,which showed that the rapid antigen detection assay has a specificity of 100%and sensitivity of 91.7%.展开更多
Objective To explore a highly sensitive and highly specific method to detect the serum MG7 antigen(Ag)level for early gastric cancer diagnosis.Methods The serum MG7-Ag level was detected by enzyme-linked immunosorbent...Objective To explore a highly sensitive and highly specific method to detect the serum MG7 antigen(Ag)level for early gastric cancer diagnosis.Methods The serum MG7-Ag level was detected by enzyme-linked immunosorbent assay(ELISA)method in 116 preoperative gastric cancer patients,63 postoperative gastric cancer patients,41 patients with precancerous lesion,37 pa-展开更多
With gene engineering EB virus membrane antigen as the diagnostic antigen, indirect immunofluo-rescence (IF) assay was used to detect IgA antibody against EB virus membrane antigen (MA-IgA) in sera from 202 nasopharyn...With gene engineering EB virus membrane antigen as the diagnostic antigen, indirect immunofluo-rescence (IF) assay was used to detect IgA antibody against EB virus membrane antigen (MA-IgA) in sera from 202 nasopharyngeal carcinoma (NPC) patients and 315 controls (normal and patients with other tumors). MA-IgA antibody was positive in 96.8% of the pretreatment NPC patients with a GMT of 1:36.3. MA-IgA detection by this method was more sensitive than EA-IgA detection by IE. In contrast, patients with tumors other than NPC were negative for MA-IgA antibody. 9.1% of VCA-IgA positive persons were MA-IgA positive with a GMT of less than 1:5. No MA-IgA positive was found in VCA-IgA negatives. The results indicated that this method was relatively specific. In the treatment group, the positive rate and GMT of MA-IgA antibody declined with increase in survival time and the decline was faster than VCA-IgA. When recurrence or distant metastasis developed, similar to VCA-IgA and EA-IgA antibodies, the positive rate and GMT of MA-IgA antibody increased to its pretreatment level. Therefore, MA-IgA detection might be valuable in the early diagnosis and monitor of NPC.展开更多
As one of the most important tumor-associated antigens of colorectal adenocarcinoma, the carcinoembryonic antigen (CEA) threatens human health seriously ali over the globe. Fast electrical and highly sensitive detec...As one of the most important tumor-associated antigens of colorectal adenocarcinoma, the carcinoembryonic antigen (CEA) threatens human health seriously ali over the globe. Fast electrical and highly sensitive detection of the CEA with A1GaN/GaN high electron mobility transistor is demonstrated experimentally. To achieve a low detection limit, the Au-gated sensing area of the sensor is functionalized with a CEA aptamer instead of the corresponding antibody. The proposed aptasensor has successfully detected different concentrations (ranging from 50picogram/milliliter (pg/ml) to 50 nanogram/milliliter (ng/ml)) of CEA and achieved a detection limit as low as 50pg/ml at Vas = 0.5 V. The drain-source current shows a c/ear increase of 11.5μA under this bias.展开更多
The formation of covalent adducts as a result of the interaction of metabolically activated chemicals with host macromolecules is a common critical event in mutagenic, carcinogenic, and immunologic phenomena. Because ...The formation of covalent adducts as a result of the interaction of metabolically activated chemicals with host macromolecules is a common critical event in mutagenic, carcinogenic, and immunologic phenomena. Because of their antigenicity and their immunogenicity, covalent adducts may be detected using sensitive immunochemical techniques. The immunochemical approaches to biomonitoring and molecular dosimetry of DNA damage are particularly attractive because they allow sensitive quantitation of specific DNA adducts present in small samples and do not rely on the use of radiolabeled adducts. Two examples of biomarker immunoassay development are presented: an avidin/biotin-amplified ELISA for the major DNA adduct of the human bladder carcinogen 4-aminobiphenyl (ABP), and a particle concentration fluorescent immunoassay (PCFIA) for the major protein adduct associated with toxicity by the prototype hepatotoxin acetaminophen. The examples illustrate critical steps in the development of biomarker immunoassays which include selection of the relevant adduct, preparation of an appropriate immunogen, immunization, characterization of antisera, and development of application-specific sample processing techniques for biomarker quantitation. Immunochemical procedures may be combined with other analytical techniques to form hybrid systems which take advantage of both the antigenicity and the physical or chemical properties of a biomarker to achieve greater specificity and/or sensitivity. The future usefulness of these new tools of molecular epidemiology will depend on a compound-by-compound validation of methods and critical evaluation of the biologic importance of the particular antigenic biomarker as an indicator of exposure and as an indicator of risk.展开更多
Monoclonal antibodies against colon and pancreatic cancer, CL-2, CL-3, PS-9, PS-10, were used to detect the associated antigens in feces of patients with gastrointestinal carcinoma and non-cancer diseases. Binding inh...Monoclonal antibodies against colon and pancreatic cancer, CL-2, CL-3, PS-9, PS-10, were used to detect the associated antigens in feces of patients with gastrointestinal carcinoma and non-cancer diseases. Binding inhibition test by SABC-ELISA method were performed for the measurement of the antigen level. Results showed that the associated antigen detected in feces of patients with colon cancer were significantly higher than that of non-cancer disease or normal subjects. The positive rates were 61.1% as detected with CL-2; 53.4% with CL-3; 55.0%, PS-9; and 53.3% PS-10 in cancer patients while that in normal subjects were 7%; 9%; 8%; and 8% respectively. When 'cocktail' of CL-2, PS-9 and PS-10 were used, the positive rates were 92.5% in colon cancer and 14% in normal subjects. In seven out of the sixty patients with colon cancer studied who were graded as Dukes A, the results were all positive. The results seem superior to the serologic detection and may provide a promising new approach in the early diagnosis of colon cancer.展开更多
During the late incubation period or initial phase of dengue virus infection,laboratory confirmation is through viral isolation in cell culture and/or molecular investigations, or immunofluorescence,or immunohistochem...During the late incubation period or initial phase of dengue virus infection,laboratory confirmation is through viral isolation in cell culture and/or molecular investigations, or immunofluorescence,or immunohistochemistry[1].The dengue virus non-structural antigen NSl that would develop before the appearance of dengue IgM and/or IgG is emerging as a suitable option for dengue diagnosis[2].Platelet therapy is a standard clinical practice for dengue patients with severe thrombocytopenia[3].However,during introductory screening,platelet count is not being done in many cases. This results in delays of platelet therapy. In the course of the current(2010) spurt of dengue in New Delhi[4],simultaneous screening for NSl,IgM and IgG and platelet enumeration has been introduced at the展开更多
Background: Children with seasonal influenza infection cause a significant burden of disease each year in the pediatric clinic. Influenza A and B viruses are the major types responsible for illness. A better understan...Background: Children with seasonal influenza infection cause a significant burden of disease each year in the pediatric clinic. Influenza A and B viruses are the major types responsible for illness. A better understanding of the periodicity facilitates the prevention and control of influenza in children. Objective: This study aims to analyze the epidemiological patterns and subtype characterization of influenza viruses among children in Shenzhen, China. Methods: Influenza samples were collected by nasopharyngeal swabs from influenza like illness patients in Shenzhen Children’s Hospital from January 2016 to December 2018. The positive cases and influenza subtypes were determined by gold labeled antigen detection and reverse transcriptase polymerase chain reaction. The influenza periodicity and age, subtype distribution as well as the association between climate parameters and different influenza subtypes were analyzed by SPSS 22.0. Results: The influenza positive rate during 2016-2018 was 21.0%, with a highest positive rate in the year 2018. The positive rate varied by month, season, and year describing a sequence of peaks presenting primarily in all year including spring, summer and winter. The characteristics of influenza peak were different in each year, with a spring peak in 2016 and a summer plus a winter-spring peaks in 2017 and 2018. In addition, influenza B exhibited a winter-spring seasonal pattern while influenza A displayed a more variable seasonality, highlighting influenza B rather than influenza A which had a negative association with climate parameters. Influenza-positive cases were older than influenza-negative cases (P P Conclusion: Influenza activity in children from Shenzhen typically displays both winter-spring and summer peaks. Influenza A epidemic occurred separately or co-circulated with influenza B, with a winter-spring pattern for influenza B and a much more variable seasonality for influenza A. Influenza B had a negative association with climate parameters. In addition, hospitalization with influenza often occurs in younger individuals infected with influenza A.展开更多
Objective To establish a highly sensitive serologic method for detecting the circulating antigen of Schistosoma japonicum for the diagnosis of this disease and the evaluation of drug therapy effect.Methods A set of ...Objective To establish a highly sensitive serologic method for detecting the circulating antigen of Schistosoma japonicum for the diagnosis of this disease and the evaluation of drug therapy effect.Methods A set of complex support made of poly vinyl chloride (PVC) film slide, adsorbed with specific antibody, was used to test serum samples collected from cases with schistosomiasis japonica, by adding specific enzyme conjugate and substrate tetra methyl benzidine (TMB) with 3% H 2O 2 in 10∶0.1, developed or prepared in our laboratory. Tests were carried out with the set on cases with malaria, paragonimiasis, clonorchiasis, visceral leishmaniasis and cysticercosis as well as normal individuals as control. Serum sample of 50 μl was used in each test and reacted with reagents at room temperature for 30 minutes. The color of positive reaction was blue while negative reaction was colorless.Results Positive rate among cases with schistosomiasis japonica was 100% (45/45) in acute stage, 94.8% (530/559) in early chronic stage and 52.4% (66/126) in late stage. False positive reaction was found neither among all normal individuals (0/513), nor among cases of 155 with malaria, 120 with clonorchiasis, 24 with visceral leishmaniasis and 110 with cysticercosis, except one among 29 cases with parasgonimiasis (1/29).Conclusions The rapid one step enzyme Immunoassay (EIA) to detect circulating schistosome antigen (CSA) has been well established in our laboratory with high sensitivity and good specificity as well as reproducibility. The reaction result can be read easily even in field conditions without power supply. Moreover, the assay is time saving, simple to handle and suitable for the diagnosis of shistosomiasis japonica and evaluation of drug therapeutic effect in practice in the control program of the disease.展开更多
At present,many researchers focused on the point-of-care testing(POCT),a method of disease markers detection without large-scale instruments and specialized persons.However,most POCT diagnostic methods were suffered...At present,many researchers focused on the point-of-care testing(POCT),a method of disease markers detection without large-scale instruments and specialized persons.However,most POCT diagnostic methods were suffered from poor detection sensitivity or inefficiency in quantitative detection.Herein,we developed a newly QD-immune fluorescence test strips(QD-IFTS) based on quantum dots(QDs) as the fluorescence nanocarrier to prepare the immune fluorescence probes in the classical immunochromatography detection system for sensing carcino-embryonic antigen(CEA),a kind of glycoprotein produced by intestinal tissue and a broad spectrum of tumor marker for cancer diagnosis.And we designed a homemade strips fluorescence reader for detection of fluorescence intensity of QDs on the QD-IFTS.Under the optimized reaction conditions,chromatographic time of the newly QD-IFTS was only25 min,sample volume of the newly QD-IFTS was only 40 m L and the LOD of the newly QD-IFTS was 0.72 ng/m L.In addition,the efficiency and robustness of the newly QD-IFTS were confirmed by successfully application in 300 clinical serum samples,and the results revealed great potential in clinical POCT of other biomarkers.展开更多
Early detection of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)infection is an efficient way to prevent the spread of coronavirus disease 2019(COVID-19).Detecting SARS-CoV-2 antigen can be rapid and con...Early detection of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)infection is an efficient way to prevent the spread of coronavirus disease 2019(COVID-19).Detecting SARS-CoV-2 antigen can be rapid and convenient,but it is still challenging to develop highly sensitive methods for effective diagnosis.Herein,a lateral flow assay(LFA)based on fluorescent nanoparticles emitting in the second near-infrared(NIR-II)window is developed for sensitive detection of SARS-CoV-2 antigen.Benefiting from the NIR-II fluorescence with high penetration and low autofluorescence,such NIR-II based LFA allows enhanced signal-to-background ratio,and the limit of detection is down to 0.01 ng·mL^(−1)of SARS-CoV-2 antigen.In the clinical swab sample tests,the NIR-II LFA outperforms the colloidal gold LFA with higher overall percent agreement with the polymerase chain reaction test.The clinical samples with low antigen concentrations(~0.015–~0.068 ng·mL^(−1))can be successfully detected by the NIR-II LFA,but fail for the colloidal gold LFA.The NIR-II LFA can provide a promising platform for highly sensitive,rapid,and cost-effective method for early diagnosis and mass screening of SARS-CoV-2 infection.展开更多
基金Shenzhen Guangming District Soft Science Research Project,No.2021R01097。
文摘BACKGROUND Influenza A and B virus detection is pivotal in epidemiological surveillance and disease management.Rapid and accurate diagnostic techniques are crucial for timely clinical intervention and outbreak prevention.Quantum dot-encoded microspheres have been widely used in immunodetection.The integration of quantum dot-encoded microspheres with flow cytometry is a well-established technique that enables rapid analysis.Thus,establishing a multiplex detection method for influenza A and B virus antigens based on flow cytometry quantum dot microspheres will help in disease diagnosis.AIM To establish a codetection method of influenza A and B virus antigens based on flow cytometry quantum dot-encoded microsphere technology,which forms the foundation for the assays of multiple respiratory virus biomarkers.METHODS Different quantum dot-encoded microspheres were used to couple the monoclonal antibodies against influenza A and B.The known influenza A and B antigens were detected both separately and simultaneously on a flow cytometer,and the detection conditions were optimized to establish the influenza A and B antigen codetection method,which was utilized for their detection in clinical samples.The results were compared with the fluorescence quantitative polymerase chain reaction(PCR)method to validate the clinical performance of this method.RESULTS The limits of detection of this method were 26.1 and 10.7 pg/mL for influenza A and B antigens,respectively,which both ranged from 15.6 to 250000 pg/mL.In the clinical sample evaluation,the proposed method well correlated with the fluorescent quantitative PCR method,with positive,negative,and overall compliance rates of 57.4%,100%,and 71.6%,respectively.CONCLUSION A multiplex assay for quantitative detection of influenza A and B virus antigens has been established,which is characterized by high sensitivity,good specificity,and a wide detection range and is promising for clinical applications.
基金supported by the Cholangiocarcinoma Research Institute,Khon Kaen University,Thailand.This project is funded by the National Science Research Council of Thailand(NRCT)the Post-Doctoral Training Program from Khon Kaen University,Thailand(PD2565-02-01)supported by the Thailand Centre of Excellence for Life Sciences(TCELS)SDT-R was funded by the Wellcome Trust ISSF grant at Imperial College London.
文摘Background Screening for opisthorchiasis,a parasitic worm infection affecting many millions of people in Southeast Asia,has traditionally relied on faecal egg examination such as the formalin-ethyl acetate concentration technique(FECT)and Kato-Katz method.Although the urinary enzyme-linked immunosorbent assay(ELISA)has been used more recently,we developed a urinary antigen-based rapid diagnostic test(RDT)to simplify diagnosis and as a point-of-care testing(POCT)and field applications for surveillance and control of opisthorchiasis.Methods A urinaryOpisthorchis viverrini(OV)-RDT was developed using immunochromatographic methodology with a specific monoclonal antibody against OV.The diagnostic performance of the urinary OV-RDT was compared to that of quantitative faecal FECT and urinary antigen ELISA(n=493).Cross-reactivities of urinary OV-RDT with other helminthiases coexisted withO.viverrini were determined(n=96).A field trial in the application of urinary OV-RDT was compared with urinary antigen ELISA at baseline screening and assessment of drug treatment outcomes in opisthorchiasis(n=1629).The McNemar chi-square,Kruskal-Wallis and Cohen’s kappa coefficient(κ-value)tests were used for statistical analyses.Results Urinary OV-RDT had sensitivity of 94.2%and specificity of 93.2%,compared to faecal FECT.Urinary OV-RDT had high diagnostic agreement(Kappa=0.842-0.874,P<0.001)and quantitative correlation with urinary antigen ELISA(Kruskal-Wallis tests=316.2,P<0.0001)and faecal FECT(Kruskal-Wallis tests=362.3,P<0.0001).The positive rates by OV-RDT,ELISA and FECT were 48.9%,52.5%and 49.3%,respectively.Cross-reactions of urinary OV-RDT with other helminthiases were few(2%).Field trials of urinary OV-RDT yielded comparable prevalence ofO.viverrini between urinary OV-RDT(53.2%)and urinary antigen ELISA(54.0%).OV screening showed high diagnostic agreement(kappa>0.8,P<0.0001)between urinary OV-RDT and urinary antigen ELISA.The cure rates of opisthorchiasis at 1 month post-praziquantel treatment determined by urinary OV-RDT(86.6%)and urinary antigen ELISA(80.5%)were similar(P>0.05).Conclusions The urinary OV-RDT test has high potential as a new tool for screening and evaluating treatment outcomes in opisthorchiasis.The ease of sample collection and simplicity of urinary OV-RDT may facilitate mass screening,control and elimination of opisthorchiasis,thereby contributing to a reduction in the disease burden in Southeast Asia.
基金supported by the National Key Research and Development Program of China (2021YFC2400904)the National Science and Technology Major Project of China (2018ZX10102001).
文摘To develop a national reference panel for severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)antigen detection kit and establish a quality standard.The cultures of SARS-CoV-2 and other pathogens were collected to establish a national reference panel for SARS-CoV-2 antigen detection.The stability and homogeneity of the reference panel were evaluated.Based on World Health Organization(WHO)guidance and nucleic acid quantitative results,a quality standard reference panel was established.Currently,three generations of SARS-CoV-2 antigen national reference materials with batch numbers 370095–202001,370095–202202,and 370095–202203 have been successfully established.These national reference panels comprised 8 positive samples,20 negative samples,1 repetitive sample,and 1 lower detection limit sample.The stability and homogeneity of the reference panel meet the requirements.The quality standards are as follows:the positive and negative coincidence rates are 8/8 and 20/20,respectively.The 10 test results of the medium and low-concentration repetitive reference materials should be positive,and the color rendering should be uniform(or the coefficient of variance should not be higher than 20.0%).The lower detection limit should be at least 5×105 U/mL(equivalent to copies/mL),and higher concentrations above the lower detection limit must be positive.A national reference panel for the SARS-CoV-2 antigen detection kit has been established.As the standard of SARS-CoV-2 antigen reagents,the reference panel has played a crucial role in the pre-marketing quality evaluation and post-marketing quality supervision in China.
基金Supported by the Key Discipline of Jiaxing Respiratory Medicine Construction Project,No.2019-zc-04.
文摘At present,with the development of technology,the detection of cryptococcal antigen(CRAG)plays an increasingly important role in the diagnosis of cryptococcosis.However,the three major CRAG detection technologies,latex agglutination test(LA),lateral flow assay(LFA)and Enzyme-linked Immunosorbent Assay,have certain limitations.Although these techniques do not often lead to false-positive results,once this result occurs in a particular group of patients(such as human immunodeficiency virus patients),it might lead to severe consequences.
基金supported by China's National Science and Technology Major Project(2018ZX10102001)the Non-profit Central Research Institute Fund of Chinese Academy of Medical Sciences(2019PT3100292020PT310004).
文摘Many factors have been identified as having the ability to affect the sensitivity of rapid antigen detection(RAD)tests for severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).This study aimed to identify the impact of sample processing on the sensitivity of the RAD tests.We explored the effect of different inactivation methods,viral transport media(VTM)solutions,and sample preservation on the sensitivity of four RAD kits based on two SARS-CoV-2 strains.Compared with non-inactivation,heat inactivation significantly impacted the sensitivity of most RAD kits;however,β-propiolactone inactivation only had a minor effect.Some of the VTM solutions(VTM2,MANTACC)had a significant influence on the sensitivity of the RAD kits,especially for low viral-loads samples.The detection value of RAD kits was slightly decreased,while most of them were still in the detection range with the extension of preservation time and the increase of freeze–thaw cycles.Our results showed that selecting the appropriate inactivation methods and VTM solutions is necessary during reagent development,performance evaluation,and clinical application。
基金This work was supported by the National Key R&D program of China(2020YFA0907800)the National Natural Science Foundation of China(31922002,31720103901,31772242 and 31870040),the 111 Project(B18022)+1 种基金the Fundamental Research Funds for the Central Universities[22221818014]the Youth Innovation Promotion Association CAS(Y202027)to W.W and the Open Project Funding of the State Key Laboratory of Bioreactor Engineering.
文摘Antigen detection provides particularly valuable information for medical diagnoses;however,the current detection methods are less sensitive and accurate than nucleic acid analysis.The combination of CRISPR/Cas12a and aptamers provides a new detection paradigm,but sensitive sensing and stable amplification in antigen detection remain challenging.Here,we present a PCR-free multiple trigger dsDNA tandem-based signal amplification strategy and a de novo designed dual aptamer synergistic sensing strategy.Integration of these two strategies endowed the CRISPR/Cas12a and aptamer-based method with ultra-sensitive,fast,and stable antigen detection.In a demonstration of this method,the limit of detection was at the single virus level(0.17 fM,approximately two copies/μL)in SARS-CoV-2 antigen nucleocapsid protein analysis of saliva or serum samples.The entire procedure required only 20 min.Given our system’s simplicity and modular setup,we believe that it could be adapted reasonably easily for general applications in CRISPR/Cas12a-aptamer-based detection.
基金supported by the Health and Medical Research Fund-Commissioned Research on Control of Infectious Diseases(Phase IV,CID-HKU6)a donation of TE Health Consultant Company Limitedthe framework of the Higher Education Sprout Project by the Ministry of Education(MOE-111-S-023-A)in Taiwan.
文摘Middle East respiratory syndrome coronavirus(MERS-CoV)infection in humans has a high mortality of>30%.Dromedaries are the reservoir of MERS-CoV and the main source of human infections.However,MERS-CoV infections in dromedaries are usually subclinical.Rapid diagnosis of MERS-CoV infection in these animals is important in preventing camel-to-human transmission of the virus.The possible cross-reactivity of a previously reported rapid nucleocapsid protein-based antigen detection assay for MERS-CoV was examined with different CoVs,including Tylonycteris bat CoV HKU4,dromedary camel CoV UAE-HKU23,human CoV-229E,human CoV-OC43,severe acute respiratory syndrome CoV-2 and rabbit CoV HKU14,where none of them showed false-positive results.The assay was further validated using quantitative real-time reverse transcription-polymerase chain reaction-confirmed MERS-CoV-positive and MERS-CoV-negative dromedary nasal samples collected in Dubai,the United Arab Emirates,which showed that the rapid antigen detection assay has a specificity of 100%and sensitivity of 91.7%.
文摘Objective To explore a highly sensitive and highly specific method to detect the serum MG7 antigen(Ag)level for early gastric cancer diagnosis.Methods The serum MG7-Ag level was detected by enzyme-linked immunosorbent assay(ELISA)method in 116 preoperative gastric cancer patients,63 postoperative gastric cancer patients,41 patients with precancerous lesion,37 pa-
文摘With gene engineering EB virus membrane antigen as the diagnostic antigen, indirect immunofluo-rescence (IF) assay was used to detect IgA antibody against EB virus membrane antigen (MA-IgA) in sera from 202 nasopharyngeal carcinoma (NPC) patients and 315 controls (normal and patients with other tumors). MA-IgA antibody was positive in 96.8% of the pretreatment NPC patients with a GMT of 1:36.3. MA-IgA detection by this method was more sensitive than EA-IgA detection by IE. In contrast, patients with tumors other than NPC were negative for MA-IgA antibody. 9.1% of VCA-IgA positive persons were MA-IgA positive with a GMT of less than 1:5. No MA-IgA positive was found in VCA-IgA negatives. The results indicated that this method was relatively specific. In the treatment group, the positive rate and GMT of MA-IgA antibody declined with increase in survival time and the decline was faster than VCA-IgA. When recurrence or distant metastasis developed, similar to VCA-IgA and EA-IgA antibodies, the positive rate and GMT of MA-IgA antibody increased to its pretreatment level. Therefore, MA-IgA detection might be valuable in the early diagnosis and monitor of NPC.
基金Supported by the National Key Research and Development Program of China under Grant Nos 2016YFB0400104 and 2016YFB0400301the National Natural Science Foundation of China under Grant No 61334002the National Science and Technology Major Project
文摘As one of the most important tumor-associated antigens of colorectal adenocarcinoma, the carcinoembryonic antigen (CEA) threatens human health seriously ali over the globe. Fast electrical and highly sensitive detection of the CEA with A1GaN/GaN high electron mobility transistor is demonstrated experimentally. To achieve a low detection limit, the Au-gated sensing area of the sensor is functionalized with a CEA aptamer instead of the corresponding antibody. The proposed aptasensor has successfully detected different concentrations (ranging from 50picogram/milliliter (pg/ml) to 50 nanogram/milliliter (ng/ml)) of CEA and achieved a detection limit as low as 50pg/ml at Vas = 0.5 V. The drain-source current shows a c/ear increase of 11.5μA under this bias.
文摘The formation of covalent adducts as a result of the interaction of metabolically activated chemicals with host macromolecules is a common critical event in mutagenic, carcinogenic, and immunologic phenomena. Because of their antigenicity and their immunogenicity, covalent adducts may be detected using sensitive immunochemical techniques. The immunochemical approaches to biomonitoring and molecular dosimetry of DNA damage are particularly attractive because they allow sensitive quantitation of specific DNA adducts present in small samples and do not rely on the use of radiolabeled adducts. Two examples of biomarker immunoassay development are presented: an avidin/biotin-amplified ELISA for the major DNA adduct of the human bladder carcinogen 4-aminobiphenyl (ABP), and a particle concentration fluorescent immunoassay (PCFIA) for the major protein adduct associated with toxicity by the prototype hepatotoxin acetaminophen. The examples illustrate critical steps in the development of biomarker immunoassays which include selection of the relevant adduct, preparation of an appropriate immunogen, immunization, characterization of antisera, and development of application-specific sample processing techniques for biomarker quantitation. Immunochemical procedures may be combined with other analytical techniques to form hybrid systems which take advantage of both the antigenicity and the physical or chemical properties of a biomarker to achieve greater specificity and/or sensitivity. The future usefulness of these new tools of molecular epidemiology will depend on a compound-by-compound validation of methods and critical evaluation of the biologic importance of the particular antigenic biomarker as an indicator of exposure and as an indicator of risk.
文摘Monoclonal antibodies against colon and pancreatic cancer, CL-2, CL-3, PS-9, PS-10, were used to detect the associated antigens in feces of patients with gastrointestinal carcinoma and non-cancer diseases. Binding inhibition test by SABC-ELISA method were performed for the measurement of the antigen level. Results showed that the associated antigen detected in feces of patients with colon cancer were significantly higher than that of non-cancer disease or normal subjects. The positive rates were 61.1% as detected with CL-2; 53.4% with CL-3; 55.0%, PS-9; and 53.3% PS-10 in cancer patients while that in normal subjects were 7%; 9%; 8%; and 8% respectively. When 'cocktail' of CL-2, PS-9 and PS-10 were used, the positive rates were 92.5% in colon cancer and 14% in normal subjects. In seven out of the sixty patients with colon cancer studied who were graded as Dukes A, the results were all positive. The results seem superior to the serologic detection and may provide a promising new approach in the early diagnosis of colon cancer.
文摘During the late incubation period or initial phase of dengue virus infection,laboratory confirmation is through viral isolation in cell culture and/or molecular investigations, or immunofluorescence,or immunohistochemistry[1].The dengue virus non-structural antigen NSl that would develop before the appearance of dengue IgM and/or IgG is emerging as a suitable option for dengue diagnosis[2].Platelet therapy is a standard clinical practice for dengue patients with severe thrombocytopenia[3].However,during introductory screening,platelet count is not being done in many cases. This results in delays of platelet therapy. In the course of the current(2010) spurt of dengue in New Delhi[4],simultaneous screening for NSl,IgM and IgG and platelet enumeration has been introduced at the
文摘Background: Children with seasonal influenza infection cause a significant burden of disease each year in the pediatric clinic. Influenza A and B viruses are the major types responsible for illness. A better understanding of the periodicity facilitates the prevention and control of influenza in children. Objective: This study aims to analyze the epidemiological patterns and subtype characterization of influenza viruses among children in Shenzhen, China. Methods: Influenza samples were collected by nasopharyngeal swabs from influenza like illness patients in Shenzhen Children’s Hospital from January 2016 to December 2018. The positive cases and influenza subtypes were determined by gold labeled antigen detection and reverse transcriptase polymerase chain reaction. The influenza periodicity and age, subtype distribution as well as the association between climate parameters and different influenza subtypes were analyzed by SPSS 22.0. Results: The influenza positive rate during 2016-2018 was 21.0%, with a highest positive rate in the year 2018. The positive rate varied by month, season, and year describing a sequence of peaks presenting primarily in all year including spring, summer and winter. The characteristics of influenza peak were different in each year, with a spring peak in 2016 and a summer plus a winter-spring peaks in 2017 and 2018. In addition, influenza B exhibited a winter-spring seasonal pattern while influenza A displayed a more variable seasonality, highlighting influenza B rather than influenza A which had a negative association with climate parameters. Influenza-positive cases were older than influenza-negative cases (P P Conclusion: Influenza activity in children from Shenzhen typically displays both winter-spring and summer peaks. Influenza A epidemic occurred separately or co-circulated with influenza B, with a winter-spring pattern for influenza B and a much more variable seasonality for influenza A. Influenza B had a negative association with climate parameters. In addition, hospitalization with influenza often occurs in younger individuals infected with influenza A.
文摘Objective To establish a highly sensitive serologic method for detecting the circulating antigen of Schistosoma japonicum for the diagnosis of this disease and the evaluation of drug therapy effect.Methods A set of complex support made of poly vinyl chloride (PVC) film slide, adsorbed with specific antibody, was used to test serum samples collected from cases with schistosomiasis japonica, by adding specific enzyme conjugate and substrate tetra methyl benzidine (TMB) with 3% H 2O 2 in 10∶0.1, developed or prepared in our laboratory. Tests were carried out with the set on cases with malaria, paragonimiasis, clonorchiasis, visceral leishmaniasis and cysticercosis as well as normal individuals as control. Serum sample of 50 μl was used in each test and reacted with reagents at room temperature for 30 minutes. The color of positive reaction was blue while negative reaction was colorless.Results Positive rate among cases with schistosomiasis japonica was 100% (45/45) in acute stage, 94.8% (530/559) in early chronic stage and 52.4% (66/126) in late stage. False positive reaction was found neither among all normal individuals (0/513), nor among cases of 155 with malaria, 120 with clonorchiasis, 24 with visceral leishmaniasis and 110 with cysticercosis, except one among 29 cases with parasgonimiasis (1/29).Conclusions The rapid one step enzyme Immunoassay (EIA) to detect circulating schistosome antigen (CSA) has been well established in our laboratory with high sensitivity and good specificity as well as reproducibility. The reaction result can be read easily even in field conditions without power supply. Moreover, the assay is time saving, simple to handle and suitable for the diagnosis of shistosomiasis japonica and evaluation of drug therapeutic effect in practice in the control program of the disease.
基金financially supported by the National Natural Science Foundation of China(Nos.51373117,51303126 and 31600800)Tianjin Natural Science and Technology Foundation(No.16ZXMJSY00010)
文摘At present,many researchers focused on the point-of-care testing(POCT),a method of disease markers detection without large-scale instruments and specialized persons.However,most POCT diagnostic methods were suffered from poor detection sensitivity or inefficiency in quantitative detection.Herein,we developed a newly QD-immune fluorescence test strips(QD-IFTS) based on quantum dots(QDs) as the fluorescence nanocarrier to prepare the immune fluorescence probes in the classical immunochromatography detection system for sensing carcino-embryonic antigen(CEA),a kind of glycoprotein produced by intestinal tissue and a broad spectrum of tumor marker for cancer diagnosis.And we designed a homemade strips fluorescence reader for detection of fluorescence intensity of QDs on the QD-IFTS.Under the optimized reaction conditions,chromatographic time of the newly QD-IFTS was only25 min,sample volume of the newly QD-IFTS was only 40 m L and the LOD of the newly QD-IFTS was 0.72 ng/m L.In addition,the efficiency and robustness of the newly QD-IFTS were confirmed by successfully application in 300 clinical serum samples,and the results revealed great potential in clinical POCT of other biomarkers.
基金Guangdong Provincial Department of Science and Technology-key research and development project(No.2020B1111160003)Shenzhen Science and Technology Innovation Commission technology breakthrough project(No.JSGG20191231141403880)+1 种基金Shenzhen San-Ming Project(No.SZSM201809085)Shenzhen Science and Technology Innovation Commission general project(No.JCYJ20180504165657443)。
文摘Early detection of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)infection is an efficient way to prevent the spread of coronavirus disease 2019(COVID-19).Detecting SARS-CoV-2 antigen can be rapid and convenient,but it is still challenging to develop highly sensitive methods for effective diagnosis.Herein,a lateral flow assay(LFA)based on fluorescent nanoparticles emitting in the second near-infrared(NIR-II)window is developed for sensitive detection of SARS-CoV-2 antigen.Benefiting from the NIR-II fluorescence with high penetration and low autofluorescence,such NIR-II based LFA allows enhanced signal-to-background ratio,and the limit of detection is down to 0.01 ng·mL^(−1)of SARS-CoV-2 antigen.In the clinical swab sample tests,the NIR-II LFA outperforms the colloidal gold LFA with higher overall percent agreement with the polymerase chain reaction test.The clinical samples with low antigen concentrations(~0.015–~0.068 ng·mL^(−1))can be successfully detected by the NIR-II LFA,but fail for the colloidal gold LFA.The NIR-II LFA can provide a promising platform for highly sensitive,rapid,and cost-effective method for early diagnosis and mass screening of SARS-CoV-2 infection.