A chemiluminescence enzyme immunoassay based on magnetic microparticles (MmPs-CLEIA) was developed to evaluate serum a-fetoprotein (AFP) in parallel with traditional colorimetric enzyme-linked immunosorbent assay (ELI...A chemiluminescence enzyme immunoassay based on magnetic microparticles (MmPs-CLEIA) was developed to evaluate serum a-fetoprotein (AFP) in parallel with traditional colorimetric enzyme-linked immunosorbent assay (ELISA).A systematic comparison between the MmPs-CLEIA and colorimetric ELISA concluded that the MPs-CLEIA exhibited fewer dosages of immunoreagents,less total assay time,and better linearity,recovery,precision,sensitivity and validity.AFP was detected in forty human serum samples by the proposed MPs-CLEIA and ELISA,and the results were compared with commercial electrochemiluminescence immunoassay (ECLIA) kit.The correlation coefficient between MPs-CLEIA and ELISA was obtained with R 2 0.6703;however,the correlation between MPs-CLEIA and ECLIA (R 2 0.9582) was obviously better than that between colorimetric ELISA and ECLIA (R 2 0.6866).展开更多
Plasma quinolone concentrations are not routinely measured in clinical practice.However,in order to optimize quinolone treatment,monitoring of plasma concentrations could sometimes be useful particularly in critically...Plasma quinolone concentrations are not routinely measured in clinical practice.However,in order to optimize quinolone treatment,monitoring of plasma concentrations could sometimes be useful particularly in critically ill patients.In this study,anti-sparfloxacin antibody was obtained by immunizing rabbits with sparfloxacin conjugated with bovine serum albumin using isobutyl chloroformate method.After the assay procedure was optimized,the standard curve of sparfloxacin was established.The practical measuring range of the competitive ELISA extended from 5 ng/mL to 2 mg/mL.The recovery rates and coefficients of variation for rat plasma,urine and tissues were 87.7-106.2% and 4.8-15.3%,respectively.To demonstrate the potential of the ELISA,a preliminary pharmacokinetics and tissue distribution study of sparfloxacin in rats and quantitative analysis of sparfloxacin in several pharmaceuticals were performed and compared with high-performance liquid chromatography(HPLC).The experimental data indicated that the proposed method would be a valuable tool in therapeutic drug monitoring(TDM) for sparfloxacin.展开更多
Vegetative propagation of seed potato often allows passaging of viruses to seed tubers,resulting in significant yield losses and reduction of potato tuber quality.Thus,virus detection approach is crucial for effective...Vegetative propagation of seed potato often allows passaging of viruses to seed tubers,resulting in significant yield losses and reduction of potato tuber quality.Thus,virus detection approach is crucial for effective virus management programs and the production of virus-free seed potatoes.Among the reported potato-infecting viruses,potato virus A(PVA)is considered as one of the most important viruses in potato-growing regions worldwide.This study prepared four hybridoma lines secreting PVA-specific monoclonal antibodies(MAbs)(2D4,8E11,14A6 and 16H10)using purified PVA virions as an immunogen.Western blotting results indicated that all the four MAbs reacted strongly and specifically with the putative capsid protein of PVA.Using these four MAbs,this study developed antigen-coated plate enzyme-linked immunosorbent assay(ACP-ELISA),Dot-ELISA and Tissue print-ELISA for detection of PVA infection in potato plants.The results indicated that PVA can be detected in crude tissue extracts from infected potato plants diluted up to 1:327680(w/v,g m L^(-1))by ACP-ELISA or up to1:10240 by Dot-ELISA.The Tissue print-ELISA is the quickest and easiest approach among the three serological assays,and is more suitable for onsite large-scale potato screening programs.Further analyses of field-collected potato samples showed that the sensitivities and specificities of the three serological approaches were similar to those of RT-PCR in PVA detection and confirmed that PVA is currently widespread in Yunnan and Zhejiang provinces of China.Hence,the results strongly suggest that these highly sensitive serological approaches based on PVA-specific MAbs are useful and powerful for PVA-free seed potato production programs and PVA field surveys.展开更多
Recombinant batroxobin(S3101)is a thrombin-like serine protease that binds to fibrinogen or is taken up by the reticuloendothelial system.A literature survey showed no adequate method that could determine sufficient c...Recombinant batroxobin(S3101)is a thrombin-like serine protease that binds to fibrinogen or is taken up by the reticuloendothelial system.A literature survey showed no adequate method that could determine sufficient concentrations to evaluate pharmacokinetic parameters for phase I clinical studies.Therefore,a sensitive method is urgently needed to support the clinical pharmacokinetic evaluation of S3101.In this study,a sensitive bioanalytical method was developed and validated,using a Quanterix single molecular array(Simoa)assay.Moreover,to thoroughly assess the platform,enzyme-linked immunosorbent assay and electrochemiluminescence assay were also developed,and their performance was compared with that of this novel technology platform.The assay was validated in compliance with the current guidelines.Measurements with the Simoa assay were precise and accurate,presenting a valid assay range from 6.55 to 4000 pg/mL.The intra-and inter-run accuracy and precision were within-19.3%to 15.3%and 5.5%to 17.0%,respectively.S3101 was stable in human serum for 280 days at-20℃and-70℃,for 2 h prior to pre-treatment and 24 h post pre-treatment at room temperature(22℃-28℃),respectively,and after five and two freeze-thaw cycles at-70℃and-20oC,respectively.The Simoa assay also demonstrated sufficient dilution linearity,assay sensitivity,and parallelism for quantifying S3101 in human serum.The Simoa assay is a sensitive and adequate method for evaluating the pharmacokinetic parameters of S3101 in human serum.展开更多
Microcystins(MCs)are a group of closely related toxic cyclic heptapeptides produced by common cyanobacte-ria,which cause lots of accidents and threatens human health.In this paper,an indirect competitive enzyme-linked...Microcystins(MCs)are a group of closely related toxic cyclic heptapeptides produced by common cyanobacte-ria,which cause lots of accidents and threatens human health.In this paper,an indirect competitive enzyme-linked immu-nosorbent assay(ic-ELISA)was established and used to detect microcystin-LR(MC-LR)in drinking and surface waters.The concentration of coating antigen was 5 mg/mL,the dilution of monoclonal antibody MC10E7 was 1:3000,the dilution of enzyme tracer(goat anti-mouse IgG-peroxidase)was 1:3000,the standard concentration of MC-LR ranged from 0.001 mg/L to 30 mg/L,and o-phenylenediamine was used as substrate.The assay showed high relativity with high performance liquid chromatography(HPLC)with a correlation coefficient of more than 99%.The relative standard deviation was less than 10%,the detection limit was achieved down to 0.01 mg/L and up to 5.1 mg/L.The quantitative detection range was from 0.03 mg/L to 3 mg/L,and the antibody had high specificity for[4-arginine]microcystins.It performed well in spite of the influence of the real samples.展开更多
Objective:To explore the feasibility of enzyme-linked immunosorbent assay(ELISA)in detecting syphilis dry blood spots.Methods:Based on dry blood spot samples,laboratory linear dial,laboratory basic dial,laboratory int...Objective:To explore the feasibility of enzyme-linked immunosorbent assay(ELISA)in detecting syphilis dry blood spots.Methods:Based on dry blood spot samples,laboratory linear dial,laboratory basic dial,laboratory interference dial,and laboratory precision dial were constructed.The linear range,sensitivity,specificity,precision and other performances of ELISA for detecting syphilis dry blood spot samples were comprehensively evaluated,and the stability of dry blood spot samples at 37℃was detected.In addition,112 suspected syphilis antibody-positive plasma samples were selected as the control,and dry blood spot samples were prepared accordingly.The clinical sensitivity,clinical specificity,positive predictive value and negative predictive value of dry blood spot samples and plasma samples in ELISA for syphilis detection were compared,and the consistency and correlation between the two samples were analyzed by the Kappa consistency test and Spearman rank correlation analysis.Results:The results of the linear analysis showed that the serial dilution of dry blood spot samples in ELISA for syphilis antibody ranged from 23to 27,and there was a good linear range[R2=0.9862(P<0.05)].Sensitivity and specificity analysis showed that the positive coincidence rate and negative coincidence rate of the two detection methods were 100%(15/15).The results of the interference dial test showed that ELISA based on dry blood spot samples could accurately detect 6 syphilis antibody samples from 18samples,and the detection accuracy rate was 100.00%(6/6).The results of the precision test showed that the RSD of syphilis antibody detection in dry blood spot samples with different dilution times(23,25and 27)was 0.24%to 3.87%between spots,0.06%to4.07%between batches and 0.49%to 3.88%between days.Within 7 days,the inter-day RSD of dry blood spots with different dilution times(23,25,27)were 0.27%,0.65%and 0.95%,respectively.The clinical sensitivity,clinical specificity,positive predictive value and negative predictive value of dry blood spot samples in ELISA detection of syphilis antibody were 96.00%(72/75),100.00%(37/37),100.00%(72/72)and 92.50%,respectively.The results of the Kappa consistency test and Spearman rank correlation analysis showed that the Kappa value of the two methods was 0.941(P<0.01),and the correlation coefficientρbetween the S/CO ratios of the two methods was 0.211(P<0.01).The comparison of S/CO ratio results showed that the distribution characteristics of the S/CO ratio between the two methods were similar,and the ratio distribution was relatively concentrated.Conclusion:using dry blood spot samples to perform ELISA for syphilis detection has good precision,strong anti-interference ability and excellent stability.Although false-positive results appear in weak positive samples,it still has a high application value in ELISA for syphilis antibody detection,which can provide an important reference for disease diagnosis.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
BACKGROUND Defective neutrophil regulation in inflammatory bowel disease(IBD)is thought to play an important role in the onset or manifestation of IBD,as it could lead to damage of the intestinal mucosal barrier by th...BACKGROUND Defective neutrophil regulation in inflammatory bowel disease(IBD)is thought to play an important role in the onset or manifestation of IBD,as it could lead to damage of the intestinal mucosal barrier by the infiltration of neutrophils in the inflamed mucosa and the accumulation of pathogens.Like neutrophils in the context of innate immune responses,immunoglobulin A(IgA)as an acquired immune response partakes in the defense of the intestinal epithelium.Under normal conditions,IgA contributes to the elimination of microbes,but in connection with the loss of tolerance to chitinase 3-like 1(CHI3L1)in IBD,IgA could participate in CHI3L1-mediated improved adhesion and invasion of potentially pathogenic microorganisms.The tolerance brake to CHI3L1 and the occurrence of IgA autoantibodies to this particular target,the exact role and underlying mechanisms of CHI3L1 in the pathogenesis of IBD are still unclear.AIM To determine the predictive potential of Ig subtypes of a novel serological marker,anti-CHI3L1 autoantibodies(aCHI3L1)in determining the disease phenotype,therapeutic strategy and long-term disease course in a prospective referral cohort of adult IBD patients.METHODS Sera of 257 Crohn’s disease(CD)and 180 ulcerative colitis(UC)patients from a tertiary IBD referral center of Hungary(Division of Gastroenterology,Department of Internal Medicine,Faculty of Medicine,University of Debrecen)were assayed for IgG,IgA,and secretory IgA(sIgA)type aCHI3L1 by enzyme-linked immunosorbent assay using recombinant CHI3L1,along with 86 healthy controls(HCONT).RESULTS The IgA type was more prevalent in CD than in UC(29.2%vs 11.1%)or HCONT(2.83%;P<0.0001 for both).However,sIgA subtype aCHI3L1 positivity was higher in both CD and UC patients than in HCONT(39.3%and 32.8%vs 4.65%,respectively;P<0.0001).The presence of both IgA and sIgA aCHI3L1 antibodies was associated with colonic involvement(P<0.0001 and P=0.038,respectively)in patients with CD.Complicated disease behavior at sample procurement was associated with aCHI3L1 sIgA positivity(57.1%vs 36.0%,P=0.009).IgA type aCH3L1 was more prevalent in patients with frequent relapse during the disease course in the CD group(46.9%vs 25.7%,P=0.005).In a group of patients with concomitant presence of pure inflammatory luminal disease and colon involvement at the time of diagnosis,positivity for IgA or sIgA type aCH3L1 predicted faster progression towards a complicated disease course in time-dependent models.This association disappeared after merging subgroups of different disease locations.CONCLUSION CHI3L1 is a novel neutrophil autoantigenic target in IBD.The consideration of antibody classes along with location-based prediction may transform the future of serology in IBD.展开更多
The ongoing Coronavirus disease 19 pandemic has likely changed the world in ways not seen in the past.Neutralizing antibody(NAb)assays play an important role in the management of the severe acute respiratory syndrome ...The ongoing Coronavirus disease 19 pandemic has likely changed the world in ways not seen in the past.Neutralizing antibody(NAb)assays play an important role in the management of the severe acute respiratory syndrome coronavirus-2(SARS-CoV-2)outbreak.Using these tools,we can assess the presence and duration of antibody-mediated protection in naturally infected individuals,screen convalescent plasma preparations for donation,test the efficacy of immunotherapy,and analyze NAb titers and persistence after vaccination to predict vaccine-induced protective effects.This review briefly summarizes the various methods used for the detection of SARS-CoV-2 NAbs and compares their advantages and disadvantages to facilitate their development and clinical application.展开更多
Objective The aim of the study was to investigate the clinicopathological characteristics of hypoxiainducible factor-1α(HIF-1α) and vascular endothelial growth factor(VEGF) expression in patients with lung cancer.Me...Objective The aim of the study was to investigate the clinicopathological characteristics of hypoxiainducible factor-1α(HIF-1α) and vascular endothelial growth factor(VEGF) expression in patients with lung cancer.Methods Cancerous and noncancerous tissues were collected post-operation from 115 patients with lung cancers by the self-control method. Total RNA was extracted from the lung tissues. The status of tissue HIF-1α expression and intercellular distribution was observed by immunochemistry using a tissue microarray. The expression levels of circulating HIF-1α and VEGF were detected by enzyme-linked immunosorbent assay(ELISA).Results The expression of serum HIF-1α [(138.3 ± 28.8) μg/L] in the group of patients with lung cancer was significantly higher(P < 0.01) than that in the group of patients with pneumonia [(58.8 ± 14.5) μg/L] and the control group of patients ((24.1 ± 3.3) μg/L)There was a strong positive correlation of serum HIF-1α levels(r = 0.937, P < 0.01) with serum VEGF levels. The specific concentration of total RNA [(1.52 ± 1.14) μg/mg wet lung tissues] in the cancerous tissues was significantly higher(t = 8.494, P < 0.001) than that in the noncancerous tissues ((0.58 ± 0.33) μg/mg)The clinicopathological features of HIF-1α expression in lung cancer tissues revealed a significant relationship between positive HIF-1α expression and patient sex(χ~2 = 4.494, P = 0.034), tumor size(χ~2 = 4.679, P = 0.031), differentiation degree(χ~2= 8.846, P = 0.012), and presence of lymphatic node metastasis(χ~2= 6.604, P = 0.037).Conclusion Abnormal HIF-1α expression in lung cancer is closely related with nucleic acid metabolism and angiogenesis, and it may be helpful in the diagnosis and identification of lung cancer.展开更多
Amebic liver abscesses(ALAs)are the most commonly encountered extraintestinal manifestation of human invasive amebiasis,which results from Entamoeba histolytica(E.histolytica)spreading extraintestinally.Amebiasis can ...Amebic liver abscesses(ALAs)are the most commonly encountered extraintestinal manifestation of human invasive amebiasis,which results from Entamoeba histolytica(E.histolytica)spreading extraintestinally.Amebiasis can be complicated by liver abscess in 9%of cases,and ALAs led to almost 50000 fatalities worldwide in 2010.Although there have been fewer and fewer cases in the past several years,ALAs remain an important public health problem in endemic areas.E.histolytica causes both amebic colitis and liver abscess by breaching the host’s innate defenses and invading the intestinal mucosa.Trophozoites often enter the circulatory system,where they are filtered in the liver and produce abscesses,and develop into severe invasive diseases such as ALAs.The clinical presentation can appear to be colitis,including upper-right abdominal pain accompanied by a fever in ALA cases.Proper diagnosis requires nonspecific liver imaging as well as detecting anti-E.histolytica antibodies;however,these antibodies cannot be used to distinguish between a previous infection and an acute infection.Therefore,diagnostics primarily aim to use PCR or enzyme-linked immunosorbent assay to detect E.histolytica.ALAs can be treated medically,and percutaneous catheter drainage is only necessary in approximately 15%of cases.The indicated treatment is to administer an amebicidal drug(such as tinidazole or metronidazole)and paromomycin or other luminal cysticidal agent for clinical disease.Prognosis is good with almost universal recovery.Establishing which diagnostic methods are most efficacious will necessitate further analysis of similar clinical cases.展开更多
Hepatitis E is caused by hepatitis E virus(HEV),which has been classifi ed into four genotypes.Genotypes 3 and 4 are regarded as zoonotic pathogens.Accumulating researches indicate that genotype 4 is the main HEV stra...Hepatitis E is caused by hepatitis E virus(HEV),which has been classifi ed into four genotypes.Genotypes 3 and 4 are regarded as zoonotic pathogens.Accumulating researches indicate that genotype 4 is the main HEV strain circulating in China,and there are high levels of seropositive pigs and human in some provinces of China.In this study,serum samples from pigs and from human occupationally exposed to pigs were obtained from pig farms in Guangdong Province,in subtropical southern China,in order to investigate for the fi rst time the prevalence of anti-HEV immunoglobulin G(Ig G)in the region.Antibodies against HEV were detected by Enzyme-Linked Immunosorbent Assay(ELISA)using a commercially marketed kit.The results showed that high numbers of pigs(74/94;78.7%)and human(50/94;53.2%)from three pig farms in Guangdong Province were positive for anti-HEV Ig G.The correlation coeffi cient relating the prevalence in pigs and human on different farms was 0.920.The seropositive rate in males(human)was 48.8%(20/41)and that in females was 47.7%(9/19),which showed no statistically signifi cant difference.These data indicated that there was a high prevalence of anti-HEV antibodies in pigs and in human with occupational exposure to pigs.The risk of infection with HEV in both human and pigs in Guangdong Province appeared to be age-dependent,to a certain extent.This study provided basic data for further researches on HEV and was a reminder that more attention should be paid to HEV infection both in pigs and workers on pig farms in the study region.展开更多
BACKGROUND The recent rise in the incidence of hepatitis B virus(HBV)infections in a densely populated city of eastern India(“mixing vessel”of people of varied socioeconomic and immune status)prompted this study.App...BACKGROUND The recent rise in the incidence of hepatitis B virus(HBV)infections in a densely populated city of eastern India(“mixing vessel”of people of varied socioeconomic and immune status)prompted this study.Applying saliva on fingers for enumerating bank notes is a common practice in the Indian subcontinent.Paper notes may be a potential source of“horizontal”transmission of this virus,especially if there are cuts/bruises on the oral mucous membrane or skin.AIM To investigate whether paper currencies could be a plausible mode of horizontal transmission of HBV infection.METHODS Polymerase chain reactions(PCR)followed by nucleotide sequencing was done for the detection of HBV.Hepatitis B virus surface antigen enzyme-linked immunosorbent assay(HBsAg ELISA)was performed on all HBV deoxyribonucleic acid-positive samples to check the detectability of the virus.Atomic force microscopy(AFM)was carried out for visual confirmation of HBV particles in ultracentrifuged/immunoprecipitated samples from currency paper washings.RESULTS HBV-specific PCRs on pellets obtained after ultracentrifugation/immunoprecipitation of the currency paper washings detected potentially intact/viable HBV(genotype D2)in 7.14%of samples(n=70).AFM gave the visual confirmation of HBV particles in ultracentrifuged/immunoprecipitated samples from currency paper washings.However,HBV isolates from the currency notes could not be detected by HBsAg ELISA.CONCLUSION It is a common practice in the Indian subcontinent to count paper currencies by applying saliva on fingertips.Paper notes may be a potential source of“horizontal”transmission of this virus,especially if there are cuts/bruises on the oral mucous membrane or skin,but it was practically not possible to demonstrate experimentally such transmission.Detection of potentially intact/viable and“occult”HBV from currency poses potential risk of silent transmission of this virus among the general population.展开更多
BACKGROUND Alpha-defensin has been widely studied for the diagnosis of periprosthetic joint infection(PJI).However,there is a lack of detailed information regarding the proper laboratory technique of the enzyme-linked...BACKGROUND Alpha-defensin has been widely studied for the diagnosis of periprosthetic joint infection(PJI).However,there is a lack of detailed information regarding the proper laboratory technique of the enzyme-linked immunosorbent assay(ELISA)method,such as sample dilution.AIM To assess the influence of dilution in the synovial fluid during ELISA for the diagnosis of knee PJI;and determine which dilution presents a better performance.METHODS Forty samples of synovial fluid from arthroplasty knees were included,17 in the infected group and 23 in the aseptic group,according to Musculoskeletal Infection Society criteria.Initially,five synovial fluid samples from each group were assessed for quantitative analysis of alpha-defensin using ELISA.Different dilution ratios(1:10,1:100,1:500,1:1000 and 1:5000)were tested based on the predetermined cutoff value of 5.2 mg/L.The dilutions that performed better were used to compare the results of all samples.RESULTS For infected cases,a gradual increase in the dilution of synovial fluid samples led to an equivalent increase in alpha-defensin level.The same was not observed in the aseptic cases.Both 1:1000 and 1:5000 dilutions presented satisfactory results to differentiate infected and aseptic cases.Further analyses were performed using 1:1000 and 1:5000 for all 40 samples.The 1:1000 dilution resulted in a sensitivity of 88.2%(95%CI,66%-98%)and specificity of 95.7%(95%CI,79%-99%),whereas the 1:5000 dilution presented a sensitivity of 94.1%(95%CI,73%-99%)and a specificity of 100%(95%CI,86%-100%).CONCLUSION The synovial fluid dilution had an important influence on the alpha-defensin ELISA results.Dilutions of 1:5000 showed the best performance for the diagnosis of knee PJI.The results of this study set the basis for a more reliable and reproducible alpha-defensin ELISA during the investigation of PJI,contributing to the expansion of this technique in different treatment centers worldwide.展开更多
The glomerulonephritis disease is characterized by inflammation of glomeruli or small blood vessels in the kidney that causes kidney diseases. The reason of glomerulonephritis disease is to deposit the anti-GBM auto a...The glomerulonephritis disease is characterized by inflammation of glomeruli or small blood vessels in the kidney that causes kidney diseases. The reason of glomerulonephritis disease is to deposit the anti-GBM auto antibody in the glomerular basement membrane. The type IV collagen is the main component of glomerular basement membrane that has α3 chain of type (IV) collagen of non-collagenous domain which contains N-terminal 7S domain, a triple helical collagenous domain and C-terminal non-collagenous glomerular domain (NC1). The amino terminal of α3 (IV) NC1 that induces the Experimental Autoimmuno Glomerulonephritis (EAG) in rat model has been identified. The recombinant rat α3 (IV) NC1 antigen has nine amino acid spans that are consistent with antibody or T cell epitope that induces in EAG. The research is carried out on the recombinant rat α3 (IV) NC1 production, purification, quantification, and characterization. The circulation of anti-GBM antibody in glomerular basement membrane can be measured by the ELISA assay. In addition, the recombinant rat antigen is secreted in HEK293 cell supernatant that is purified by Anti-FLAG M2 monoclonal IgG antibody affinity column and characterized and quantified by SDS-PAGE gel electrophoresis and Western blotting techniques.展开更多
Canine herpesvirus (CHV-1) causes disease associated with high mortality in infect-ed puppies, which represents large financial losses for dog breeders. Since CHV-1 at the time of the study he had not been reported in...Canine herpesvirus (CHV-1) causes disease associated with high mortality in infect-ed puppies, which represents large financial losses for dog breeders. Since CHV-1 at the time of the study he had not been reported in Mexico, the main objective of this study was to determine the prevalence of antibodies against CHV-1 in canine kennels in the metropolitan area of Mexico City. A commercial enzyme-linked immuno-sorbent assay (ELISA) was used, and the results were compared to those of a viral neutralization test. The ELISA kit uses the complete viral particle as the antigen. The plaque reduction neutralization test was combined with the immunoperoxidase technique because of the low cytopathic effect of CHV-1. Neutralizing antibodies were also detected in 20 randomly selected samples. The prevalence of CHV-1 with ELISA was 87%. The concordance between ELISA and serum neutralization (SN) was 0.1129, the sensitivity of the ELISA against SN was 1.0 (100%), the positive predic-tive value was 0.39 (39%), and the negative predictive value was 1 (100%). These results show that ELISA is useful for monitoring the dog population for CHV-1;a positive test result requires confirmation with an SN test, and a negative ELISA result indicates a high probability of being SN-negative. The only variables that were sta-tistically associated with CHV-1 prevalence were breed and kennel. A statistically significant relationship between the degree of ELISA and SN titer was obtained, with a confidence level of 95%. None of the clinical presentation factors was statistically significant. These results suggest that most of the canine population studied in Mex-ico is in a herpesvirus latency state.展开更多
The global pandemic caused by severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)virus has necessitated rapid,easy-to-use,and accurate diagnostic methods to monitor the virus infection.Herein,a ratiometric flu...The global pandemic caused by severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)virus has necessitated rapid,easy-to-use,and accurate diagnostic methods to monitor the virus infection.Herein,a ratiometric fluorescence enzyme-linked immunosorbent assay(ELISA)was developed using Si-fluorescein isothiocyanate nanoparticles(FITC NPs)for detecting SARSCoV-2 nucleocapsid(N)protein.Si-FITC NPs were prepared by a one-pot hydrothermal method using 3-aminopropyl triethoxysilane(APTES)-FITC as the Si source.This method did not need post-modification and avoided the reduction in quantum yield and stability.The p-nitrophenyl(pNP)produced by the alkaline phosphatase(ALP)-mediated hydrolysis of pnitrophenyl phosphate(pNPP)could quench Si fluorescence in Si-FITC NPs via the inner filter effect.In ELISA,an immunocomplex was formed by the recognition of capture antibody/N protein/reporter antibody.ALP-linked secondary antibody bound to the reporter antibody and induced pNPP hydrolysis to specifically quench Si fluorescence in Si-FITC NPs.The change in fluorescence intensity ratio could be used for detecting N protein,with a wide linearity range(0.01-10.0 and 50-300 ng/mL)and low detection limit(0.002 ng/mL).The concentration of spiked SARS-CoV-2 N protein could be determined accurately in human serum.Moreover,this proposed method can accurately distinguish coronavirus disease 2019(COVID-19)and non-COVID-19 patient samples.Therefore,this simple,sensitive,and accurate method can be applied for the early diagnosis of SARS-CoV-2 virus infection.展开更多
Anti-idiotypic antibodies which can mimic antigens have many potential applications in the immunoassay.This research used the monoclonal antibody of the conserved chemical region of pyrethroids,3-phenoxybenzoic acid(P...Anti-idiotypic antibodies which can mimic antigens have many potential applications in the immunoassay.This research used the monoclonal antibody of the conserved chemical region of pyrethroids,3-phenoxybenzoic acid(PBA),and the domain antibody library to develop an environmentally-friendly immunoassay for the detection of pyrethroids.The domain antibodies(A8,B8,and C6)which bound to anti-PBA monoclonal antibody(MAb)were isolated from a naive phage display human domain antibody library.This domain antibody is cloneable,pyrethroid-free,and applicable as a competitive mimetic antigen in the immunoassay.The best immunoassay was achieved using A8,resulting in IC50 of 0.714μg/mL for PBA,1.775μg/mL for Cypermethrin,1.624μg/mL forβ-cypermethrin,3.675μg/mL for Fenvalerate,and 4.895μg/mL for Flucythrinate.This way of selecting anti-idiotypic antibodies to detect pyrethroids could provide potential applications in developing immunoassays for identifying various chemical contaminants in food.展开更多
This research describes a fast detection method on the basis of enzyme-linked immunosorbent assay(ELISA)for Escherichia coli in drainage of wastewater treatment plants.Optimized conditions such as the reaction format(...This research describes a fast detection method on the basis of enzyme-linked immunosorbent assay(ELISA)for Escherichia coli in drainage of wastewater treatment plants.Optimized conditions such as the reaction format(sandwich or direct),the concentrations of diluted horse radish peroxidase(HRP)-E.coli conjugate,and anti-HPR antibody and pretreatment of E.coli were studied.Those results showed that the linear range of detection for E.coli was 10 cfu/mL-6×10^(4) cfu/mL.Compared with conventional methods,it is a convenient and sensitive detection method with low cost.展开更多
OBJECTIVE:To investigate the effects of acupoint catgut embedding for 3 weeks on lung tissue,blood immunoglobulin E(IgE)and interleutin-4(IL-4),brain tissue microglia x-42(OX-42)and toll-like receptor-2(TLR-2)in rats ...OBJECTIVE:To investigate the effects of acupoint catgut embedding for 3 weeks on lung tissue,blood immunoglobulin E(IgE)and interleutin-4(IL-4),brain tissue microglia x-42(OX-42)and toll-like receptor-2(TLR-2)in rats with allergic rhinitis of lung deficiency type.METHODS:Forty-five female Sprague-Dawley rats were randomly divided for two times.The first time,they were randomly divided into model group and blank group(Group C)according to 2∶1,and the second time,the model group were randomly divided into model control group(Group B)and intervention treatment group(Group A)according to 1∶1.15 in each group.For Group A and Group B,the lung deficiency model was made by"sulfur–moxa fumigation",and then the allergic rhinitis model was established by"ovalbumin(OVA)sensitization".Then catgut embedding was performed at acupoints in Group A and not in Group B.After 3 weeks,collect lung tissue samples for hematoxylin-eosin staining,then take blood to observe the concentration of IgE and IL-4,and finally take brain tissue to observe the results of OX-42 and TLR-2.RESULTS:IgE level(μg/m L)was(3.11±0.20)in the Group A,(4.19±0.44)in the Group B,and(2.29±0.30)in the Group C(all P<0.001).IL-4 level(pg/mL)was(14.2±0.7)in the Group A,(18.6±2.4)in the Group B,and(11.4±1.2)for the Group C(all P<0.001).The mean OD for OX-42 is(0.1728±0.0016)in the Group A,(0.1810±0.0046)in the Group B and(0.1674±0.0025)in the Group C(all P<0.001).CONCLUSION:Although 3 weeks of acupoint catgut embedding already showed obvious efficacy on rats with allergic rhinitis,the allergic reaction in the body still continued.To achieve further treatment,prolonging the catgut embedding time is necessary.展开更多
基金supported by the National Basic Research Program of China (973 Program,no. 2007CB714507)National Nature Science Foundation of China (no. 90813015)
文摘A chemiluminescence enzyme immunoassay based on magnetic microparticles (MmPs-CLEIA) was developed to evaluate serum a-fetoprotein (AFP) in parallel with traditional colorimetric enzyme-linked immunosorbent assay (ELISA).A systematic comparison between the MmPs-CLEIA and colorimetric ELISA concluded that the MPs-CLEIA exhibited fewer dosages of immunoreagents,less total assay time,and better linearity,recovery,precision,sensitivity and validity.AFP was detected in forty human serum samples by the proposed MPs-CLEIA and ELISA,and the results were compared with commercial electrochemiluminescence immunoassay (ECLIA) kit.The correlation coefficient between MPs-CLEIA and ELISA was obtained with R 2 0.6703;however,the correlation between MPs-CLEIA and ECLIA (R 2 0.9582) was obviously better than that between colorimetric ELISA and ECLIA (R 2 0.6866).
文摘Plasma quinolone concentrations are not routinely measured in clinical practice.However,in order to optimize quinolone treatment,monitoring of plasma concentrations could sometimes be useful particularly in critically ill patients.In this study,anti-sparfloxacin antibody was obtained by immunizing rabbits with sparfloxacin conjugated with bovine serum albumin using isobutyl chloroformate method.After the assay procedure was optimized,the standard curve of sparfloxacin was established.The practical measuring range of the competitive ELISA extended from 5 ng/mL to 2 mg/mL.The recovery rates and coefficients of variation for rat plasma,urine and tissues were 87.7-106.2% and 4.8-15.3%,respectively.To demonstrate the potential of the ELISA,a preliminary pharmacokinetics and tissue distribution study of sparfloxacin in rats and quantitative analysis of sparfloxacin in several pharmaceuticals were performed and compared with high-performance liquid chromatography(HPLC).The experimental data indicated that the proposed method would be a valuable tool in therapeutic drug monitoring(TDM) for sparfloxacin.
基金supported by the National Key Research and Development Program of China(2017YFD0201604)the National Natural Science Foundation of China(31571976)。
文摘Vegetative propagation of seed potato often allows passaging of viruses to seed tubers,resulting in significant yield losses and reduction of potato tuber quality.Thus,virus detection approach is crucial for effective virus management programs and the production of virus-free seed potatoes.Among the reported potato-infecting viruses,potato virus A(PVA)is considered as one of the most important viruses in potato-growing regions worldwide.This study prepared four hybridoma lines secreting PVA-specific monoclonal antibodies(MAbs)(2D4,8E11,14A6 and 16H10)using purified PVA virions as an immunogen.Western blotting results indicated that all the four MAbs reacted strongly and specifically with the putative capsid protein of PVA.Using these four MAbs,this study developed antigen-coated plate enzyme-linked immunosorbent assay(ACP-ELISA),Dot-ELISA and Tissue print-ELISA for detection of PVA infection in potato plants.The results indicated that PVA can be detected in crude tissue extracts from infected potato plants diluted up to 1:327680(w/v,g m L^(-1))by ACP-ELISA or up to1:10240 by Dot-ELISA.The Tissue print-ELISA is the quickest and easiest approach among the three serological assays,and is more suitable for onsite large-scale potato screening programs.Further analyses of field-collected potato samples showed that the sensitivities and specificities of the three serological approaches were similar to those of RT-PCR in PVA detection and confirmed that PVA is currently widespread in Yunnan and Zhejiang provinces of China.Hence,the results strongly suggest that these highly sensitive serological approaches based on PVA-specific MAbs are useful and powerful for PVA-free seed potato production programs and PVA field surveys.
文摘Recombinant batroxobin(S3101)is a thrombin-like serine protease that binds to fibrinogen or is taken up by the reticuloendothelial system.A literature survey showed no adequate method that could determine sufficient concentrations to evaluate pharmacokinetic parameters for phase I clinical studies.Therefore,a sensitive method is urgently needed to support the clinical pharmacokinetic evaluation of S3101.In this study,a sensitive bioanalytical method was developed and validated,using a Quanterix single molecular array(Simoa)assay.Moreover,to thoroughly assess the platform,enzyme-linked immunosorbent assay and electrochemiluminescence assay were also developed,and their performance was compared with that of this novel technology platform.The assay was validated in compliance with the current guidelines.Measurements with the Simoa assay were precise and accurate,presenting a valid assay range from 6.55 to 4000 pg/mL.The intra-and inter-run accuracy and precision were within-19.3%to 15.3%and 5.5%to 17.0%,respectively.S3101 was stable in human serum for 280 days at-20℃and-70℃,for 2 h prior to pre-treatment and 24 h post pre-treatment at room temperature(22℃-28℃),respectively,and after five and two freeze-thaw cycles at-70℃and-20oC,respectively.The Simoa assay also demonstrated sufficient dilution linearity,assay sensitivity,and parallelism for quantifying S3101 in human serum.The Simoa assay is a sensitive and adequate method for evaluating the pharmacokinetic parameters of S3101 in human serum.
基金This work was supported by the National High-Tech Research and Development(863)Program of China(2002AA649160).
文摘Microcystins(MCs)are a group of closely related toxic cyclic heptapeptides produced by common cyanobacte-ria,which cause lots of accidents and threatens human health.In this paper,an indirect competitive enzyme-linked immu-nosorbent assay(ic-ELISA)was established and used to detect microcystin-LR(MC-LR)in drinking and surface waters.The concentration of coating antigen was 5 mg/mL,the dilution of monoclonal antibody MC10E7 was 1:3000,the dilution of enzyme tracer(goat anti-mouse IgG-peroxidase)was 1:3000,the standard concentration of MC-LR ranged from 0.001 mg/L to 30 mg/L,and o-phenylenediamine was used as substrate.The assay showed high relativity with high performance liquid chromatography(HPLC)with a correlation coefficient of more than 99%.The relative standard deviation was less than 10%,the detection limit was achieved down to 0.01 mg/L and up to 5.1 mg/L.The quantitative detection range was from 0.03 mg/L to 3 mg/L,and the antibody had high specificity for[4-arginine]microcystins.It performed well in spite of the influence of the real samples.
文摘Objective:To explore the feasibility of enzyme-linked immunosorbent assay(ELISA)in detecting syphilis dry blood spots.Methods:Based on dry blood spot samples,laboratory linear dial,laboratory basic dial,laboratory interference dial,and laboratory precision dial were constructed.The linear range,sensitivity,specificity,precision and other performances of ELISA for detecting syphilis dry blood spot samples were comprehensively evaluated,and the stability of dry blood spot samples at 37℃was detected.In addition,112 suspected syphilis antibody-positive plasma samples were selected as the control,and dry blood spot samples were prepared accordingly.The clinical sensitivity,clinical specificity,positive predictive value and negative predictive value of dry blood spot samples and plasma samples in ELISA for syphilis detection were compared,and the consistency and correlation between the two samples were analyzed by the Kappa consistency test and Spearman rank correlation analysis.Results:The results of the linear analysis showed that the serial dilution of dry blood spot samples in ELISA for syphilis antibody ranged from 23to 27,and there was a good linear range[R2=0.9862(P<0.05)].Sensitivity and specificity analysis showed that the positive coincidence rate and negative coincidence rate of the two detection methods were 100%(15/15).The results of the interference dial test showed that ELISA based on dry blood spot samples could accurately detect 6 syphilis antibody samples from 18samples,and the detection accuracy rate was 100.00%(6/6).The results of the precision test showed that the RSD of syphilis antibody detection in dry blood spot samples with different dilution times(23,25and 27)was 0.24%to 3.87%between spots,0.06%to4.07%between batches and 0.49%to 3.88%between days.Within 7 days,the inter-day RSD of dry blood spots with different dilution times(23,25,27)were 0.27%,0.65%and 0.95%,respectively.The clinical sensitivity,clinical specificity,positive predictive value and negative predictive value of dry blood spot samples in ELISA detection of syphilis antibody were 96.00%(72/75),100.00%(37/37),100.00%(72/72)and 92.50%,respectively.The results of the Kappa consistency test and Spearman rank correlation analysis showed that the Kappa value of the two methods was 0.941(P<0.01),and the correlation coefficientρbetween the S/CO ratios of the two methods was 0.211(P<0.01).The comparison of S/CO ratio results showed that the distribution characteristics of the S/CO ratio between the two methods were similar,and the ratio distribution was relatively concentrated.Conclusion:using dry blood spot samples to perform ELISA for syphilis detection has good precision,strong anti-interference ability and excellent stability.Although false-positive results appear in weak positive samples,it still has a high application value in ELISA for syphilis antibody detection,which can provide an important reference for disease diagnosis.
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-
基金Supported by the German Federal Ministry of Education and Research(BMBF-Wachstumskern-PRAEMED.BIO),03WKDB2Csupported by the Janos Bolyai Research Scholarship of the Hungarian Academy of Sciences,BO/00232/17/5+1 种基金Research Grants of National Research Development and Innovation Office,K115818/2015/1New National Excellence Program of the Ministry of Human Capacities,ÚNKP-18-4 Bolyai Plus.
文摘BACKGROUND Defective neutrophil regulation in inflammatory bowel disease(IBD)is thought to play an important role in the onset or manifestation of IBD,as it could lead to damage of the intestinal mucosal barrier by the infiltration of neutrophils in the inflamed mucosa and the accumulation of pathogens.Like neutrophils in the context of innate immune responses,immunoglobulin A(IgA)as an acquired immune response partakes in the defense of the intestinal epithelium.Under normal conditions,IgA contributes to the elimination of microbes,but in connection with the loss of tolerance to chitinase 3-like 1(CHI3L1)in IBD,IgA could participate in CHI3L1-mediated improved adhesion and invasion of potentially pathogenic microorganisms.The tolerance brake to CHI3L1 and the occurrence of IgA autoantibodies to this particular target,the exact role and underlying mechanisms of CHI3L1 in the pathogenesis of IBD are still unclear.AIM To determine the predictive potential of Ig subtypes of a novel serological marker,anti-CHI3L1 autoantibodies(aCHI3L1)in determining the disease phenotype,therapeutic strategy and long-term disease course in a prospective referral cohort of adult IBD patients.METHODS Sera of 257 Crohn’s disease(CD)and 180 ulcerative colitis(UC)patients from a tertiary IBD referral center of Hungary(Division of Gastroenterology,Department of Internal Medicine,Faculty of Medicine,University of Debrecen)were assayed for IgG,IgA,and secretory IgA(sIgA)type aCHI3L1 by enzyme-linked immunosorbent assay using recombinant CHI3L1,along with 86 healthy controls(HCONT).RESULTS The IgA type was more prevalent in CD than in UC(29.2%vs 11.1%)or HCONT(2.83%;P<0.0001 for both).However,sIgA subtype aCHI3L1 positivity was higher in both CD and UC patients than in HCONT(39.3%and 32.8%vs 4.65%,respectively;P<0.0001).The presence of both IgA and sIgA aCHI3L1 antibodies was associated with colonic involvement(P<0.0001 and P=0.038,respectively)in patients with CD.Complicated disease behavior at sample procurement was associated with aCHI3L1 sIgA positivity(57.1%vs 36.0%,P=0.009).IgA type aCH3L1 was more prevalent in patients with frequent relapse during the disease course in the CD group(46.9%vs 25.7%,P=0.005).In a group of patients with concomitant presence of pure inflammatory luminal disease and colon involvement at the time of diagnosis,positivity for IgA or sIgA type aCH3L1 predicted faster progression towards a complicated disease course in time-dependent models.This association disappeared after merging subgroups of different disease locations.CONCLUSION CHI3L1 is a novel neutrophil autoantigenic target in IBD.The consideration of antibody classes along with location-based prediction may transform the future of serology in IBD.
基金supported by grants from the Applied Basic Research Key Project of Wuhan Municipal Bureau of Science and Technology(2020020601012218)the Fundamental Research Funds for the Central Universities(HUST COVID-19 Rapid Response Call No.2020kfyXGYJ040).
文摘The ongoing Coronavirus disease 19 pandemic has likely changed the world in ways not seen in the past.Neutralizing antibody(NAb)assays play an important role in the management of the severe acute respiratory syndrome coronavirus-2(SARS-CoV-2)outbreak.Using these tools,we can assess the presence and duration of antibody-mediated protection in naturally infected individuals,screen convalescent plasma preparations for donation,test the efficacy of immunotherapy,and analyze NAb titers and persistence after vaccination to predict vaccine-induced protective effects.This review briefly summarizes the various methods used for the detection of SARS-CoV-2 NAbs and compares their advantages and disadvantages to facilitate their development and clinical application.
基金Supported by grants-in-aid from Projects of the Society Development(No.BK2013048) of Nantong Citythe Departments of Jiangsu S&T or Health(No.WSW-011)the International S&T Cooperation Program of China(No.2013DFA32150)
文摘Objective The aim of the study was to investigate the clinicopathological characteristics of hypoxiainducible factor-1α(HIF-1α) and vascular endothelial growth factor(VEGF) expression in patients with lung cancer.Methods Cancerous and noncancerous tissues were collected post-operation from 115 patients with lung cancers by the self-control method. Total RNA was extracted from the lung tissues. The status of tissue HIF-1α expression and intercellular distribution was observed by immunochemistry using a tissue microarray. The expression levels of circulating HIF-1α and VEGF were detected by enzyme-linked immunosorbent assay(ELISA).Results The expression of serum HIF-1α [(138.3 ± 28.8) μg/L] in the group of patients with lung cancer was significantly higher(P < 0.01) than that in the group of patients with pneumonia [(58.8 ± 14.5) μg/L] and the control group of patients ((24.1 ± 3.3) μg/L)There was a strong positive correlation of serum HIF-1α levels(r = 0.937, P < 0.01) with serum VEGF levels. The specific concentration of total RNA [(1.52 ± 1.14) μg/mg wet lung tissues] in the cancerous tissues was significantly higher(t = 8.494, P < 0.001) than that in the noncancerous tissues ((0.58 ± 0.33) μg/mg)The clinicopathological features of HIF-1α expression in lung cancer tissues revealed a significant relationship between positive HIF-1α expression and patient sex(χ~2 = 4.494, P = 0.034), tumor size(χ~2 = 4.679, P = 0.031), differentiation degree(χ~2= 8.846, P = 0.012), and presence of lymphatic node metastasis(χ~2= 6.604, P = 0.037).Conclusion Abnormal HIF-1α expression in lung cancer is closely related with nucleic acid metabolism and angiogenesis, and it may be helpful in the diagnosis and identification of lung cancer.
文摘Amebic liver abscesses(ALAs)are the most commonly encountered extraintestinal manifestation of human invasive amebiasis,which results from Entamoeba histolytica(E.histolytica)spreading extraintestinally.Amebiasis can be complicated by liver abscess in 9%of cases,and ALAs led to almost 50000 fatalities worldwide in 2010.Although there have been fewer and fewer cases in the past several years,ALAs remain an important public health problem in endemic areas.E.histolytica causes both amebic colitis and liver abscess by breaching the host’s innate defenses and invading the intestinal mucosa.Trophozoites often enter the circulatory system,where they are filtered in the liver and produce abscesses,and develop into severe invasive diseases such as ALAs.The clinical presentation can appear to be colitis,including upper-right abdominal pain accompanied by a fever in ALA cases.Proper diagnosis requires nonspecific liver imaging as well as detecting anti-E.histolytica antibodies;however,these antibodies cannot be used to distinguish between a previous infection and an acute infection.Therefore,diagnostics primarily aim to use PCR or enzyme-linked immunosorbent assay to detect E.histolytica.ALAs can be treated medically,and percutaneous catheter drainage is only necessary in approximately 15%of cases.The indicated treatment is to administer an amebicidal drug(such as tinidazole or metronidazole)and paromomycin or other luminal cysticidal agent for clinical disease.Prognosis is good with almost universal recovery.Establishing which diagnostic methods are most efficacious will necessitate further analysis of similar clinical cases.
基金Supported by the National Key R&D Program(2016YFD0500707)Department of Education of Guangdong Province(YQ2015030)the Industry Technology System of Modern Agriculture Construction Fund of China(CARS-36)
文摘Hepatitis E is caused by hepatitis E virus(HEV),which has been classifi ed into four genotypes.Genotypes 3 and 4 are regarded as zoonotic pathogens.Accumulating researches indicate that genotype 4 is the main HEV strain circulating in China,and there are high levels of seropositive pigs and human in some provinces of China.In this study,serum samples from pigs and from human occupationally exposed to pigs were obtained from pig farms in Guangdong Province,in subtropical southern China,in order to investigate for the fi rst time the prevalence of anti-HEV immunoglobulin G(Ig G)in the region.Antibodies against HEV were detected by Enzyme-Linked Immunosorbent Assay(ELISA)using a commercially marketed kit.The results showed that high numbers of pigs(74/94;78.7%)and human(50/94;53.2%)from three pig farms in Guangdong Province were positive for anti-HEV Ig G.The correlation coeffi cient relating the prevalence in pigs and human on different farms was 0.920.The seropositive rate in males(human)was 48.8%(20/41)and that in females was 47.7%(9/19),which showed no statistically signifi cant difference.These data indicated that there was a high prevalence of anti-HEV antibodies in pigs and in human with occupational exposure to pigs.The risk of infection with HEV in both human and pigs in Guangdong Province appeared to be age-dependent,to a certain extent.This study provided basic data for further researches on HEV and was a reminder that more attention should be paid to HEV infection both in pigs and workers on pig farms in the study region.
基金Supported by Institutional Grant by Council of Scientific and Industrial Research-Indian Institute of Chemical Biology,Kolkata,No.MLP-118。
文摘BACKGROUND The recent rise in the incidence of hepatitis B virus(HBV)infections in a densely populated city of eastern India(“mixing vessel”of people of varied socioeconomic and immune status)prompted this study.Applying saliva on fingers for enumerating bank notes is a common practice in the Indian subcontinent.Paper notes may be a potential source of“horizontal”transmission of this virus,especially if there are cuts/bruises on the oral mucous membrane or skin.AIM To investigate whether paper currencies could be a plausible mode of horizontal transmission of HBV infection.METHODS Polymerase chain reactions(PCR)followed by nucleotide sequencing was done for the detection of HBV.Hepatitis B virus surface antigen enzyme-linked immunosorbent assay(HBsAg ELISA)was performed on all HBV deoxyribonucleic acid-positive samples to check the detectability of the virus.Atomic force microscopy(AFM)was carried out for visual confirmation of HBV particles in ultracentrifuged/immunoprecipitated samples from currency paper washings.RESULTS HBV-specific PCRs on pellets obtained after ultracentrifugation/immunoprecipitation of the currency paper washings detected potentially intact/viable HBV(genotype D2)in 7.14%of samples(n=70).AFM gave the visual confirmation of HBV particles in ultracentrifuged/immunoprecipitated samples from currency paper washings.However,HBV isolates from the currency notes could not be detected by HBsAg ELISA.CONCLUSION It is a common practice in the Indian subcontinent to count paper currencies by applying saliva on fingertips.Paper notes may be a potential source of“horizontal”transmission of this virus,especially if there are cuts/bruises on the oral mucous membrane or skin,but it was practically not possible to demonstrate experimentally such transmission.Detection of potentially intact/viable and“occult”HBV from currency poses potential risk of silent transmission of this virus among the general population.
文摘BACKGROUND Alpha-defensin has been widely studied for the diagnosis of periprosthetic joint infection(PJI).However,there is a lack of detailed information regarding the proper laboratory technique of the enzyme-linked immunosorbent assay(ELISA)method,such as sample dilution.AIM To assess the influence of dilution in the synovial fluid during ELISA for the diagnosis of knee PJI;and determine which dilution presents a better performance.METHODS Forty samples of synovial fluid from arthroplasty knees were included,17 in the infected group and 23 in the aseptic group,according to Musculoskeletal Infection Society criteria.Initially,five synovial fluid samples from each group were assessed for quantitative analysis of alpha-defensin using ELISA.Different dilution ratios(1:10,1:100,1:500,1:1000 and 1:5000)were tested based on the predetermined cutoff value of 5.2 mg/L.The dilutions that performed better were used to compare the results of all samples.RESULTS For infected cases,a gradual increase in the dilution of synovial fluid samples led to an equivalent increase in alpha-defensin level.The same was not observed in the aseptic cases.Both 1:1000 and 1:5000 dilutions presented satisfactory results to differentiate infected and aseptic cases.Further analyses were performed using 1:1000 and 1:5000 for all 40 samples.The 1:1000 dilution resulted in a sensitivity of 88.2%(95%CI,66%-98%)and specificity of 95.7%(95%CI,79%-99%),whereas the 1:5000 dilution presented a sensitivity of 94.1%(95%CI,73%-99%)and a specificity of 100%(95%CI,86%-100%).CONCLUSION The synovial fluid dilution had an important influence on the alpha-defensin ELISA results.Dilutions of 1:5000 showed the best performance for the diagnosis of knee PJI.The results of this study set the basis for a more reliable and reproducible alpha-defensin ELISA during the investigation of PJI,contributing to the expansion of this technique in different treatment centers worldwide.
文摘The glomerulonephritis disease is characterized by inflammation of glomeruli or small blood vessels in the kidney that causes kidney diseases. The reason of glomerulonephritis disease is to deposit the anti-GBM auto antibody in the glomerular basement membrane. The type IV collagen is the main component of glomerular basement membrane that has α3 chain of type (IV) collagen of non-collagenous domain which contains N-terminal 7S domain, a triple helical collagenous domain and C-terminal non-collagenous glomerular domain (NC1). The amino terminal of α3 (IV) NC1 that induces the Experimental Autoimmuno Glomerulonephritis (EAG) in rat model has been identified. The recombinant rat α3 (IV) NC1 antigen has nine amino acid spans that are consistent with antibody or T cell epitope that induces in EAG. The research is carried out on the recombinant rat α3 (IV) NC1 production, purification, quantification, and characterization. The circulation of anti-GBM antibody in glomerular basement membrane can be measured by the ELISA assay. In addition, the recombinant rat antigen is secreted in HEK293 cell supernatant that is purified by Anti-FLAG M2 monoclonal IgG antibody affinity column and characterized and quantified by SDS-PAGE gel electrophoresis and Western blotting techniques.
文摘Canine herpesvirus (CHV-1) causes disease associated with high mortality in infect-ed puppies, which represents large financial losses for dog breeders. Since CHV-1 at the time of the study he had not been reported in Mexico, the main objective of this study was to determine the prevalence of antibodies against CHV-1 in canine kennels in the metropolitan area of Mexico City. A commercial enzyme-linked immuno-sorbent assay (ELISA) was used, and the results were compared to those of a viral neutralization test. The ELISA kit uses the complete viral particle as the antigen. The plaque reduction neutralization test was combined with the immunoperoxidase technique because of the low cytopathic effect of CHV-1. Neutralizing antibodies were also detected in 20 randomly selected samples. The prevalence of CHV-1 with ELISA was 87%. The concordance between ELISA and serum neutralization (SN) was 0.1129, the sensitivity of the ELISA against SN was 1.0 (100%), the positive predic-tive value was 0.39 (39%), and the negative predictive value was 1 (100%). These results show that ELISA is useful for monitoring the dog population for CHV-1;a positive test result requires confirmation with an SN test, and a negative ELISA result indicates a high probability of being SN-negative. The only variables that were sta-tistically associated with CHV-1 prevalence were breed and kennel. A statistically significant relationship between the degree of ELISA and SN titer was obtained, with a confidence level of 95%. None of the clinical presentation factors was statistically significant. These results suggest that most of the canine population studied in Mex-ico is in a herpesvirus latency state.
基金supported by the National Key Research and Development Program of China(No.2021YFA0910900)the National Natural Science Foundation(No.22104147)+4 种基金Youth Innovation Promotion Association CAS(No.2021359)the Natural Science Foundation of Guangdong(Nos.2018B030306046 and 2020A1515111130)Guangdong Provincial Key Laboratory of Synthetic Genomics(No.2019B030301006)Shenzhen Science and Technology Program(No.KQTD20180413181837372)Shenzhen Outstanding Talents Training Fund.
文摘The global pandemic caused by severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)virus has necessitated rapid,easy-to-use,and accurate diagnostic methods to monitor the virus infection.Herein,a ratiometric fluorescence enzyme-linked immunosorbent assay(ELISA)was developed using Si-fluorescein isothiocyanate nanoparticles(FITC NPs)for detecting SARSCoV-2 nucleocapsid(N)protein.Si-FITC NPs were prepared by a one-pot hydrothermal method using 3-aminopropyl triethoxysilane(APTES)-FITC as the Si source.This method did not need post-modification and avoided the reduction in quantum yield and stability.The p-nitrophenyl(pNP)produced by the alkaline phosphatase(ALP)-mediated hydrolysis of pnitrophenyl phosphate(pNPP)could quench Si fluorescence in Si-FITC NPs via the inner filter effect.In ELISA,an immunocomplex was formed by the recognition of capture antibody/N protein/reporter antibody.ALP-linked secondary antibody bound to the reporter antibody and induced pNPP hydrolysis to specifically quench Si fluorescence in Si-FITC NPs.The change in fluorescence intensity ratio could be used for detecting N protein,with a wide linearity range(0.01-10.0 and 50-300 ng/mL)and low detection limit(0.002 ng/mL).The concentration of spiked SARS-CoV-2 N protein could be determined accurately in human serum.Moreover,this proposed method can accurately distinguish coronavirus disease 2019(COVID-19)and non-COVID-19 patient samples.Therefore,this simple,sensitive,and accurate method can be applied for the early diagnosis of SARS-CoV-2 virus infection.
基金This work was supported by Key Science and Technology Project in Henan Province(202102110061,202102110060,182102110423)High Level Talent Research Project of Henan Institute of Science and Technology(2016018,2016019).
文摘Anti-idiotypic antibodies which can mimic antigens have many potential applications in the immunoassay.This research used the monoclonal antibody of the conserved chemical region of pyrethroids,3-phenoxybenzoic acid(PBA),and the domain antibody library to develop an environmentally-friendly immunoassay for the detection of pyrethroids.The domain antibodies(A8,B8,and C6)which bound to anti-PBA monoclonal antibody(MAb)were isolated from a naive phage display human domain antibody library.This domain antibody is cloneable,pyrethroid-free,and applicable as a competitive mimetic antigen in the immunoassay.The best immunoassay was achieved using A8,resulting in IC50 of 0.714μg/mL for PBA,1.775μg/mL for Cypermethrin,1.624μg/mL forβ-cypermethrin,3.675μg/mL for Fenvalerate,and 4.895μg/mL for Flucythrinate.This way of selecting anti-idiotypic antibodies to detect pyrethroids could provide potential applications in developing immunoassays for identifying various chemical contaminants in food.
基金This work was supported by the National High-Tech Research and Development(863)Program of China,985 Project from Tsinghua University,and China Postdoctoral Science Foundation.
文摘This research describes a fast detection method on the basis of enzyme-linked immunosorbent assay(ELISA)for Escherichia coli in drainage of wastewater treatment plants.Optimized conditions such as the reaction format(sandwich or direct),the concentrations of diluted horse radish peroxidase(HRP)-E.coli conjugate,and anti-HPR antibody and pretreatment of E.coli were studied.Those results showed that the linear range of detection for E.coli was 10 cfu/mL-6×10^(4) cfu/mL.Compared with conventional methods,it is a convenient and sensitive detection method with low cost.
基金General Program of the National Natural Science Foundation of China:Regulatory Mechanism of Acupoint Catgut Embedding via Axonal Reflex on Neurogenic Inflammation in Allergic Rhinitis(No.81273985)General Program of the National Natural Science Foundation of China:the Study on the Mechanism of Acupoint Catgut Embedding Therapy for Allergic Rhinitis Based on the Coupling of"Acupoint Nasal Mucosal Sensory Nerve Dendritic Cells"(No.81473523)National Science and Technology Support Program for the Twelfth Five-Year Plan:the Study on Clinical Effect and Operation Standard of Catgut Embedding at Acupoints in Treating Allergic Rhinitis(No.2015BA I04B00)。
文摘OBJECTIVE:To investigate the effects of acupoint catgut embedding for 3 weeks on lung tissue,blood immunoglobulin E(IgE)and interleutin-4(IL-4),brain tissue microglia x-42(OX-42)and toll-like receptor-2(TLR-2)in rats with allergic rhinitis of lung deficiency type.METHODS:Forty-five female Sprague-Dawley rats were randomly divided for two times.The first time,they were randomly divided into model group and blank group(Group C)according to 2∶1,and the second time,the model group were randomly divided into model control group(Group B)and intervention treatment group(Group A)according to 1∶1.15 in each group.For Group A and Group B,the lung deficiency model was made by"sulfur–moxa fumigation",and then the allergic rhinitis model was established by"ovalbumin(OVA)sensitization".Then catgut embedding was performed at acupoints in Group A and not in Group B.After 3 weeks,collect lung tissue samples for hematoxylin-eosin staining,then take blood to observe the concentration of IgE and IL-4,and finally take brain tissue to observe the results of OX-42 and TLR-2.RESULTS:IgE level(μg/m L)was(3.11±0.20)in the Group A,(4.19±0.44)in the Group B,and(2.29±0.30)in the Group C(all P<0.001).IL-4 level(pg/mL)was(14.2±0.7)in the Group A,(18.6±2.4)in the Group B,and(11.4±1.2)for the Group C(all P<0.001).The mean OD for OX-42 is(0.1728±0.0016)in the Group A,(0.1810±0.0046)in the Group B and(0.1674±0.0025)in the Group C(all P<0.001).CONCLUSION:Although 3 weeks of acupoint catgut embedding already showed obvious efficacy on rats with allergic rhinitis,the allergic reaction in the body still continued.To achieve further treatment,prolonging the catgut embedding time is necessary.
