Gold immunochromatographic assay for simultaneous detection of carbofuran and triazophos in water samples

Using a simple test for rapid identification and quantification of pesticide multiresidues in food and environmental samples is a long-cherished approach for practical monitoring purposes. Here two gold-based lateral-flow strips (strip A and strip B) were investigated for simultaneous detection of carbofuran and triazophos. For the strip A format, a bispecific monoclonal antibody (BsMcAb) against both carbofuran and triazophos was employed to prepare the immunogold probe. For the strip B format, anti-carbofuran monoclonal antibody (McAb) and anti-triazophos McAb separately labeled with colloidal gold were combined as detector reagents. By comparison of visual results from pesticide standard tests between the two formats, the strip B assay manifested higher sensitivities for both pesticides. Analysis of spiked water samples by the preferable strip indicated that the detection limits for carbofuran and triazophos were 32 and 4 μg/L, respectively. The strength of the portable one-step strip assay was in the simultaneous screening for two pesticides within a short time (8-10 min) without any equipment.     

Rapid determination of hazardous compounds in food based on a competitive fluorescence microsphere immunoassay

Development of a microsphere-based competitive fluorescence immunoassay for the determination of hazardous low-molecular-weight compounds in food is described. In this method, antigens are covalently bound to carboxy-modified microspheres to compete monoclonal antibody with low-molecular-weight compounds in food samples; mouse IgG/fluorescein isothiocyanate conjugate is used as the fluorescent molecular probe. Thus, the hazardous low-molecular-weight compounds are quantified using a multiparameter flow cytometer. This method has been evaluated using clenbuterol as a model compound. It has a sensitivity of 0.01 ng/mL with dynamic range of 0.01-100 ng/mL, and the concentration of clenbuterol providing 50% inhibition (IC₅₀) is 1.1 ng/mL. The main advantages of this method are its high efficiency, biocompatibility, and selectivity, as well as ultralow trace sample consumption and low cost.     

三聚氰胺胶体金免疫层析试纸条的研制

通过胶体金免疫层析技术建立一种特异、便捷、快速的三聚氰胺抗原检测方法,对奶制品及饲料中的三聚氰胺残留水平监测提供参考。用柠檬酸三钠还原法制备的胶体金,标记纯化的三聚氰胺单克隆抗体,喷于试纸的金标垫。将MEL-OVA(三聚氰胺和卵清白蛋白的偶练物)和纯化的羊抗鼠IgG分别喷于试纸的T(检测线)处和C(质控线)处,通过挑选试纸条材料和调试工艺参数,并最终组装成试纸条。结果显示,制备的试纸监测体系方法检出限为50 g/L。试纸条对牛奶、奶粉和饲料中的三聚氰胺残留的检出限分别为100 g/L、100 ng/g和200 ng/g。将该法与LC-MS/MS法对比检测牛奶、奶粉和饲料样品,在试纸条检测范围内,与LC-MS/MS法检测结果一致性好,从而验证了该方法的有效性。制备的三聚氰胺胶体金检测试纸在常温干燥环境下至少可保质6个月,能够检测出三聚氰胺含量大于50 g/L的样品,适用于现场实际样品三聚氰胺残留水平监测,具有良好的应用前景。     

Rapid analytical method for the determination of 220 pesticide with their isomers by GCMS-TIC

This paper presents a cost-effective and validated multi residue modified and miniaturized method for the determination of 220 chemically different groups of pesticides and their isomers. This determination method is performed with single Quaid Gas Chromatography Mass Spectrometry -Total Ion Chromatogram GCMS-TIC. Two methods was experimented and modified with different GCMS parameters to analyses most common used pesticide and their residues in the standers solution and can be applied for real environmental samples. The results showed by single Quaid GCMS-TIC it can analyze 220 pesticides including their isomers within 49.6 min and low detection limit by using modified method 2 as described in this research. Limit of detection (LOD) was ranged from 0.78 to 14.74 ng/ml (ppb) with good separation and resolution. Limit of quantification (LOQ) was ranged between 2.34 and 44.22 ng/ml (ppb). Method 2 was more accurate, shorter, and clear separation rather than method 1. This method can be successfully applied in real environmental samples proven to be a good option for routine analysis of pesticide within the maximum residue limits (MRL) referenced to European commission especially with the most common GCMS-TIC which exists in most of labs and low income countries.     

Development of a direct competitive enzyme-linked immunosorbent assay for parathion residue in food samples

A heterologous direct competitive enzyme-linked immunosorbent assay (ELISA) for parathion residue determination is described based on a monoclonal antibody and a new competitor. The effects of several physicochemical factors, such as methanol concentration, ionic strength, pH value, and sample matrix, on the performance of the ELISA were optimized for the sake of obtaining a satisfactory assay sensitivity. Results showed that when the assay medium was in the optimized condition (phosphate buffer solution [PBS] containing 10% [v/v] methanol and 0.2 mol/L NaCl at a pH value of 5.0), the sensitivity (estimated as the IC₅₀ value) and the limit of detection (LOD, estimated as the IC₁₀ value) were 1.19 and 0.08 ng/ml, respectively. The precision investigation indicated that the intraassay precision values all were below 10% and that the interassay precision values ranged from 4.89 to 19.12%. In addition, the developed ELISA showed a good linear correlation (r² = 0.9962) to gas chromatography within the analyte’s concentration range of 0.1 to 16 ng/ml. When applied to the fortified samples (parathion adding level: 5-15 μg/kg), the developed ELISA presented mean recoveries of 127.46, 122.52, 91.92, 124.01, 129.72, 99.37, and 87.17% for tomato, cucumber, banana, apple, orange, pear, and sugarcane, respectively. Results indicated that the established ELISA is a potential tool for parathion residue determination.     

食品中农药残留的新型生物检测技术研究进展

食品安全是关系国计民生的重大战略问题。农药残留（简称农残）是造成食品质量与安全问题的重要因素。食品中农药残留不仅发生率高，而且超标严重，涉及范围广，长期接触或食用含有农药残留的食品会危害人体健康。传统的农残分析技术已经很难满足多样化食品安全检测的实际需求，现代新型生物技术因特异性强、灵敏度高、简单快速等优势，在食品的农残检测中得到广泛应用和持续发展。分析了食品中农药残留及检测技术现状，综述了酶联免疫吸附测定法、荧光免疫法和生物条形码免疫法等分析技术，及电化学、光学、压电传感器等生物传感技术在食品农残检测中的应用进展，讨论了各种技术的优势和局限，以期为相关研究提供参考，从而进一步提高我国食品农残的检测效率，保障食品安全，促进我国食品行业的可持续发展。     

Analysis of amphetamines and metabolites in urine with ultra performance liquid chromatography tandem mass spectrometry

A simple, rapid and sensitive ultra performance liquid chromatography tandem mass spectrometry method was developed and fully validated for the quantitative determination of seven amphetamines and metabolites in urine. The method was validated for selectivity, linearity, LOQ, LOD, imprecision, bias, analyte and processed sample stability, matrix effect, recovery, carryover and dilution integrity. A classic liquid–liquid extraction with ethyl acetate was used as sample preparation procedure. The compounds were separated on an Acquity UPLC HSS C₁₈ column in 6.8 min. The linear dynamic range was established from 25 to 500 ng/mL. The limit of quantification was fixed to the lowest calibrator level and the limit of detection ranged from 0.125 to 2.5 ng/mL. The method presented an excellent intra- and inter-assay imprecision and bias (<10.7%) at each measured concentration of two external quality controls (QC) and three “in house” QC. No matrix effects were observed and good recoveries (>70%) were obtained for all the compounds. No carryover was observed after the analysis of high concentrated samples (8000 ng/mL). The method was subsequently applied to authentic samples.     

Fluorescent competitive aptasensor for detection of aflatoxin B1

Aflatoxin B1 (AFB₁) is one of the most commonly found mycotoxins in food commodities, particularly cereals, oilseeds, spices and tree nuts. In the past decade, aptamers have come into limelight and emerged as a new biosensing element replacing antibodies in various detection formats. Herein we report a faster, more sensitive, high throughput method for the detection of AFB₁ using AFB₁‐specific aptamers. The assay format was based on a competitive reaction of the fluorescent tagged aptamer specific to AFB₁ with the aflatoxin conjugate. Under optimal conditions, a linear range of detection (50 ng to 50 pg) was achieved with a limit of detection (LOD) of 10 pg/mL in the buffer system. Results of inter‐ and intra‐assay revealed that the assay was repeatable with standard deviation in acceptable range. The assay was also validated in food samples such as dried red chilies, groundnut and whole pepper with recovery in the range of 92 to 102% at 10 ng/mL and 100 pg/mL levels. The aptasensor assay was also compared with standard analytical method of HPLC and was found to be more sensitive. This detection technique has the potential to be developed into a biosensor platform for AFB₁ detection.     

An aptamer-based chromatographic strip assay for sensitive toxin semi-quantitative detection

An aptamer-based chromatographic strip assay method for rapid toxin detection was developed. The aptamer-based strip assay was based on the competition for the aptamer between ochratoxin A and DNA probes. The sensing results indicated that the sensitivity of the aptamer-based strip was better than that of conventional antibody-based strips. The visual limit of detection of the strip for qualitative detection was 1 ng/mL while the LOD for semi-quantitative detection could down to 0.18 ng/mL by using scanning reader. The recoveries of test samples were from 96% to 110%. All detections could be achieved in less than 10 min, indicating that the aptamer-based strip could be a potential useful tool for rapid on-site detections.     

抗吡虫啉单克隆抗体的制备及鉴定

为制备灵敏度高,特异性强的抗吡虫啉单克隆抗体,建立经济、快速、准确的吡虫啉残留免疫学分析方法,采用分子模拟技术分析吡虫啉及其类似农药的电荷分布后,选择1-[6-(2-羧乙硫基-3-吡啶)甲基]-N-硝基-2-咪唑啉亚胺 (H1) 作为免疫半抗原,1-(6-氯-3-吡啶甲基)-3-羧丙基-N-硝基-2-咪唑啉亚胺 (H2) 作为包被半抗原,利用NHS酯法将H1和H2分别与牛血清蛋白 (BSA) 和卵清蛋白 (OVA) 偶联合成免疫原与包被原。免疫BALB/c小鼠后,采用常规杂交瘤技术共获得2株稳定分泌抗吡虫     

Development of a one-step immunochromatographic strip test using gold nanoparticles for the rapid detection of Salmonella typhi in human serum

An immunochromatographic strip test using gold nanoparticles was developed for the rapid detection of Salmonella typhi (S. typhi) in human serum. The strip test based on the principle of sandwich immunoassay by the specific binding of antigens from S. typhi O901 and antibody of S. typhi O901 on a nitrocellulose membrane. Antibody-gold nanoparticle conjugate was used as the label and was coated onto a glass fiber membrane, which was used as a conjugate pad. To create a test and control zone, antibody of S. typhi O901 and an anti-IgG were dotted on the nitrocellulose membrane, respectively. Positive samples were displayed as red dots at the test and control zones of the nitrocellulose membrane, while negative samples resulted in a red dot only in the control zone. The limit of detection (LOD) was found to be 1.14×10(5) cfu mL(-1), which could be visually detected by the naked eye within 15 min. This strip test provided a lower detection limit and analysis time than a dot blot immunoassay (8.88×10(6) cfu mL(-1) for LOD and 110 min for reaction time). In addition, our immunochromatographic strip test was employed to detect S. typhi in human serum effectively, with high accuracy. This strip test offers great promise for a rapid, simple and low-cost analysis of S. typhi.     

Application of one-step liquid chromatography–electrospray tandem MS/MS and collision-induced dissociation to quantification of ezetimibe and identification of its glucuronated metabolite in human serum: A pharmacokinetic study

A new one-step liquid chromatography–electrospray tandem MS/MS method is described to quantify ezetimibe (EZM) a novel lipid lowering drug in human serum. Also using collision-induced dissociation (CID) of the analyte, identification and chromatographic separation of its major metabolite, ezetimibe glucuronide (EZM-G) is achieved in this study. A thawed serum aliquot of 100 μL was deproteinated by addition of 500 μL methanol containing omeprazole as internal standard (I.S.). Separation of the drug, its metabolite and the I.S. were achieved using acetonitrile–water (70:30, v/v) as mobile phase at flow rate of 0.5 mL/min on a MZ PerfectSil target C18 column. Multiple reaction monitoring (MRM) mode of precursor–product ion transition (408.7 → 272.0 for EZM and 345 → 194.5 for the I.S.) was applied for detection and quantification of the drug while, EZM-G was chromatographically separated and identified using CID. The analytical method was linear over the concentration range of 1–32 ng/mL of EZM in human serum with a limit of quantification of 1 ng/mL. The coefficient variation values of both inter- and intra-day analysis were less than 8% whereas the percentage error was less than 3.7. The validated method was applied in a randomized cross-over bioequivalence study of two different EZM preparations in 24 healthy volunteers.     

农药残留检测关键用酶固定化研究进展

为保障消费者食用安全,迫切需要研发农产品和食品中的农药残留快速检测技术.酶抑制法检测是目前农药残留快速检测技术中的主要研究方向之一,而酶的固定化是用基于酶抑制法原理对农药残留检测研究中的重要步骤.通过物理或化学的方法高效地将酶固定于载体上,同时保持酶的催化活性是开发各类基于酶抑制法检测农药残留传感器的关键.本文将从固定...     

Determination of deoxynivalenol in organic and conventional food and feed by sol–gel immunoaffinity chromatography and HPLC–UV detection

The paper describes the determination of deoxynivalenol (DON) in 55 wheat food and feed samples, 26 from conventional and 29 from organic production. Immunoaffinity columns prepared by entrapping anti-DON antibodies by the sol–gel method were used for sample clean-up. DON was quantified by high performance liquid chromatography (HPLC) and ultraviolet (UV) detection. In general, the incidence of DON contamination was rather low. In eight samples (14.5%) the DON concentration was above the LOQ (380 ng/g), in six samples (10.9%) DON was detected but could not be quantified (>LOD (200 ng/g), <LOQ). In seven conventional samples (two pasta, two cookie, two snack and one feed sample) but only in one organic sample (a snack) the DON concentration was >LOQ. The data indicate both a higher incidence of DON contamination and higher DON concentrations in food and feed samples from conventional than in those from organic production.     

Occurrence and risk assessment of organophosphorus pesticide residues in Chinese tea

Tea is the second widely consumed beverage next to water. Tea drinking is one of the important pathways for human exposure of organonphosphorus pesticide. Consequently, incidence of organonphosphorus pesticide residues and risk assessment should be clear. In this study, the level of organonphosphorus pesticide residues in 810 Chinese teas manufactured between 2010–2013 was investigated using gas chromatography coupled with tandem mass spectrometry and a flame photometric detector. Incidence of organonphosphorus pesticide residues occurred with a frequency of 29% and the average concentration of 93 μg kg^?1. The residue levels varied from tea types, sale spots, and production area. Chlorpyrifos, isocarbophos, and triazophos were the only three organonphosphorus pesticides with detectable residues, and the detectable rates were 13.0%, 13.6%, and 17.4%, respectively. The corresponding average daily intake of chlorpyrifos, isocarbophos, and triazophos by tea drinking was 0.000083 μg kg^?1 bw day^?1, 0.0036 μg kg^?1 bw day^?1, and 0.0022 μg kg^?1 bw day^?1. These results showed that the total hazard quotient of organonphosphorus pesticide pesticides from tea drinking was less than 0.02 and that the tea-drinking originated organonphosphorus pesticide exposure had a little adverse health effect for human being.     

A coordination polymer nanobelt (CPNB)-based aptasensor for sulfadimethoxine

A polymer-based aptasensor, which consisted of fluorescein amidite (FAM)-modified aptamers and coordination polymer nanobelts (CPNBs), was developed utilizing the fluorescence quenching effect to detect sulfadimethoxine residue in food products. A single-stranded DNA (ssDNA) aptamer, which was a specific bio-probe for sulfadimethoxine (Su13; 5'-GAGGGCAACGAGTGTTTATAGA-3'), was discovered by a magnetic bead-based systematic evolution of ligands by exponential enrichment (SELEX) technique, and the fluorescent quenchers CPNBs were produced by mixing AgNO(3) and 4,4'-bipyridine. This aptasensor easily and sensitively detected sulfadimethoxine in solution with a limit of detection (LOD) of 10ng/mL. Furthermore, the antibiotic dissolved in milk was also effectively detected with the same LOD value. In addition, this aptamer probe offered high specificity for sulfadimethoxine compared to other antibiotics. These valuable results provide ample evidence that the CPNB-based aptasensor can be used to quantify sulfadimethoxine residue in food products.     

Simultaneous determination of aminomethylbenzoic acid,cefminox sodium and etamsylate in human urine by capillary electrophoresis

A sensitive and selective capillary electrophoresis method is developed, for the first time, for effective separation and simultaneous determination of aminomethylbezoic acid (PAMBA), cefminox sodium (CMNX) and etamsylate (ETM). The electrophoresis conditions were investigated and optimized. A 25 mM phosphate solution (pH 8.5) was used as a buffer and the peak area was determined with UV detection at 216 nm wavelength under 18 kV separation voltage. Under optimal conditions, the three drugs can be separated effectively. Good linearity was achieved in 3.13–150 μg/mL for PAMBA, 6.25–150 μg/mL for CMNX and 3.13–150 μg/mL for ETM, with the correlation coefficients of >0.999. The limit of detection (LOD) for PAMBA, CMNX and ETM was 1.04, 2.08 and 1.04 μg/mL, respectively. Their recoveries in human urine were in the range from 90.2% to 101% with the RSD (n = 5) of 0.7–3.1%. The proposed method is simple, rapid and accurate, and provides the sensitivity and linearity necessary for analysis of the test drugs in human urine at clinically relevant concentrations.     

Rapid detection of glycyrrhizin by immunochromatographic assay

An immunochromatographic assay was developed for detecting glycyrrhizin (1). The qualitative assay is based on a competitive immunoassay using anti-1 monoclonal antibody (MAb) and a detector reagent that contains colloidal gold particles coated with anti-1 MAb. The immunochromatographic strip test, which has a detection limit of 250 ng/mL, is useful as a rapid screening method for detecting glycyrrhizin in plants, biological fluids and food samples.     

Monoclonal antibodies labeled with colloidal gold for immunochromatographic express analysis of diphtheria toxin

A one-step immunochromatographic method for the detection of the diphtheria toxin in different water samples (phosphate buffer, milk, and human nasopharyngeal swab) was developed using a conjugate of monoclonal antibodies labeled with colloidal gold. The detection limit of diphtheria toxin was 10 ng/ml, the time of the analysis was 15 min. The use of silver to enhance the reaction sensitivity and scanning equipment to evaluate the immunochromatography results decreased the detection limit to 1.25 ng/ml.     

Development of immunochromatography strip-test using nanocolloidal gold-antibody probe for the rapid detection of aflatoxin B1 in grain and feed samples

An immunochromatography (ICG) strip test using a nanocolloidal gold-antibody probe was developed and optimized for the rapid detection of aflatoxin B1 (AFB1). A monoclonal antibody specific to AFB1 was produced from the cloned hybridoma cell (AF78), coupled with nanocolloidal gold, and distributed on the conjugate pad of the ICG strip test. The visual detection limit of the ICG strip test was 0.5 ng/ml, and this method showed a cross-reaction to aflatoxin B2, G1, and G2. In total, 172 grain and feed samples were collected and analyzed by both the ICG strip test and HPLC. The results of the ICG strip test showed a good agreement with those obtained by HPLC. These results indicated that the ICG strip test has a potential use as a rapid and cost-effective screening tool for the determination of AFB1 in real samples and could be applied to the preliminary screening of mycotoxin in food and agricultural products, generating results within 15 min without complicated steps.