Rapid detection of tulathromycin in pure milk and honey with an immunochromatographic test strip

In this paper, we describe the development of a rapid, simple, specific, and sensitive immunochromatographic strip test based on an antibody–antigen reaction for detection of tulathromycin (TULA) in samples of 0.01 M PBS pH 7.4, pure milk and honey. The monoclonal antibodies specifically recognized the corresponding antigens produced in our laboratory. Under optimized conditions, the cut-off limits of the test strips for TULA in 0.01 M PBS pH 7.4 was found to be 2.5?ng/mL. Meanwhile, with simple preparation for detection in pure milk and honey samples, the cut-off limits for TULA were found to be 5 and 10?ng/mL, respectively. Each test can be evaluated in 5?min. In summary, this lateral-flow device provides a sensitive, effective, specific, and rapid method for detection of TULA residues in food matrix.     

Development of an immunochromatographic test strip for the detection of ochratoxin A in red wine

Humans and animals are exposed to mycotoxins naturally present in foods. Ochratoxin A (OTA), one of the most abundant and toxic mycotoxins, is ubiquitous in nature and harmful to humans and animals. In this study, we developed a rapid, simple, specific, and sensitive lateral-flow immunochromatographic assay for detection of OTA in 0.01?M phosphate-buffered saline solution (PBS) (pH 7.4) and in red wine. We produced sensitive and selective monoclonal antibody against OTA. Under optimized conditions, the cut-off limit of the test strip was 10?ng/mL OTA in 0.01?M PBS (pH 7.4) and in red wine. The results were obtained within 10 min. Therefore, our developed method is useful for the detection of OTA in red wine.     

Rapid detection of zearalenone and its metabolite in corn flour with the immunochromatographic test strip

Zearalenone (ZEN) is a mycotoxin produced by fungal species which are widespread in nature and commonly contaminate many cereal grains, such as wheat, maize, corn, barley, and other cereal grains. An immunochromatographic (ICA) test strip was developed for the rapid simultaneous detection of ZEN and its metabolite in corn flour samples. For the ICA test, antigen (ZEN-BSA) and goat anti-mouse IgG were, respectively, drawn on nitrocellulose membrane as control line and test line. Under the optimized condition, the cut-off limits of test strips for ZEN, α-ZAL, and β-ZAL were found to be 0.5?ng/mL in 0.01?M PBS and 50?ng/mL in corn flour samples, meanwhile α-ZOL, β-ZOL, and ZEA were found to be 1?ng/mL in 0.01?M PBS and 75?ng/mL in corn flour samples. Our data indicate that the ICA is sensitive, rapid, cost-effective, and specific on-site screening for ZEN and its metabolite.     

Rapid detection of aldicarb in cucumber with an immunochromatographic test strip

A rapid, simple, and sensitive immunochromatographic strip test has been developed to detect aldicarb (ALD) in 0.01 M phosphate buffer solution (PBS) (pH 7.4) and cucumber samples. The test was developed with a sensitive monoclonal antibody specific for ALD, generated by immunizing BALB/c mice with a well-characterized ALD – keyhole limpet hemocyanin conjugate, produced in our laboratory. Under the optimized condition, the cut-off limits of test strip for ALD were found to be 25?ng/mL in 0.01 M PBS (pH 7.4) and 100?ng/mL in cucumber samples, and the highest standard relative deviation was 9.05%. Both results were obtained within 5 min. The data indicate that the method is sensitive, rapid, and specific, so this immunochromatographic test strip has utility for the rapid high-throughput screening of the pesticide, especially in food samples.     

Development of an immunochromatographic strip for detection of acetamiprid in cucumber and apple samples

An immunochromatographic strip was developed for detection of acetamiprid (AC) and cross-reactivity with thiacloprid (TC) in cucumber and apple samples. This detection system was developed based on a sensitive and specific monoclonal antibody produced in our laboratory. Under optimized conditions, the cut-off limits of the test strip for AC were found to be 1 and 0.25?ng/mL for TC in phosphate-buffered saline. Meanwhile, with simple preparation for detection cucumber and apple samples, the cut-off limits for AC were found to be 5 and 2.5?ng/mL for TC in cucumber samples. The immunochromatographic assay also revealed AC cut-off values of 30 and 15?ng/mL in apple samples. Each test can be evaluated within 5?min. The data indicate that the method is sensitive, rapid, specific, and has the advantages of simplicity; therefore, this immunochromatographic assay is a useful screening method for detection of AC and TC residues in cucumber and apple samples.     

Development of ic-ELISA and an immunochromatographic strip assay for the detection of methylmercury

In this study, we developed an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and an immunochromatographic strip assay based on a monoclonal antibody (mAb) against methylmercury (MeHg) to detect the presence of MeHg in tap water. Under optimum conditions (pH 8.0, 0.8% NaCl, and 0.1% Tween 20), the 50% half maximal inhibitory concentration (IC₅₀) and limit of detection (LOD) were 16.64 and 2.03?ng/mL, respectively. The anti-MeHg mAb was speci?c to mercury with no cross-reactivity with other metal ions. The cut-off value of the immunochromatographic strip assay was 500?ng/mL for semi-quantitative detection, and the LOD was 11.3?ng/mL for quantitative detection. The average recovery rates of the ic-ELISA and immunochromatographic strip assay were 98.13% and 107.87%, respectively, in tap water. Therefore, ic-ELISA and the immunochromatographic strip assay can be used to detect MeHg in tap water.     

Rapid detection of clonidine and its cross-reactivity with apraclonidine in pig urine using an immunochromatographic test strip

Clonidine (CLO) preferentially stimulates central α-2-adrenoceptors and causes both desirable and undesirable effects. Undesirable effects include sedation and withdrawal reactions, which occur as a sudden rise in arterial pressure, nervousness, agitation, and increased heart rate. An immunochromatographic test strip was developed for the rapid simultaneous detection of CLO and its cross-reactivity with apraclonidine (ACLO) in pig urine. The antigen CLO-OVA and goat anti-mouse IgG were attached to a nitrocellulose membrane as control line and test line. The cut-off limits of the test strips for CLO and ACLO were found to be 2.5?ng/mL in both 0.01?M PBS (pH 7.4) and pig urine. All the results were obtained within 5 min. The results revealed that the developed method is a sensitive, rapid, and simple tool for the detection of CLO and ACLO.     

Development of a monoclonal antibody assay and immunochromatographic test strip for the detection of amikacin residues in milk and eggs

An amikacin-sensitive monoclonal antibody (MAb) assay and immunochromatographic test strip were developed and applied for the detection of amikacin residues in bovine milk and chicken eggs. The immunoassay was specific to amikacin and showed no cross-reactivity with other aminoglycosides. The half maximum inhibitory concentration (IC₅₀) of the assay was 0.65?ng/mL and the results were obtained within 90?min. Recoveries from spiked food matrices were within the range of 73.55–84.61% for bovine milk and 73.70–105.75% for whole egg. The strip test results were obtained within 10?min and showed a visual detection limit of 5.0?ng/mL for both food matrices. These results show that the MAb immunoassay and strip test developed in this study are very specific to amikacin and sufficiently sensitive for detection and routine monitoring of amikacin residues in food.     

Development of an immunochromatographic strip assay for ractopamine detection using an ultrasensitive monoclonal antibody

A sensitive gold nanoparticle immunochromatographic strip assay and indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) were developed for the detection of ractopamine (RCT), a beta-adrenergic agonist. The immunogen and coating antigen were synthesized by the carbodiimide method and conjugated with keyhole limpet hemocyanin and ovalbumin, respectively. The highly sensitive and specific monoclonal antibody was prepared for RCT, with a 50% inhibition concentration of 0.05?ng/ml and had no cross-reactivity with other beta-adrenergic agonists. An ultrasensitive and rapid immunochromatographic strip assay was developed with an RCT cutoff value of 2?ng/ml. Both developed methods can be used for RCT detection in swine urine.     

Development and validation of an immunochromatographic test strip for rapid detection of doxycycline residues in swine muscle and liver

A one-step lateral flow immunochromatographic technique using colloidal gold-labelled monoclonal antibody (Mab) was developed for the rapid detection of doxycycline (DOX) residues in swine tissues. For this purpose, a Mab against DOX named as 2.3/3A6 with low cross-reactivity (CR) to other tetracyclines (TCs) was produced. The sensitivity of the test strip was as low as 7 ng/mL for DOX and the half maximal inhibitory concentration (IC₅₀) was calculated to be 22±2 ng/mL by relative optical density. The test can be completed within 10 min and the detection limit was 20 ng/mL in unaided visual assessment. Recoveries in samples spiked with DOX at concentrations of 20, 40 and 80 ng/g, were demonstrated to be from 81% to 95% for muscle samples and from 81% to 92% for liver samples. The intra-assay and inter-assay coefficients of variation (CVs) ranged from 3% to 6% and from 4% to 8%, respectively. Comparing with high performance liquid chromatography (HPLC) method in sensitivity and accuracy for the detection of DOX in swine tissues, the test strip showed good agreement. Therefore, the test strip is suitable for rapid and reliable determination of DOX residues in swine tissues semi-quantitatively or qualitatively.     

Development of an ultrasensitive ic-ELISA and immunochromatographic strip assay for the simultaneous detection of florfenicol and thiamphenicol in eggs

An ultrasensitive monoclonal antibody-based gold nanoparticle immunochromatographic strip assay and indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) were developed to detect florfenicol (FF) and thiamphenicol (TAP) in egg samples. The ic-ELISA, with optimized pH, methanol content and sodium chloride content, exhibited an IC₅₀ value of 0.2?ng/mL for FF and 0.27?ng/mL for TAP, with the working range of 0.05–0.77 and 0.05–1.42?ng/mL, respectively. The optimized ic-ELISA showed negligible cross-reactivity with other phenols and broad-spectrum antibiotics. The recoveries in egg samples using the ic-ELISA ranged from 84% to 115% with a coefficient of variation of less than 5%. Based on this monoclonal antibody, a rapid and ultrasensitive immunochromatographic strip assay was developed with a cutoff value of 1?ng/mL for FF and TAP. Our results indicated that both developed methods were highly useful for screening FF and TAP in eggs.     

Development of a monoclonal antibody-based immunochromatographic strip for cephalexin

An anti-cephalexin (CEX) monoclonal antibody was prepared by the immunogen of CEX-keyhole limpet hemocyanin connected by disuccinimidyl suberate, which was used for the establishment of a rapid immunochromatographic test strip. With this semi-quantitative detection method, a low limit of deduction of 10 ng mL^?1 was obtained with the naked eyes and 1.3 ± 0.1 ng mL^?1 by a strip reader in unprocessed milk within 5 min, which was much lower than the European Union Maximum Residue Limit of 100 µg kg^?1 in milk. The recovery in the range of 87–120% was measured in another milk sample. In conclusion, this method with high sensitivity and satisfactory recovery shows significant potential for CEX residue detection in milk.     

Development of IC-ELISA and immunochromatographic strip assay for the detection of flunixin meglumine in milk

Flunixin meglumine (FM) is a novel nonsteroidal anti-inflammatory drug for animals, which has antipyretic, analgesic, and anti-inflammatory effects. The drug, which was originally used to relieve inflammation in horses, musculoskeletal disorders, and pain, has been approved for use in endotoxemia, infectious diseases in swine, etc. A sensitive anti-FM monoclonal antibody 2H4 was prepared and used to develop an indirect competitive enzyme-linked immunosorbent assay and immunochromatographic strip assay for the detection of FM in milk. The complete antigen and coating antigen were conjugated with bovine serum albumin and ovalbumin, respectively. The monoclonal antibody 2H4, with a half inhibition concentration of 0.29?ng/mL, had a limit of detection of 0.432?ng/mL and a linear range of detection of 0.08664–0.97226?ng/mL. A sensitive and convenient immunochromatographic strip assay was developed with an FM cutoff value of 0.29?ng/mL. The developed methods were suitable for the detection of FM in milk.     

Development of an ic-ELISA and immunochromatographic strip for detection of sparfloxacin in honey

In this study, a sensitive monoclonal antibody (mAb) 3D5 against sparfloxacin (SPFX) was generated. Based on the mAb 3D5, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) method was developed for the detection of SPFX in honey. The half maximal inhibitory concentration and limit of detection for the ic-ELISA method were 0.12 and 0.02?ng/mL, respectively. The experiments showed a negligible cross-reaction with other drugs. The average recovery rates for spiked SPFX honey extracts ranged from 90% to 101%, indicating an accepted accuracy. Furthermore, a lateral-flow immunochromatographic assay strip method with a cut-off value of 2?ng/mL was developed. Therefore, both of the developed methods are suitable for future use as rapid screening methods to detect and control the content of SPFX residues in honey samples.     

Development of indirect competitive enzyme-linked immunosorbent and immunochromatographic strip assays for carbofuran detection in fruits and vegetables

Carbofuran is a highly toxic pesticide used in fruits and vegetables. In this study, we produced a specific and sensitive monoclonal antibody (mAb) which was prepared based on a hapten that was derivatized with benzofuranol against carbofuran. Following mice immunization and cell fusion, we obtained three monoclonal cell lines: 6D5, 3H2, and 3C6. The cell line 3H2 generated mAb with the highest affinity and sensitivity. The half maximal inhibitory concentration was 0.3?ng/mL, and the cross-reactivity was <1%. Based on this mAb, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and immunochromatographic test strip (ICS) assay were developed to detect carbofuran residues in cucumbers and apples. The working range of ic-ELISA was 0.1–1?ng/mL, and the cutoff value of ICS was 1?ng/mL. The analytical recovery of carbofuran in cucumber and apple samples ranged from 81% to 97%. Both methods represent rapid screening tools for carbofuran detection in fruits and vegetables.     

胶体金免疫层析试纸条快速检测乙型流感病毒

目的建立有效、简便的胶体金免疫层析试纸条快速检测乙型流感病毒感染的方法。方法通过对乙型流感病毒核蛋白单克隆抗体进行胶体金标记,成功研制了乙型流感病毒免疫层析检测试纸条。结果该试纸条操作简单,肉眼于10～15 min内判定结果,对乙型流感病毒具有高度特异性,与甲型H1N1、H3N2亚型流感病毒等其他重要呼吸道病毒无交叉反应。试纸条在室温保存12个月、2～8℃保存18个月,其特异性和灵敏度无明显变化。对从内蒙自治区医院收集的流感样症状病人的702份鼻咽部分泌物进行检测,与美国Quidel公司同类产品的符合率为95%。结论建立的乙型流感病毒免疫层析检测方法具有简便、快速、特异、敏感和稳定等特点,对乙型流感病毒感染疾病的临床检测与早期诊断具有重要意义。     

Development of an antibody-based colloidal gold immunochromatographic lateral flow strip test for natamycin in milk and yoghurt samples

An immunochromatographic lateral flow strip test was developed for the detection of natamycin (Nata) residues in milk and cheese samples. Monoclonal antibody (mAb) against Nata was produced with a half maximal inhibitory concentration of 1.85?μg?L^?1. MAb conjugated with gold nanoparticles was the detection reagent. Nata conjugated with ovalbumin via the 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide method immobilized on a nitrocellulose membrane was the capture reagent. This semi-quantitative method required only 15?min to complete. The optimum concentrations of coating antigen and mAb were 0.15?µg?mL^?1 and 0.2?µg?mL^?1, respectively. Based on an optical density scanner, the visual limit of detection of Nata was 5?μg?L^?1 and 10?μg?kg^?1 in milk and yoghurt samples, respectively.     

Development of ic-ELISA and lateral-flow immunochromatographic assay strip for the detection of citrinin in cereals

A sensitive and specific anti-citrinin (anti-CIT) monoclonal antibody 1F2 was obtained following immunization and cell fusion. An indirect competitive enzyme-linked immunosorbent assay was developed with a 50% inhibitory concentration of 0.761?ng/mL and a limit of detection of 0.089?ng/mL. The recovery rates for CIT-spiked cereals (maize, wheat, and rice) ranged from 112% to 123%. A lateral-flow immunochromatographic assay was developed for both semi-quantitative and quantitative detection. With CIT-spiked cereals, the visual limit of detection was 8?ng/g and the cut-off value was 40?ng/g (semi-quantitative analysis with naked-eye detection). Using a strip scan reader, the calculated limit of detection was 1.28–1.8?ng/g for different CIT-spiked cereals. The recovery rates ranged from 110% to 127%. Therefore, both methods were effective for CIT detection and suitable for on-site detection and rapid screening of samples.     

Development of a monoclonal antibody assay and a lateral flow strip test for the detection of paromomycin residues in food matrices

A monoclonal antibody (MAb) was produced against paromomycin and used in the development of an immunoassay and a lateral flow strip test for the detection of paromomycin residues in food matrices. The MAb had 25.1% cross-reactivity with neomycin, but was insensitive to other aminoglycosides. Tested under optimized conditions in 0.01 M phosphate-buffered saline, the half maximum inhibitory concentration (IC₅₀) of the MAb was 0.61?ng/ml for paromomycin and 2.43?ng/ml for neomycin, with results obtained within 90?min. The mean recoveries from spiked food matrices were within the range of 64.56–105.85% for paromomycin and 54.08–100.55% for neomycin. The strip test results for different food matrices were obtained within 5?min and showed visual detection limits of 2.5–20?ng/ml (µg/kg) for paromomycin and 10–30?ng/ml for neomycin. Therefore, the developed immunoassay and strip test can be used in food analysis for routinely monitoring not only paromomycin but also neomycin residues.     

Development of a gold nanoparticle immunochromatographic assay for the on-site analysis of 6-benzylaminopurine residues in bean sprouts

The presence of pesticide residues in crops is a significant public concern. Due to the large number of samples and pesticides, a reliable, simple, cost-effective, and high-throughput analytic method is required. In this study, we developed a gold nanoparticle (GNP) immunochromatographic assay (ICA) for the rapid detection of 6-benzylaminopurine (6-BA) residues in bean sprouts. A monoclonal antibody against 6-BA, having a half-maximum inhibition concentration of 2.25?ng/mL, was labeled with GNPs and fixed onto a glass fiber membrane to create a conjugate pad. Coating antigen and goat anti-mouse antibody were sprayed onto a nitrocellulose membrane to form the test line and control line, respectively. The visual limit of detection and cutoff limit of ICA were 10 and 80?ng/g in bean sprouts, respectively. The results of ICA were obtained within 10 min with the naked eye. Therefore, ICA is a rapid tool for the on-site screening and detection of 6-BA in bean sprouts.