Sensitive Monoclonal Antibody Research Articles

Development of time-resolved fluorescence immunochromatography and ic-ELISA based on monoclonal antibodies specifically recognizing fumonisin B1

Fumonisins (FBs) are mycotoxins primarily synthesized by Fusarium moniliformis. Among these, FB1 exhibits not only high toxicity towards humans and animals but also carcinogenic properties. The global prevalence of FB1 contamination in cereals and related products, particularly maize, is alarmingly significant. Consequently, the accurate determination of FB1 levels in cereals holds immense importance. In this study, highly sensitive monoclonal antibodies specifically targeting FB1 were prepared and utilized for the establishment of a time-resolved fluorescence immunochromatographic assay (TRFIC) to detect FB1. The parameters of antibody labeling with time-resolved fluorescent microspheres were optimized. The detection time was significantly reduced to 6 min. The limits of detection (LOD) for corn, rice, and feed were determined as 0.496–0.844 μg/kg, and the quantification (LOQ) was 0.788–1.322 μg/kg. In addition, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) was successfully developed. Under optimized conditions, the half inhibitory concentration (IC50) value for FB1 was determined as 2.137 μg/L. A strong correlation between the results obtained from these two methods and HPLC-MS/MS analysis was observed in the same samples tested. In conclusion, both immunological methods developed in this work are highly suitable for rapid FB1 detection in real field samples.

Nanoparticles-based lateral flow strip for halofuginone ultrasensitive detection in chicken and beef

Halofuginone (HFG) is a widely used anti-coccidiosis drug in animal husbandry. There is an urgent need for a rapid detection method to detect HFG residues in chicken and beef to ensure human health. In this study, suitable HFG haptens were selected using computer simulation, and a highly sensitive monoclonal antibody (mAb) was developed with the 50%-inhibitory concentration (IC50) of 0.66 ng/mL. Then, gold nanoparticle-based lateral flow immunochromatographic (ICA) strips for HFG detection were produced. Under optimization conditions, the ICA strips exhibited outstanding performance with a visual limit of detection (vLOD) of 50 μg/kg and 5 μg/kg, and cut-off values of 100 μg/kg and 10 μg/kg, for chicken and beef samples, respectively. In addition, the reliability of strips was verified by enzyme linked immunosorbent assay (ELISA) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Therefore, the developed strip can be used as an effective tool for HFG residues screening in chicken and beef samples.

Cx1Mab-1: A Novel Anti-mouse CXCR1 Monoclonal Antibody for Flow Cytometry.

The C-X-C motif chemokine receptor-1 (CXCR1) is a rhodopsin-like G-protein-coupled receptor, expressed on the cell surface of immune cells and tumors. CXCR1 interacts with some C-X-C chemokines, such as CXCL6, CXCL7, and CXCL8/interleukin-8, which are produced by various cells. Since CXCR1 is involved in several diseases including tumors and diabetes mellitus, drugs targeting CXCR1 have been developed. Therefore, the development of sensitive monoclonal antibodies (mAbs) for CXCR1 has been desired for the diagnosis and treatment. This study established a novel anti-mouse CXCR1 (mCXCR1) mAb, Cx1Mab-1 (rat IgG1, kappa), using the Cell-Based Immunization and Screening method. Cx1Mab-1 reacted with mCXCR1-overexpressed Chinese hamster ovary-K1 (CHO/mCXCR1) and mCXCR1-overexpressed LN229 glioblastoma (LN229/mCXCR1) in flow cytometry. Cx1Mab-1 demonstrated a high binding affinity for CHO/mCXCR1 and LN229/mCXCR1 with a dissociation constant of 2.6 × 10-9 M and 2.1 × 10-8 M, respectively. Furthermore, Cx1Mab-1 could detect mCXCR1 by Western blot analysis. These results indicated that Cx1Mab-1 is useful for detecting mCXCR1, and provides a possibility for targeting mCXCR1-expressing cells in vivo experiments.

Abstract 1857: Development of monoclonal antibodies against CD44v specifically expressed in cancers

Abstract CD44 is a transmembrane glycoprotein that binds to the extracellular matrix including hyaluronic acid. Human CD44 has 19 exons, 10 of which are commonly present in all isoforms. CD44 standard isoform (CD44s) is composed of the common exons, and is expressed in many cell types. CD44 variants (CD44v) are produced by alternative splicing, in which the remaining nine variant exons are inserted into the CD44 standard isoform in various combinations. The CD44 isoforms have overlapping and unique functions. Both CD44s and CD44v harbor hyaluronan-binding motifs that promote interaction with the tumor microenvironment, which activate various signaling pathways in tumor cells. CD44v is mainly expressed in epithelial cells and is involved in cell adhesion, proliferation, and survival. Overexpression of CD44v is observed in many solid tumors and plays an important role in invasion, metastasis, and acquisition of stemness. In this study, we employed the Cell-Based Immunization and Screening method (CBIS method), and established highly sensitive anti-CD44 monoclonal antibodies (mAbs) using CD44v3-10-overexpressed cell lines as immunogens. After that, we determined the epitopes by enzyme-linked immunosorbent assay and investigated their applications. As a result, we successfully established eight mAbs against each CD44v (v3 to v10), which can be used for flow cytometry, western blotting, and immunohistochemistry. Furthermore, some of them exhibited cancer specificity against pancreatic cancers, colorectal cancers, gastric cancers, or oral cancers, indicating that established anti-CD44v mAbs are useful for the antibody therapies. Citation Format: Yukinari Kato, Hiroyuki Suzuki, Mika K. Kaneko. Development of monoclonal antibodies against CD44v specifically expressed in cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1857.

Development of a sensitive lateral flow immunoassay for the rapid detection of butralin in fruits

Butralin is a dinitrobenzene pesticide, has been widely used as a herbicide in many crops. Its high levels of residues threaten the ecological balance and human health. Therefore, establishment of a sensitive monoclonal antibody (mAb) for rapid detection of butralin residues is critical. In this study, special haptens were designed with good preservation of the characteristic structure of target compound. After mice immunization, a highly sensitive monoclonal antibody was produced. The half-maximum inhibitory concentration (IC50) was 1.936 ng/mL. And we applied it to develop a method of lateral flow immunoassay (LFIA), with vLOD value of the LFIA strip to be 10 ng/mL. In watermelon and pear samples, the cLOD values of the LFIA strip were 0.432 ng/mL and 0.426 ng/mL, respectively. The recoveries of the developed method ranged from 94.4% ± 2.1%–112.1% ± 3.4% and 89% ± 5.4%–103% ± 4.1% in watermelons and pears samples, with a CV of 2.9%–3.4% and 2.2%–5.2%, respectively. The results demonstrated that the developed method showed great prospect in the detection and monitoring of butralin in real food samples.

Development of a colloidal gold immunochromatographic strip for rapid detection of ribavirin in pig urine

ABSTRACT A specific and sensitive monoclonal antibody (mAb) for ribavirin (RBV) was generated by immunizing BALB/c mice with RBV – bovine serum albumin. Using the mAb, a highly sensitive indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and a colloidal gold immunochromatographic (CGI) test strip were developed for detecting RBV in pig urine. Under optimum assay conditions, IC50 was 0.08 ng/mL in 0.05 M phosphate-buffered saline and the recovery rate in real-world samples ranged between 77.3% and 99%. The cut-off limit of the CGI test strip for RBV was 5 ng/mL in pig urine. The cross-reactivity rate of the test strip to the tested antiviral drugs including acyclovir and moroxydine hydrochloride and the metabolites 1,2,4-triazole-3-carboxamide and 1-β-D-ribofuranosyl-1,2,4-triazole-3-carboxylic acid was below 0.1%. The results indicate that ic-ELISA and the CGI test strip have high sensitivity and specificity and that these methods are suitable for the rapid detection of RBV in pig urine.

Ultrasensitive detection of imatinib in human serum using a gold-based paper sensor

Imatinib is the tyrosine kinase inhibitor of choice for the treatment of chronic myeloid leukemia and gastrointestinal stromal tumors. However, imatinib has drawbacks such as drug resistance and significant differences in pharmacokinetics within patients. Therefore, a colloidal gold-based immunochromatographic assay (CG-IA) was developed for measuring and monitoring imatinib in human serum. An imatinib derivative containing carboxyl groups was used for the synthesis of the immunogen, and 4-(4-methyl-1-piperazinylmethyl) benzoic acid was selected as the hapten for the heterologous coating antigen. Next, a highly sensitive and specific monoclonal antibody (mAb), 2F7 was screened for the construction of a CG-IA, with an IC50 value of 0.091 ng/mL. For the qualification of imatinib in human serum, the visual limit of detection (vLOD) and cut-off values of the CG-IA were 2 and 20 ng/mL, respectively. For quantitative detection, the calculated LOD value of the CG-IA was 0.068 ng/mL, with a linearity range of 1.004 and 23.087 ng/mL. The recovery rate of spiked serum samples was between 88.24 % and 104.75 %. In addition, the concentration of imatinib in the serum samples from 10 patients was detected by CG-IA and revealed a good correlation with those from LC-MS/MS. These results indicated that the developed gold-based paper sensor could become an effective tool for the rapid monitoring of imatinib in human serum samples.

Quantitative Determination of Aflatoxin B1 in Maize and Feed by ELISA and Time-Resolved Fluorescent Immunoassay Based on Monoclonal Antibodies.

In this study, a highly sensitive monoclonal antibody (mAb) was developed for the detection of aflatoxin B1 (AFB1) in maize and feed. Additionally, indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and time-resolved fluorescence immunoassay assay (TRFICA) were established. Firstly, the hapten AFB1-CMO was synthesized and conjugated with carrier proteins to prepare the immunogen for mouse immunization. Subsequently, mAb was generated using the classical hybridoma technique. The lowest half-maximal inhibitory concentration (IC50) of ic-ELISA was 38.6 ng/kg with a linear range of 6.25-100 ng/kg. The limits of detections (LODs) were 6.58 ng/kg and 5.54 ng/kg in maize and feed, respectively, with the recoveries ranging from 72% to 94%. The TRFICA was developed with a significantly reduced detection time of only 21 min, from sample processing to reading. Additionally, the limits of detection (LODs) for maize and feed were determined to be 62.7 ng/kg and 121 ng/kg, respectively. The linear ranges were 100-4000 ng/kg, with the recoveries ranging from 90% to 98%. In conclusion, the development of AFB1 mAb and the establishment of ic-ELISA for high-throughput sample detection, as well as TRFICA for rapid detection presented robust tools for versatile AFB1 detection in different scenarios.

An immunochromatographic sensor for the detection of 5-fluorouracil in human plasma

This study prepared a colloidal gold-based immunochromatographic strip (CG-ICS) for the detection of 5-fluorouracil (5-FU) in human plasma, for therapeutic drug monitoring. Three haptens were synthesized with different carboxyl introductory positions and with different carbon chain lengths for the spacer arm, and were used to screen the coating antigens and immunogens. A highly sensitive and specific monoclonal antibody (mAb) 3C12 with an IC50 value of 4.359 ng/mL detected by ic-ELISA was produced for the construction of the strip. The visual limit of detection (vLOD) for semi-quantitative analysis of the CG-ICS was 50 ng/mL and the cut-off value was 2000 ng/mL. Furthermore, the calculated LOD (cLOD) value and linear range for quantitative detection were 1.464 ng/mL and 5.432–367.627 ng/mL, respectively. This detection required at most 10 min. In addition, the recovery rate of the spiked 5-FU in human plasma detected by the CG-ICS was 91.77 % to 94.04 %, which matched very well the results from LC-MS/MS. For real positive samples, the test results of the CG-ICS were consistent with those of LC-MS/MS. These findings suggest that the CG-ICS detection method is suitable for the rapid detection of 5-FU.

Development of a Sensitive Anti-Mouse CD39 Monoclonal Antibody (C39Mab-1) for Flow Cytometry and Western Blot Analyses.

CD39 is involved in adenosine metabolism by converting extracellular ATP to adenosine. As extracellular adenosine plays a critical role in the immune suppression of the tumor microenvironment, the inhibition of CD39 activity by monoclonal antibodies (mAbs) is one of the important strategies for tumor therapy. This study developed specific and sensitive mAbs for mouse CD39 (mCD39) using the Cell-Based Immunization and Screening method. The established anti-mCD39 mAb, C39Mab-1 (rat IgG2a, kappa), reacted with mCD39-overexpressed Chinese hamster ovary-K1 (CHO/mCD39) by flow cytometry. The kinetic analysis using flow cytometry indicated that the dissociation constant of C39Mab-1 for CHO/mCD39 was 7.3 × 10-9 M. Furthermore, C39Mab-1 detected the lysate of CHO/mCD39 by western blot analysis. These results indicated that C39Mab-1 is useful for the detection of mCD39 in many functional studies.

An immunochromatographic strip sensor for rapid and sensitive detection of candesartan, olmesartan medoxomil, and irbesartan in herbal beverages.

Sartans, as a class of antihypertensive drugs, pose a threat to human health when illegally added to herbal beverages. It is crucial to detect sartans in herbal beverages. We have developed a highly sensitive monoclonal antibody against candesartan (CAN), olmesartan medoxomil (OLM), and irbesartan (IRB), with 50% inhibitory concentrations (IC50) that were obtained via indirect enzyme-linked immunosorbent assay (ic-ELISA) as 0.178 ng mL-1, 0.185 ng mL-1, and 0.262 ng mL-1 against CAN, OLM, and IRB, respectively. Based on this monoclonal antibody, we developed a rapid screening method for CAN, OLM, and IRB in herbal beverage samples using an immunochromatographic assay (ICA) strip. Test for 15 minutes after simple and rapid sample pre-treatment and the results of this method can be obtained through naked eye observation. The detection limits (LODs) of the ICA strip for CAN, OLM, and IRB in herbal beverage samples are lower than 0.15 ng mL-1, and the results of the ICA strip and ic-ELISA are consistent in spiked samples and recovery experiments. Therefore, this method can quickly, efficiently, and reliably achieve high-throughput on-site rapid detection of illegally added CAN, OLM, and IRB in herbal beverages.

Gold nanoparticle-based immunochromatographic assay for rapid detection of imazalil.

Imazalil (IMZ) is a commonly used fungicide for controlling fungus in agriculture, leaving residual IMZ in crops that could be hazardous to human health. In this work, we designed IMZ haptens for mice immunization and prepared sensitive monoclonal antibody (mAb) against IMZ. The subtype of anti-IMZ mAb is IgG2a. It possessed a half inhibition concentration (IC50) of 0.95 ng mL-1 and showed no cross-reactivity against other chemicals in ic-ELISA. Taking advantage of the mAb, we developed a gold nanoparticle-based immunochromatographic assay (GICA) for the rapid detection of IMZ in grapes and tomatoes. The assay gave a visual limit of detection (vLOD) of 25 ng g-1 and cut-off value of 500 ng g-1 in both samples. According to the calibration curves, the calculated LOD were 4.12 ng g-1 and 4.70 ng g-1 in grapes and tomatoes, respectively. The recovery rates of IMZ ranged from 84.7% to 104.4% with variation coefficients (CVs) of 5.7-11.8% in spiked samples, indicating a potent practicability of the GICA. The whole GICA process took 30 min. Therefore, the developed assay can be used for on-site detection and quantitation of IMZ in grape and tomato samples.

Establishment and application of a gold nanoparticle-based immunochromatographic test strip for the detection of avian leukosis virus P27 antigen in egg white samples.

Avian leukemia is an infectious tumorous disease of chickens caused by subgroup A of the avian leukemia virus (ALV-A), which mainly causes long-term viremia, slow growth, immune suppression, decreased production performance, multi-tissue tumors, and even death. The infection rate of this disease is very high in chicken herds in China, causing huge economic losses to the poultry industry every year. We successfully expressed the specific antigen protein of ALV (P27) through recombinant protein technology and screened a pair of highly sensitive monoclonal antibodies (mAbs) through mouse immunity, cell fusion, and antibody pairing. Based on this pair of antibodies, we established a dual antibody sandwich ELISA and gold nanoparticle immunochromatographic strip (AuNP-ICS) detection method. In addition, the parameters of the dual antibody sandwich ELISA and AuNP-ICS were optimized under different reaction conditions, which resulted in the minimum detection limits of 0.2 ng mL-1 and 1.53 ng ml-1, respectively. Commonly available ELISA and AuNP-ICS products on the market were compared, and we found that our established immune rapid chromatography had higher sensitivity. This established AuNP-ICS had no cross-reactivity with Influenza A (H1N1), Influenza A (H9N2), respiratory syncytial virus (RSV), varicella-zoster virus (VZV), Listeria monocytogenes listeriolysin (LLO), and Staphylococcal enterotoxin SED or SEC. Finally, the established AuNP-ICS was used to analyze 35 egg samples, and the results showed 5 positive samples and 30 negative samples. The AuNP-ICS rapid detection method established by our group had good specificity, high sensitivity, and convenience, and could be applied to the clinical sample detection of ALV-A.

Preparation of a highly specific and sensitive monoclonal antibody against tilmicosin and its application in lateral flow immunoassay

To reduce the false positive results caused by cross reactivity of the antibodies with other structural analogues, it is crucial to prepare a high specificity and sensitivity antibody against target for developing an accurate immunoassay. In this study, tilmicosin (TM) was selected as a model molecule. Firstly, two-dimensional similarity, electrostatic potential energy, mulliken atomic charges and overlapping of different haptens with TM were calculated using Gaussian 09W and Discovery studio, and the newly designed TM-HS was selected as the optimal hapten. Furthermore, a monoclonal antibody (mAb 12C8) was produced with the half maximal inhibitory concentration (IC50) of 0.36 ng/mL, and negligible cross-reactivity (CR) with other antibiotics. Finally, a lateral flow immunoassay (LFA) for the detection of TM based on amorphous carbon nanoparticles (ACNPs) labeled mAb 12C8 was developed by the reflectance value under natural light. The recoveries of TM ranged from 83.18% to 103.25% with a coefficient of variation (CV) < 12.47%. The results showed that the cut-off value of TM in milk samples was 1 ng/mL, and the limits of detection (LODs) for chicken muscle, bovine muscle, porcine muscle and porcine liver samples were 5.23, 5.98, 6.85 and 7.31 μg/kg, respectively. In addition, 40 real samples were tested by the LFA, and the detection results were consisted with that of high-performance liquid chromatography-UV detector (HPLC–UV). Those results indicated that the developed LFA is an accurate and useful tool for on-site screening of TM in milk and animal tissues.

Novel hapten design, highly sensitive monoclonal antibody production, and immunoassay development for rapid screening of illegally added chloramphenicol in cosmetics

Hapten design and synthesis have been regarded as the key factor to generate high-quality antibodies. In the present study, a novel hapten of chloramphenicol was synthesized, characterized and compared with two conventional haptens. The new hapten generated mAb 4B5 showed higher sensitivity and titer than the other two haptens-based mAbs. The haptens synthesized with the structure of chloramphenicol base generated more sensitive antibodies than the hapten with chloramphenicol succinate, and the spacer arm linked to the phenyl group hapten elicited the strongest antibody response. After optimization, a direct competitive enzyme-linked immunosorbent assay (dcELISA) and a lateral flow immunoassay (LFIA), both based on the mAb 4B5, were developed. The dcELISA had a half maximum inhibition concentration of 0.23 ng/mL and the LFIA showed a cutoff value of 5–10 ng/mL. The LFIA was applied to detect illegally-added chloramphenicol samples in anti-acne cosmetics, five out of 19 samples were tested chloramphenicol containing within 10 min, which result was confirmed with the dcELISA and HPLC. The LFIA has an adequate sensitivity and can be used as a point of care diagnostic device for rapidly screening chloramphenicol in cosmetics.

A Sensitive and Specific Monoclonal Antibody Based Enzyme-Linked Immunosorbent Assay for the Rapid Detection of Pretilachlor in Grains and the Environment.

Pretilachlor is a chloroacetamide herbicide mainly used for weed and broadleaf weed control in rice, that is widely utilized in China. In order to detect the residue of pretilachlor in the environment and food, a highly sensitive and specific monoclonal antibody (mAb) against pretilachlor was prepared, and the half maximum inhibitory concentration (IC50) of the monoclonal antibody was validated to be 31.47 ± 2.35 μg/L. An indirect competitive ELISA (ic-ELISA) based on the antibody with a linear range of 6.25~100 μg/L was developed. The specificity of the antibody was explained by computer simulations and experimental validation. The mAb exhibited negligible cross-reactivity towards alachlor, acetochlor, propisochlor, butachlor, and metalaxyl, and the limits of detection (LOD) for pretilachlor in lake, rice, and soil samples were 4.83~5.23 μg/L. The recoveries of all samples were 78.3~91.3%. The reliability of the ic-ELISA method for residue detection of pretilachlor in the environment and grains was confirmed using high performance liquid chromatography.

Development and characterization of anti-G12V mutated K-Ras monoclonal antibodies using hybridoma technology

Background. Kirsten rat sarcoma oncogene, KRAS, is a gene that encodes for K-Ras protein, a membrane-anchored protein which is involved in intracellular signal transduction. Cells harbouring KRAS mutations have been reported to progress toward cancer development and several studies have suggested the importance of KRAS mutational screening prior to cancer treatment. However, currently available diagnostic methods are costly, time consuming and laborious. Thus, monoclonal antibodies are proposed as an alternative in K-Ras mutational testing. Objective. This study aims to generate and characterise anti-K-Ras monoclonal antibodies based on their specificity and sensitivity towards G12V-mutated K-Ras proteins, which is one of the most commonly mutated KRAS isoforms. Methods. Mice were first immunised with G12V-mutated K-Ras peptides and spleens were harvested. Hybridoma cells which secreted K-Ras-specific antibodies were generated by fusing splenocytes with X63-Ag 8.6539 myeloma cells. Hybridoma polyclonal wells secreting G12V-mutated K-Ras antibodies were subcloned into a single clone producing monoclonal antibodies. The specificity and sensitivity of monoclonal antibodies were evaluated by ELISA and the reactivity of monoclonal antibodies were tested using immunoblotting and immunocytochemistry. Results. The cross-reactivity results indicated that anti-G12V monoclonal antibodies developed in this study is highly specific to G12V mimotopes with a cross-reactivity of 4.2-16.7% towards wild-type and other mutated K-Ras isoforms. The limit of detection of this monoclonal antibody was determined as 3.28 μg/mL. It was also found to be reactive in immunocytochemical assays and native G12V-mutated K-Ras in immunoblotting. Conclusion. A highly specific and sensitive monoclonal antibody was successfully developed, characterised and applied to several assays such as indirect ELISA, Western Blot and immunocytochemistry. These data indicate the potential for this anti-G12V KRAS monoclonal antibody to be further developed for use in various research, diagnostic and therapeutic applications.

Development and application of monoclonal antibody-based dot-ELISA and colloidal gold immunochromatographic strip for rapid, specific, and sensitive detection of tomato brown rugose fruit virus

Tomato brown rugose fruit virus (ToBRFV) is an emerging tobamovirus that has become a great concern to tomato production industry. Due to the lack of resistant cultivars, precise detection of ToBRFV is essential to prevent the spread of ToBRFV. In this study, we produced highly sensitive and specific monoclonal antibodies against ToBRFV and established dot-enzyme-linked immunosorbent assay (dot-ELISA) and colloidal gold immunochromatographic strip (CGICS)-based methods for ToBRFV detection. These two methods could specifically detect ToBRFV without cross-reaction with seven tested tobamoviruses and three frequently occurring tomato-infecting viruses. Sensitivity analysis showed that the limit of detection of the established dot-ELISA and CGICS methods reached up to 1:6400 and 1:10,000 (w/v, g/mL) dilution of ToBRFV-infected tomato tissue, respectively. Further analyses using field-collected tomato foliar and fruit samples showed that the results obtained by dot-ELISA and CGICS were consistent with those obtained by reverse transcription polymerase chain reaction. The established methods here allow for specific, sensitive, and robust detection of ToBRFV, and will be helpful for precise monitoring and early warning of ToBRFV.

Immunochromatographic assay for rapid detection of flupyradifurone in grape, blueberry, and tomato samples

Flupyradifurone (FPF) is a neonicotinoid insecticide that effectively controls the spread of various pests. In this study, we established an immunochromatographic assay based on a highly specific and sensitive anti-FPF monoclonal antibody (mAb) to screen for FPF residues in grapes, blueberries, and tomatoes. The cut-off value for the immunochromatographic assay was 5 mg/kg for grapes and 10 mg/kg for blueberries and tomatoes. The calculated limit of detection of the immunochromatographic assay was 0.009 mg/kg, 0.033 mg/kg, and 0.040 mg/kg for grapes, blueberries, and tomatoes, respectively. The recovery rates of the immunochromatographic assay were 97.0–108.2 % in grape samples, 90.9–105.1 % in blueberry samples, and 94.0–103.7 % in tomato samples, and the detection results were highly consistent with LC-MS/MS results. Therefore, this immunochromatographic assay was an effective and rapid tool for screening for FPF in grapes, blueberries, and tomatoes.

Rapid and sensitive detection of cyhalofop-butyl in foods using a gold nanoparticle-based lateral-flow strip

In this study, we prepared a highly sensitive monoclonal antibody (mAb) against Cyhalofop-butyl (Cyh) by indirect competitive enzyme-linked immunosorbent assay (ic-ELISA). Based on this mAb, an immunochromatographic strip (ICA) was established to detect Cyh in corn and brown rice samples. Through screening different coating antigens and antibodies, the half-maximum inhibitory concentration (IC50) of the highly specific Cyh mAb 1A3 was 2.2 ng/mL, and the detection linear range (IC20-IC80) was 0.5–5.2 ng/mL. When Cyh in corn samples was detected by ICA, the detection limit of the T line elimination value was 100 μg/kg, and the detection limit for colorimetric value was 25 μg/kg. When the detection sample was brown rice, the detection limit of the T line elimination value was 100 μg/kg, and the detection limit for colorimetric value was 12 μg/kg. The results of recovery experiments for both ic-ELISA and LC-MS/MS were consistent with those of the test results when using the ICA. The ICA test strip met the requirements for on-site rapid detection of Cyh in corn and brown rice samples. Therefore, this method is suitable for real-time detection and monitoring of agricultural products as well as during plant growth, harvesting, transportation, storage, and sales, thereby guaranteeing the safety of consumers.