Swine Urine Samples Research Articles

Optimization, validation, and application of a liquid chromatography-tandem mass spectrometry method for the determination of 47 banned drug and related chemical residues in livestock urine using graphitized carboxyl multi-walled carbon nanotubes-based QuEChERS extraction

The establishment of an efficient method for the analysis of drug residues in animal urine facilitates the real-time monitoring of drugs used in the production of animal-derived food. A modified QuEChERS extraction–liquid chromatography-tandem mass spectrometry (LC–MS/MS) method was established for the determination of 47 banned drug and related chemical residues in livestock urine. The sample was extracted with acetonitrile by converting the acid-base environment. The sample cleanup effects of seven solid phase extraction cartridges and two EMR-Lipid products were compared, and three materials, including graphitized carboxyl multi-walled carbon nanotubes (MWCNTs), PSA, and C18, were selected as QuEChERS adsorbents from 24 materials. All analytes showed good linearity, with correlation coefficients (R2) greater than 0.9936. Low limits of quantification could be obtained, ranging from 0.2 to 5.5 ng/mL. The average recoveries at low, medium, and high spiked levels were in the range of 70.8–114.9 %, with intra-day precision ranging from 2.4 % to 11.2 % and inter-day precision ranging from 4.5 % to 16.1 %. Swine urine and bovine urine samples collected from different farms were effectively analyzed using the developed method, and metronidazole was detected in three swine urine samples.

Sensitive and simultaneous detection of ractopamine and salbutamol using multiplex lateral flow immunoassay based on polyethyleneimine-mediated SiO2@QDs nanocomposites: Comparison and application

The abuse of illegal additives poses a severe threat to human health. As such, it is of great significance to develop a simple, rapid and high-throughput analysis method for ensuring food safety from source to terminal. Herein, we proposed a multiplex fluorescent lateral flow immunoassay (LFIA) using the nanoassembly of silicon dioxide (SiO2) and quantum dots (QDs) as the advanced signal probe that can simultaneously detect ractopamine (RAC) and salbutamol (SAL) in swine urine and pork samples, and compared with other four conventional labels. The core–shell SiO2@QDs nanocomposites with strong luminescence, high stability, and abundant surface carboxyl groups were conjugated with antibodies to form high-performance SiO2@QDs immunoprobes, which improve the sensitivity and reliability of the LFIA. The developed SiO2@QDs-based LFIA achieved simultaneous detection of RAC and SAL in one strip within 13 min, with the limit of detection at 0.007 ng/mL and 0.032 ng/mL, respectively. The sensitivity of SiO2@QDs was superior to gold nanoparticles (AuNPs) and fluorescent microspheres (FMs), and exhibited comparative sensitivity with time-resolved microspheres (TRFMs), while overcoming the shortcomings of QDs that tend to aggregate. The recoveries in spiked swine urine and pork samples ranged from 73.7 to 111.0% with relative standard deviations between 4.7% and 15.5%. Good correlations were observed between the SiO2@QDs-based LFIA and commercial enzyme-linked immunosorbent assay for RAC (R2 = 0.936) and SAL (R2 = 0.979) detection. Briefly, the proposed method could be a promising screening on-site tool for convenient, rapid and sensitive detection of multiple illegal additives residues from the source to the terminal in food safety.

In vitro induced pluripotency from urine-derived cells in porcine.

BACKGROUNDThe generation of induced pluripotent stem cells (iPSC) has been a game-changer in translational and regenerative medicine; however, their large-scale applicability is still hampered by the scarcity of accessible, safe, and reproducible protocols. The porcine model is a large biomedical model that enables translational applications, including gene editing, long term in vivo and offspring analysis; therefore, suitable for both medicine and animal production.AIMTo reprogramme in vitro into pluripotency, and herein urine-derived cells (UDCs) were isolated from porcine urine.METHODSThe UDCs were reprogrammed in vitro using human or murine octamer-binding transcription factor 4 (OCT4), SRY-box2 (SOX2), Kruppel-like factor 4 (KLF4), and C-MYC, and cultured with basic fibroblast growth factor (bFGF) supplementation. To characterize the putative porcine iPSCs three clonal lineages were submitted to immunocytochemistry for alkaline phosphatase (AP), OCT4, SOX2, NANOG, TRA1 81 and SSEA 1 detection. Endogenous transcripts related to the pluripotency (OCT4, SOX2 and NANOG) were analyzed via reverse transcription quantitative real-time polymerase chain reaction in different time points during the culture, and all three lineages formed embryoid bodies (EBs) when cultured in suspension without bFGF supplementation.RESULTSThe UDCs were isolated from swine urine samples and when at passage 2 submitted to in vitro reprogramming. Colonies of putative iPSCs were obtained only from UDCs transduced with the murine factors (mOSKM), but not from human factors (hOSKM). Three clonal lineages were isolated and further cultured for at least 28 passages, all the lineages were positive for AP detection, the OCT4, SOX2, NANOG markers, albeit the immunocytochemical analysis also revealed heterogeneous phenotypic profiles among lineages and passages for NANOG and SSEA1, similar results were observed in the abundance of the endogenous transcripts related to pluripotent state. All the clonal lineages when cultured in suspension without bFGF were able to form EBs expressing ectoderm and mesoderm layers transcripts.CONCLUSIONFor the first time UDCs were isolated in the swine model and reprogrammed into a pluripotent-like state, enabling new numerous applications in both human or veterinary regenerative medicine.

An ultrasensitive colloidal gold immunosensor to simultaneously detect 12 beta (2)-adrenergic agonists

In this study, we first prepared a selective monoclonal antibody against 12 beta (2)-adrenergic agonists (Salbutamol, Clenbuterol, Brombuterol, Clenpenterol, Mabuterol, Carbuterol, Cimbuterol, Mapenterol, Pirbuterol, Terbutaline, Cimaterol, and Clenproperol). Then three haptens were designed and derived, among which, haptenS3 used the amino group of the salbutamol analog to derive a carboxyl group containing a spacer, which is unique to this study. The half-maximal inhibitory concentration (IC50) values were 0.35 ng/mL (Salbutamol), 0.42 ng/mL (Clenbuterol), 0.78 ng/mL (Brombuterol), 0.88 ng/mL (Clenpenterol), 1.34 ng/mL (Mabuterol), 1.38 ng/mL (Carbuterol), 1.71 ng/mL (Cimbuterol), 2.24 ng/mL (Mapenterol), 2.25 ng/mL (Pirbuterol), 2.27 ng/mL (Terbutaline), 3.49 ng/mL (Cimaterol), and 4.89 ng/mL (Clenproperol). We further developed a monoclonal antibody-based colloidal gold immunochromatographic test strip for screening and detecting 12 beta (2)-adrenergic agonists in swine urine and lamb samples. The immunochromatographic method developed in this study is a suitable tool for the on-site rapid detection and screening of beta (2)-adrenergic agonists in swine urine and lamb samples.

Influence of Selective Agents (EMJH-STAFF), Sample Filtration and pH on Leptospira interrogans Serovar Icterohaemorrhagiae Cultivation and Isolation from Swine Urine.

Leptospira spp. cause the zoonotic disease leptospirosis, which occurs in numerous mammalians worldwide. Isolation is still important for serotyping and genotyping of Leptospira, which in turn is essential for epidemiological surveillance of leptospirosis and the development of diagnostic tests and vaccines. However, isolation of Leptospira from clinical specimens is inherently insensitive. This study was conducted to examine the influence of selective agents, sample filtration, sample pH and the use of phosphate buffered saline (PBS) buffer for sample storage to improve the success of cultivation and isolation of Leptospira interrogans serovar Icterohaemorrhagiae from swine urine. EMJH (Ellinghausen McCullough, Johnson and Harris) medium including the selective agents sulfamethoxazole, trimethoprim, amphotericin, fosfomycin and 5-fluorouracil (STAFF) increased the success of Leptospira isolation from spiked swine urine samples. Sample filtration yielded only negative results. Isolation in EMJH-STAFF was successful from swine urine with a density as low as 104 Leptospira/mL, and urine with pH ≤ 7 impaired the cultivation rate. Cultivation and isolation were not improved by the addition of PBS to spiked urine samples prior to storage for 24 h at 4 °C. The results of the study demonstrate that cultivation and isolation of leptospires from swine urine can be improved by enhanced methods.

Fabrication of Fe3O4/ZIF-8 nanocomposite for simultaneous removal of copper and arsenic from water/soil/swine urine

In this study, a magnetic nanocomposite (denoted as FZ) was fabricated using Fe3O4and zeolitic imidazolate framework-8 (ZIF-8), based on a coprecipitation method. FZ could efficiently remove Cu2+(Cu(II)) and AsO2− (As(III)) ions simultaneously from water, soil, and swine urine samples through hydrogen bonding and electrostatic interactions. The Cu(II) and As(III) removal efficiencies of the optimal FZ sample increased gradually with time and reached 99.1% and 98.4%, respectively, in 180 min. The maximum adsorption capacities of FZ4 for Cu(II) and As(III) were determined to be 33.48 mg/g and 21.12 mg/g, respectively. Additionally, the FZ with a high saturation magnetization (49.8 emu/g) was easily recovered from aqueous solutions and soil samples. Furthermore, zebrafish experiments indicated that FZ possessed a high biosafety. Thus, this study introduces a promising method for treating water, soil, and swine urine samples contaminated with Cu(II) and As(III), and verifies that FZ is suitable for practical application.

A highly specific and sensitive indirect competitive monoclonal ELISA for the detection of brombuterol in animal feed, swine urine, pork and liver samples

ABSTRACT Brombuterol, a new β-adrenergic agonist to enhance animal growth and increase feeding efficiency, is forbidden as an additive in animal feed for livestock production due to its adverse effects on consumers. In this study, a highly specific and sensitive monoclonal antibody was obtained and an indirect competitive monoclonal ELISA was developed to detect brombuterol, with an IC50 value of 0.1 μg/kg (μg/L) and no cross-reactivity to other structurally related β-adrenergic agonists. The average recovery of brombuterol using the icELISA method ranged from 72.9% to 106.4% with the coefficient of variation lower than 18.9%, which was determined by analysing spiked animal feed, swine urine, pork and liver samples (n = 5). Finally, the icELISA gave results having a good correlation with those obtained by liquid chromatography-tandem mass spectrometry. These results demonstrated that the developed icELISA for the detection of brombuterol is highly specific, sensitive, and reliable, indicating good potential for use in the area of food safety to improve consumer protection.

An enzyme-free fluorometric nanoprobe for chloramphenicol based on signal amplification using graphene oxide sheets.

A sensitive and selective method for the determination of the antibiotic chloramphenicol (CAP) is described, which is based on double signal amplification and GO as an efficient fluorescence quencher. The nucleic acid probe is composedof three well-defined regions, viz. the signal probe I, the signal probe II, and the capture probe. The capture probe will bind to CAP specifically and the signal probes produce a significantfluorescence signal. One end of the signal probes is labeled with the fluorophore 6-carboxyfluorescein (FAM). The labeled probes can be adsorbed on graphene oxide (GO) via π-stacking interactions, upon which the green fluorescence of FAM (measured at excitation/emission wavelengths of 490/514nm) is quenched. On addition of CAP, the aptamer/CAP complexes are formed, and this leads to the restoration of fluorescence due to the removal of the probes from GO. The double signal probes, together with GO as quencher, improve the fluorescence signal significantly and lower the detection limit. Under optimized conditions, the assay works in the 20- to 200-ppb CAP concentration range and has a 0.3-ppb detection limit. It is also successfully applied to the determination of CAP in spiked swine urine samples. The recoveries from spiked swine urine samples are between 97.73 and 108.56%, and the repeatability (expressed as the RSD) is between 4.66 and 8.90%. Graphical abstract The constructed DNA probes form a stable structure and bind to chloramphenicol specifically. One end of signal probes was labeled with the fluorophore 6-carboxyfluorescein (FAM). The detection sensitivity of chloramphenicol was significantly enhanced by using double signal amplification, which was superior to the traditional methods. The quantities of CAP can be achieved by fluorescence increment.

Selective extraction and detection of β-agonists in swine urine for monitoring illegal use in livestock breeding

The illegal use of β-agonists often endangers animal-derived food safety. In this study, a selective detection method for β-agonists in swine urine was established via the combination of polymeric ionic liquid-molecularly imprinted graphene oxide-miniaturized pipette tip solid-phase extraction and high-performance liquid chromatography. It is worth noting that this method relied mainly on the designed adsorbent, which presented a rich adsorption mechanism, fast mass transfer rate, and high selectivity, and was successfully utilized in the selective extraction of β-agonists from swine urine samples. The proposed method has low LOD (0.20–0.56 ng/mL), high recovery (94.9–107.9%), and high reusability (4 times, 91.9–108.8%), which indicates its high potential as a selective, sensitive, accurate, and nonfatal method for monitoring the illegal use of β-agonists in the livestock breeding stage.

Ultrasonic‐assisted dispersive liquid–liquid microextraction based on a simple and green deep eutectic solvent for preconcentration of macrolides from swine urine samples

AbstractIn this study, a simple and green deep eutectic solvent based dispersive liquid‐liquid microextraction methodology had been proposed for the assessment of four macrolides in swine urine samples. Some important parameters including type and volume of extraction solvent and demulsifier solvent, sample pH, salt concentration affecting extraction efficiency were investigated. The quantitative recoveries were achieved at pH 9 and 20% (w/v) NaCl by using 0.3 mL of deep eutectic solvent consisting of choline chloride‐phenol (1:2, molar ratio) and 0.3 mL of tetrahydrofuran. Under optimal conditions, the linear ranges of four macrolides were 0.5–20.0 µg/L with correlation coefficient higher than 0.999. Limits of detection and quantification were in the ranges of 0.02–0.1 and 0.1–0.5 µg/L, respectively. The relative recoveries for macrolides in spiked swine urine samples ranged from 80 to 93% with relative standard deviations less than 6%.

Computer-Aided Design of Molecularly Imprinted Polymers for Simultaneous Detection of Clenbuterol and Its Metabolites.

Molecular imprinting technology (MIT) offers an effective technique for efficient separation and enrichment of specific analytes from complicated matrices and has been used for illicit veterinary drug detectionin recent years due to its high selectivity, good chemical stability, and simple preparation. The development of in silico-based approaches has enabled the simulation of molecularly imprinted polymers (MIPs) to facilitate the selection of imprinting conditions such as template, functional monomer, and the best suitable solvent. In this work, using density functional theory (DFT), the molecularly imprinted polymers of clenbuterol and its metabolites were designed by computer-aided at B3LYP/6-31 + G (d, p) level. Screening molecular imprinting components such as functional monomers, cross-linkers, and solvents has been achieved in the computational simulation considerations. The simulation results showed that methacrylic acid (MAA) is the best functional monomer; the optimal imprinting ratio for both clenbuterol (CLB) and its dummy template molecule of phenylephrine (PE) to functional monomer is 1:3, while the optimal imprinting ratio for the two dummy template molecules of CLB’s metabolites is 1:5. Choosin gethyleneglycol dimethacrylate (EDGMA) as a crosslinker and aprotic solvents could increase the selectivity of the molecularly imprinted system. Atoms in Molecules (AIM) topology analysis was applied to investigate the template-monomer complexes bonding situation and helped to explain the nature of the reaction in the imprinting process. These theoretical predictions were also verified by the experimental results and found to be in good agreement with the computational results. The computer-simulated imprinting process compensates for the lack of clarity in the mechanism of the molecular imprinting process, and provides an important reference and direction for developing better recognition pattern towards CLB and its metabolite analytes in swine urine samples at the same time.

Colorimetric detection of the β-agonist ractopamine in animal feed, tissue and urine samples using gold–silver alloy nanoparticles modified with sulfanilic acid

ABSTRACTA highly sensitive, selective and simple method was proposed for colorimetric detection of ractopamine on the basis of the interaction between ractopamine and sulfanilic acid-modified gold–silver alloy nanoparticles (AuAgNPs). The AuAgNPs were prepared by the reduction of HAuCl4 and AgNO3 with sodium citrate in aqueous medium and further modified by sulfanilic acid. The interaction of ractopamine with sulfanilic acid induced rapid aggregation of sulfanilic acid-modified AuAgNPs along with an optical colour change, leading to precise quantification which could be detected by absorptiometry. Under the optimum conditions, the absorbance ratio (A600/A435) of sulfanilic acid-modified AuAgNPs exhibited a linear relationship with the concentration of ractopamine in the range of 4.5–31.6 ng/mL. The detection limit of ractopamine was 1.5 ng/mL. The established novel colorimetric detection method showed high selectivity towards ractopamine. The method was successfully applied to detect ractopamine in spiked pork, swine feed and swine urine samples with excellent recoveries from 94.4% to 112.5%. These results demonstrated that the proposed new method has a good potential for practical applications.

Highly luminescent green-emitting Au nanocluster-based multiplex lateral flow immunoassay for ultrasensitive detection of clenbuterol and ractopamine

A multiplex lateral flow immunoassay sensor based on highly luminescent green-emitting Au nanoclusters (AuNCs-MLFIA sensor) was successfully established for the simultaneous and quantitative determination of clenbuterol (Clen) and ractopamine (RAC) in swine urine. The antigens of Clen and RAC were dispersed on a nitrocellulose membrane as two test lines, and the Au nanoclusters were synthesized from 6-aza-2-thiothymine and l-arginine to obtain highly green luminescence and ultra-small nanoparticles (Arg/ATT/AuNCs). Free carboxyl groups on Arg/ATT/AuNCs enabled conjugation with biomolecules to afford an indicator for the biosensor. The AuNCs-MLFIA sensor is based on the indirect competition assay and could successfully detect samples within 18 min without sample pretreatment, qualitative results can be obtained by visual inspection under a UV lamp. The limits of detection of Clen and RAC by the naked eye were both 0.25 μg L−1. In addition, the AuNCs-MLFIA sensor allowed quantitative detection combined with a portable fluorescence reader. The half-maximal inhibitory concentrations of Clen and RAC were 0.06 and 0.32 μg L−1, respectively, with detection limits of 0.003 and 0.023 μg L−1. Thirty blind-spiked swine urine samples were analyzed by the AuNCs-MLFIA sensor and liquid chromatography–tandem mass spectrometry, and the results of the two methods showed a significant correlation. The newly developed AuNCs-MLFIA sensor overcomes several limitations of conventional LFIA sensors, including their low sensitivity, limitation to quantify analytes, and single-analyte detection.

A fluorometric clenbuterol immunoassay based on the use of organic/inorganic hybrid nanoflowers modified with gold nanoclusters and artificial antigen.

Organic/inorganic hybrid nanoflowers were synthesized from calcium phosphate and protein modified fluorescent gold nanoclusters and antigens. These nanoflowers are shown to be well suited labels for bioassay because they fulfill the functions of biological recognition and signal output. A fluorometric immunoassay was developed that was combined with immunomagnetic separation. In the detection system, the red fluorescence of the supernatant (measured at excitation/emission wavelengths of 360/640nm) is found to be proportional to the clenbuterol (Clen) concentration after two immunomagnetic separations. The assay has a linear response in the 0.5μgL-1 to 40μgL-1 Clen concentration range, and 0.167μgL-1 limit of detection. This makes it well suited for food safety monitoring. The average recoveries from spiked samples range from 92.7 to 109.1% (intra-assay) and 101.2 to 125.7% (inter-assay) with relative standard deviations of <11.6%. Spiked swine urine samples were analyzed by this method, and the results correlated well with data obtained by LC-MS/MS. Graphical abstract Fluorescenthybrid nanoflowers were fabricated with gold nanoclusters (BSA-AuNCs) and antigens. A fluorometric immunoassay based on the use of such nanoflowers and based on immunomagnetic separation was developed to detect clenbuterol residues in swine urine with satisfactory recoveries and acceptable accuracy.

Matrix effect of swine urine on time-resolved fluorescent nanobeads and colloidal gold immunochromatographic assay

ABSTRACT Immunochromatographic assay (ICA) is an efficient analytical technique and rapid, convenient, easy-to-use, low-cost, and on-site detection method that has been widely used to evaluate food safety. However, an important issue to be addressed for this method is its matrix effect. In this work, time-resolved fluorescent nanobead ICA (TRFN-ICA) and colloidal gold ICA (CG-ICA) were developed to detect clenbuterol in swine urine. Under optimized working conditions, the limits of detection of TRFN-ICA and CG-ICA were 16 and 68 pg/mL, respectively. The matrix effect on TRFN-ICA and CG-ICA was assessed in 20 swine urine samples. Results indicated that the sensitivity of TRFN-ICA was better than that of CG-ICA, and the detection time of the former was shorter than that of the latter. The matrix effect on TRFN-ICA was more serious than that on CG-ICA.

A novel quartz crystal microbalance sensor array based on molecular imprinted polymers for simultaneous detection of clenbuterol and its metabolites

For the rapid and robust detection of both parent clenbuterol (CLB) and its metabolites in swine urine samples, a novel quartz crystal microbalance (QCM) sensor array for CLB detection based on molecularly imprinted polymers (MIPs) was developed in this investigation. At first, clenbuterol and the structural analogs of its metabolites, 4-Aminohippuric acid (AHA) and 4-hydroxymandelic acid (HMA), were chosen as molecular templates. Through computational molecular modeling, the optimum ratio between the functional monomer and molecular template was selected. The surface imprinting method was applied to modify QCM electrode surface to graft a thin MIP film. The grafting polymer was characterized by Fourier-transformed infrared spectrometry (FTIR) and atomic force microscopy (AFM), respectively. After then, an array system composed of three sensors was employed to test the responses with different solutions and the principal component analysis (PCA) was adopted to analyze the corresponding data. As a result, for the designed sensor to clenbuterol, a linear equation y=100.07x-722.96 (R2=0.9928) was found between the sensor frequency shift ΔF and negative logarithm of clenbuterol concentration (-lgC). The limitation of detection (LOD) was 3.0ng/mL, which is lower than the Codex Alimentarius Commission regulations residue limit 10μg/L. The corresponding data of the three template solutions were analyzed by PCA, obtaining 100% recognition. The result demonstrated the feasibility that the developed method could be applied to detect whether the livestock was feed with CLB nutrient redistribution agent by checking the urine samples.

Advantages of time-resolved fluorescent nanobeads compared with fluorescent submicrospheres, quantum dots, and colloidal gold as label in lateral flow assays for detection of ractopamine

Label selection is a critical factor for improving the sensitivity of lateral flow assay. Time-resolved fluorescent nanobeads, fluorescent submicrospheres, quantum dots, and colloidal gold-based lateral flow assay (TRFN-LFA, FM-LFA, QD-LFA, and CG-LFA) were first systematically compared for the quantitative detection of ractopamine in swine urine based on competitive format. The limits of detection (LOD) of TRFN-LFA, FM-LFA, QD-LFA, and CG-LFA were 7.2, 14.7, 23.6, and 40.1pg/mL in swine urine samples, respectively. The sensitivity of TRFN-LFA was highest. In the quantitative determination of ractopamine (RAC) in swine urine samples, TRFN-LFA exhibited a wide linear range of 5pg/mL to 2500pg/mL with a reliable coefficient of correlation (R2=0.9803). Relatively narrow linear ranges of 10–500pg/mL (FM-LFA) and 25–2500pg/mL (QD-LFA and CG-LFA) were acquired. Approximately 0.005µg of anti-RAC poly antibody (pAb) was used in each TRFN-LFA test strip, whereas 0.02, 0.054, and 0.15µg of pAb were used in each of the FM-LFA, QD-LFA, and CG-LFA test strips, respectively. In addition, TRFN-LFA required the least RAC-BSA antigens and exhibited the shortest detection time compared with the other lateral flow assays. Analysis of the RAC in swine urine samples showed that the result of TRFN-LFA was consistent with that of liquid chromatography-tandem mass spectrometry (LC-MS/MS) and a commercial enzyme-linked immunosorbent assay (ELISA) kit.

Development of molecular imprinted column-on line-two dimensional liquid chromatography for selective determination of clenbuterol residues in biological samples

A novel method coupling molecular imprinted monolithic column with two-dimensional liquid chromatography was developed and validated for the analysis of clenbuterol in pork liver and swine urine samples. The polymers were characterized by using Fourier transform infrared spectroscopy, nitrogen adsorption desorption analyses, frontal analysis and the adsorption of selectivity. The results indicated that the imprinted columns were well prepared and possessed high selectivity adsorption capacity. Subsequently, the MIMC-2D-LC (molecular imprinted monolithic column-two dimensional liquid chromatography) method was developed for the selective analysis of clenbuterol in practical samples. The accuracy ranged from 94.3% to 99.7% and from 93.7% to 99.6% for liver and urine, respectively. The relative standard deviation (RSD) of repeatability was lower than 8.6% for both analyses. The limit of detections was 16ng·mL−1 for liver and 25ng·mL−1 for urine, respectively. Compared with the reported methods, the disturbance of endogenous impurity could be avoided by the 2D-LC method.

A competitive luminol chemiluminescence immunosensor based on a microfluidic chip for the determination of ractopamine.

Herein, a competitive luminol chemiluminescence immunosensor based on a microfluidic chip was developed to detect ractopamine (RCT) both in phosphate buffer and swine urine samples. The immunosensor can provide a liner range of 0.5-40 ng/mL and a high sensitivity with a limit of detection of 0.97 ng/mL for RCT detection in swine urine. Good rates of recovery in negative swine urine samples were achieved over the RCT concentration ranging from 0.5 to 40 ng/mL. The proposed method offered a promising analytical scheme for the on-site determination of RCT.

Development of a Monoclonal Antibody-Based Immunochromatographic Assay Detecting Ractopamine Residues in Swine Urine

A lateral-flow immunochromatographic assay (LFIA) using monoclonal antibody was developed for the rapid detection of ractopamine residues in swine urine. The assay procedure could be accomplished within 5 min, and the results of this qualitative one-step assay were evaluated visually according to whether test lines appeared or not. When applied to the swine urines, the detection limit and the half maximal inhibitory concentration (IC50) of the lateral-flow immunochromatographic test strip under an optical density scanner were calculated to be 0.1 ± 0.013 and 0.71 ± 0.056 ng/mL, respectively. The cut-off level was observed with the naked eye of 1 ng/mL for detection of ractopamine residue in swine urine. Parallel analysis of swine urine samples with ractopamine showed comparable results obtained from the LFIA and LC-MS/MS. Therefore, the described lateral-flow test strip can be used as a reliable, rapid and cost-effective on-site screening technique for the determination of ractopamine residues in swine urine.