Highly sensitive multianalyte immunochromatographic test strip for rapid chemiluminescent detection of ractopamine and salbutamol.

G Model ACA 233268 No. of Pages 6

Analytica Chimica Acta xxx (2014) xxx–xxx

Contents lists available at ScienceDirect

Analytica Chimica Acta journal homepage: www.elsevier.com/locate/aca

Highly sensitive multianalyte immunochromatographic test strip for rapid chemiluminescent detection of ractopamine and salbutamol Hongfei Gao, Jing Han, Shijia Yang, Zhenxing Wang, Lin Wang, Zhifeng Fu * Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Southwest University), Ministry of Education, College of Pharmaceutical Sciences, Southwest University, Chongqing 400716, China

H I G H L I G H T S

G R A P H I C A L A B S T R A C T

An immunochromatographic test strip was developed for detection of multiple b2-agonists. The whole assay process can be completed within 20 min. The proposed method shows much higher sensitivity due to the application of CL detection. It is a portable analytical tool suitable for field analysis and rapid screening.

A multianalyte immunochromatographic test strip was developed for the rapid detection of two b2agonists. Due to the application of chemiluminescent detection, this quantitative method shows much higher sensitivity.

A R T I C L E I N F O

A B S T R A C T

Article history: Received 2 March 2014 Received in revised form 12 May 2014 Accepted 16 May 2014 Available online xxx

A novel immunochromatographic assay (ICA) was proposed for rapid and multiple assay of b2-agonists, by utilizing ractopamine (RAC) and salbutamol (SAL) as the models. Owing to the introduction of chemiluminescent (CL) approach, the proposed protocol shows much higher sensitivity. In this work, the described ICA was based on a competitive format, and horseradish peroxidase-tagged antibodies were used as highly sensitive CL probes. Quantitative analysis of b2-agonists was achieved by recording the CL signals of the probes captured on the two test zones of the nitrocellulose membrane. Under the optimum conditions, RAC and SAL could be detected within the linear ranges of 0.50–40 and 0.10–50 ng mL 1, with the detection limits of 0.20 and 0.040 ng mL 1 (S/N = 3), respectively. The whole process for multianalyte immunoassay of RAC and SAL can be completed within 20 min. Furthermore, the test strip was validated with spiked swine urine samples and the results showed that this method was reliable in measuring b2agonists in swine urine. This CL-based multianalyte test strip shows a series of advantages such as high sensitivity, ideal selectivity, simple manipulation, high assay efficiency and low cost. Thus, it opens up new pathway for rapid screening and field analysis, and shows a promising prospect in food safety. ã 2014 Elsevier B.V. All rights reserved.

Keywords: Lean meat agent b2-agonist Multianalyte immunoassay Chemiluminescence Immunochromatographic test strip

1. Introduction

b2-agonists are a group of synthetic agents which were found to promote animal growth and increase feeding efficiency by reducing

* Corresponding author at: Southwest Univesity, College of Pharmaceutical Sciences, No. 2 Tiansheng Road, Beibei, Chongqing 400716, China. Tel.: +86 23 6825 0184; fax: +86 23 6825 1048. E-mail address: [email protected] (Z. Fu).

fat deposition and enhancing protein accretion [1]. In recent years, they have been frequently reported to be abused illegally in livestock production as “lean meat agents”. It has demonstrated that longterm or high-dose use of “lean meat agents” such as salbutamol (SAL), clenbuterol, terbutaline, and ractopamine (RAC) is hazardous to human health [2]. Thus b2-agonists have been rigorously forbidden to be used as feed additives for growth promotion in China, European Union and many other countries [3]. Thus, it is urgently needed to develop a simple, rapid, sensitive and low-cost screening technology for multianalyte detection of b2-agonists.

http://dx.doi.org/10.1016/j.aca.2014.05.024 0003-2670/ ã 2014 Elsevier B.V. All rights reserved.

Please cite this article in press as: H. Gao, et al., Highly sensitive multianalyte immunochromatographic test strip for rapid chemiluminescent detection of ractopamine and salbutamol, Anal. Chim. Acta (2014), http://dx.doi.org/10.1016/j.aca.2014.05.024


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H. Gao et al. / Analytica Chimica Acta xxx (2014) xxx–xxx

There are a variety of chromatographic methods reported for b2agonists detection, such as HPLC [4,5], GC–MS [6,7], LC–MS [8,9], LCchemiluminescent (CL) [5] and capillary electrophoresis (CE) [10– 12]. Whereas these methods are not suitable for field analysis and rapid screening, since they inherently rely on sophisticated and large instruments, time-consuming sample pretreatments, and complicated manipulations. More recently, a number of immunoassay methods that are especially fit for rapid screening have already been developed to detect various b2-agonists, including immunoassay based on ELISA [13,14], electrochemistry [15,16], surface plasmon resonance [17,18], fluorescence [19,20], CL [21] and surfaceenhanced Raman scattering [22]. However, despite the high sensitivity, ideal selectivity and low cost that can be achieved by these methods, they require multiple incubation, separation and washing steps, and are mostly designed for single-analyte assay. The immunochromatographic assay (ICA) is a membrane-based immunoassay method rising in recent years, which combines thin layer chromatography with conventional immunoassay to provide a novel analytical approach [23–34]. The principle of ICA is based on the migration of a sample solution along the antibodyimmobilized test strip where the corresponding immune recognition reaction takes place. The assay procedure could be significantly simplified for lack of a long incubation time and multiple washing steps. ICA shows outstanding advantages including userfriendly manipulation, short assay time, long-term stability, low cost, thus is fit for field assay and extensive screening. Unfortunately, up to now, most reported ICAs are designed to assay only one analyte with one single test strip, thus ICAs for multianalyte detection is urgently needed. Furthermore, the previous study of this method usually employed visible colorimetric readout strategy for qualitative or semi-quantitative detection of the analytes [23–27]. For example, Nara et al. [28] used colloidal gold nanoparticle as the signal probe and developed a rapid and semiquantitative immunochromatographic strip for cortisol analysis in serum. These reported ICAs using visual readout often provide only a yes/no response and suffer from insufficiently sensitivity. Therefore, to perform sensitive quantitative analysis, some ICAs coupled with fluorescent [29–31] and electrochemical detection

[32–34] have been developed, wherein organic dyes and quantum dots were adopted as the signal probes. CL detection is considered as a more powerful technology utilized for many bioassay applications because it only requires a simple detector without light source, and thus results in a lower background and an improved sensitivity. ICA strategy coupling with CL detection has already been reported for single-analyte immunoassay of cardiac troponin I [35]. However, no CL ICA method has been developed to detect multiple analytes up to now. Herein, we present a multianalyte immunochromatographic test strip with two test zones for CL detection of two b2-agonists. Horseradish peroxidase (HRP) was used as a highly sensitive CL probe to detect RAC and SAL. This proposed ICA should be one of the most rapid and sensitive methods which can quantify multiple b2-agonists at low concentrations for the purpose of food safety. 2. Experimental 2.1. Materials and equipments HRP-tagged mouse monoclonal antibody for RAC (anti-RAC mAb), HRP-tagged mouse monoclonal antibody for SAL (anti-SAL mAb), SAL-bovine serum albumin (BSA) and RAC-BSA conjugates were all provided by Guangzhou Ucando Biotechnology Co., Ltd. RAC and SAL standard samples were obtain from China Institute of Veterinary Drug Control. Luminol and p-iodophenol (PIP) used as HRP CL substrate were supplied by Sigma–Aldrich. SuperBlock1 T20 utilized as the blocking buffer was purchased from Thermo Fisher Scientific Inc. The dilution buffer for the antibodies and the antigens was phosphate buffer saline (PBS) at 0.10 M and pH 7.0. Ultrapure water (18.2 MV) produced by an ELGA PURELAB Classic system was employed to prepare all aqueous solutions. The swine urine samples were obtained from the local pig farms. They had been proved to be free of any b2-agonist by TSQ Quantum Ultra LC– MS/MS (Thermo Finnigin Co., Ltd.). All other reagents were of the best grade available and used as receiving. Nitrocellulose membrane (M180), glass fiber used as sample loading and conjugate pads, and cotton pulp used as absorbent paper

Fig. 1. Schematic illustration of enzyme-based CL ICA for multianalyte detection of RAC and SAL.




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Fig. 2. Effect of the concentrations of (A) HRP-tagged anti-RAC mAb and (B) HRP-tagged anti-SAL mAb on their corresponding signal-to-blank ratios, effect of the blocking time on the CL responses for (C) RAC and (D) SAL at 10 ng mL 1. All other conditions were the optimum conditions, n = 5.

were purchased from Millipore Corp. The CL responses were recorded by a MPI-A CL analyzer (Xi’an Remax Electronic Science & Technology Co., Ltd.) equipping a photomultiplier biased at 800 V. 2.2. Preparation of the ICA strip As seen in Fig. 1, the ICA strip with a width of 4 mm is composed of a sample loading pad (17 mm length), a conjugate pad (8 mm length), a nitrocellulose membrane (25 mm length), and an absorbent paper (17 mm length). The fabrication of the ICA strip was illustrated as follows. The sample loading pad was saturated with PBS (pH 8.0) containing BSA (2.0%), sucrose (2.0%), sodium azide (0.10%) and sodium borate (20 mM), and then dried and stored in ambient. HRP-tagged anti-RAC mAb (20 mg mL 1) and anti-SAL mAb (10 mg mL 1) were prepared in PBS containing 0.50% BSA, and mixed at the ratio of 1:1. Four microliters of the above mixed solution was deposited onto another glass fiber pad, followed by drying at 4 C to prepare the conjugate pad. Then 1.0 mL of RAC-BSA at 2.0 mg mL 1 and 1.0 mL of SAL-BSA at 2.0 mg mL 1 were loaded onto the nitrocellulose membrane to form two test zones (T1 and T2), respectively. T1 and T2 were 7 mm and 15 mm from the left edge of the nitrocellulose membrane, respectively. After drying overnight at 4 C, the nitrocellulose membrane was treated with 30 mL of blocking buffer for 90 min at 37 C to minimize the non-specific adsorption. Afterward, a sample loading pad, a conjugate pad, a nitrocellulose membrane, and an absorbent pad were sequentially assembled on a 60 mm long plastic adhesive backing plate as shown in Fig. 1. Each part overlapped 2 mm with the adjacent part to make sure that the solution could migrate through the whole test strip.

immuno-bindings on the two test zones were completed. Then T1 and T2 were cut down and placed into two reaction cells containing CL substrate consisted of PIP and luminol (100 mL for each cell). Finally, 15 mL of H2O2 solution was injected, and the CL responses were recorded at 60 s to accomplish immunoassay of RAC and SAL in turn. All the quantificational data for CL responses were calibrated by deducting the background. 3. Results and discussion 3.1. Principle of ICA The principle of ICA is illustrated in Fig. 1. The assay was based on a competitive immunoassay format. During the assay, a sample

2.3. Sample assay procedure Fifty microliters of sample solution composed of RAC and SAL was dropped onto the sample loading pad, and allowed to migrate along the whole test strip. After a reaction time of 15 min, the competitive

Fig. 3. CL responses from T1 and T2 when (A) RAC (10 ng mL 1) or (B) SAL (10 ng mL 1) were applied. All the conditions were the optimum conditions, n = 5.



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Fig. 4. The structures of five b2-agonists.

solution containing RAC and SAL was applied onto the sample loading pad and migrated towards another end of the strip owing to the capillary action driven by the absorbent paper. As the sample solution reached the conjugation pad, RAC and SAL reacted with their corresponding tracer antibodies (HRP-tagged anti-RAC and anti-SAL mAbs) that were pre-loaded on this pad. When the solution continued to migrate along the strip and reached T1, the excess unbinding HRP-tagged anti-RAC mAb was selectively captured by the RAC-BSA conjugate immobilized on this test zone. Then the solution continued to migrate to T2, thus the excess unbinding HRP-tagged anti-SAL mAb was selectively captured by the SAL-BSA conjugate immobilized on this zone. After the competitive immunoreactions, the two test zones were ready for CL detection. The concentrations of RAC and SAL were detected by the CL responses from T1 and T2, respectively. 3.2. Optimization of the experimental parameters The performance of ICA is always influenced by some parameters including the amount of the tracer antibody (HRPtagged antibody) and the blocking time. Fig. 2A and B showed the dependences of the CL responses on the concentrations of the tracer antibodies by using RAC at 10 ng mL 1, SAL at 10 ng mL 1 and PBS (as blank). It was found that the signal-to-blank ratios were minimum while the concentrations of the tracer antibodies for RAC and SAL were 20 and 10 mg mL 1, respectively, implying that the

competition capability of the analytes in the sample against the immobilized antigens was the strongest under this condition. The blocking buffer (SuperBlock1 T20) was applied to reduce the nonspecific adsorption. Its performance greatly influenced the background signals since ICA was conducted with a washing-free protocol. It was found that the CL intensities decreased with the blocking time and reached the steady values after 90 min (Fig. 2C and D), showing that the interference of background signals was minimum over this period. Therefore, the concentrations of 20 and 10 mg mL 1 for the tracer antibodies of RAC and SAL, respectively, and the blocking period of 90 min were finally chosen for the further investigation. Some conditions influencing the CL reaction were also studied, such as the CL substrate pH, the concentrations of luminol, PIP and H2O2. It was found that the optimum pH value for the CL reaction was 8.5 and the optimum concentrations for the three chemicals were 0.050, 0.050 and 10 mM, respectively (data not shown). 3.3. Evaluation of cross-reactivity Cross-reactivity was investigated to estimate the selectivity of this immunochromatographic test strip for multianalyte assay. In this protocol, the cross-reactivity between the two analytes and their noncognate antibodies was examined by the standard samples which contained only one analyte (RAC or SAL) at a fixed concentration (10 ng mL 1) for each time. The blank signals were

Fig. 5. (A) CL responses of RAC at the concentrations of (a) 0, (b) 0.50, (c) 1.0, (d) 10, (e) 20, and (f) 40 ng mL 1; (B) CL responses of SAL at the concentrations of (a) 0, (b) 0.10, (c) 1.0, (d) 10, (e) 30, and (f) 50 ng mL 1. Inset: calibration curves, where n = 5 for each point. The CL signals in the curves were calibrated by deducting the background response. All other conditions were the optimum conditions.




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Table 1 Comparison of analytical parameters resulting from different methods for determination of RAC or SAL. Method

Analyte

DL (ng mL

HPLC GC–MS LC–MS

RAC SAL RAC SAL RAC SAL SAL RAC SAL RAC RAC SAL SAL RAC SAL

3.0 0.50 5.0 2.0 0.60 0.30 0.060 0.12 3.5 0.50 0.39 0.43 0.14 0.20 0.040

CE CE–MS Electrochemical immunosensor Surface plasmon resonance immunoassay Flurescence sensor Time-resolved chemiluminescence strategy Colloidal gold-based immunochromatographic strip Silica nanoparticle-based immunochromatographic strip Molecularly imprinted polymers CL-based immunochromatographic strip

1

)

Reference [4] [6] [8] [10] [12] [16] [18] [20] [21] [27] [29] [37] The proposed method

Table 2 Recovery tests of RAC and SAL spiked in blank swine urine samples obtained by the proposed method and the LC–MS/MS method (n = 5). The proposed method (n = 5) Sample number

Added (ng mL 1) Found (ng mL RSD (%) Recovery (%)

1

)

1

LC–MS/MS (n = 5)

2

3

RAC

SAL

20.0

20.0

5.00

5.00

19.6 4.9 98.0

18.0 4.0 90.0

5.43 5.9 108.6

5.76 7.8 115.2

RAC

SAL

1

RAC 1.00 0.887 4.8 88.7

detected from a dilution buffer without RAC and SAL. As presented in Fig. 3, upon the addition of RAC the CL response from T1 showed a significant decrease of 34% due to the competitive binding, while the CL response from T2 only showed slight change of 0.49% compared to the blank signal. The two values were 0.52% and 32%, respectively, when SAL was applied in the further investigation. These results indicated that the cross-reactivity was negligible and the two analytes could be detected in a single run using this ICA strategy without noticeable interference to each other. The potential interference from several other analogues utilizing as “lean meat agents”, such as clenbuterol, terbutaline and phenylethanolamine A, was also investigated in detail. It was found that the cross-reactivity of anti-RAC mAb with terbutaline and clenbuterol was negligible, while that with phenylethanolamine A was 9.1%. Similarly, anti-RAC mAb was found to only crossreact with terbutaline, clenbuterol for 42% and 65%, respectively. These cross-reactivities should be resulted from the very similar molecule structures between these agents (Fig. 4). Nevertheless, this CL ICA is still of value in “lean meat agents” screening because all these b2-agonists have been rigorously forbidden to be abused illegally in animal feeding in most countries. 3.4. Performance of CL ICA Under the selected optimum conditions, the CL signals linearly decreased with the increase of the concentrations of RAC and SAL (Fig. 5) because a competitive format was employed. The calibration curves for RAC and SAL showed the linear ranges of 0.50–40 and 0.10–50 ng mL 1, with the correlation coefficients (R2) of 0.995 and 0.992, respectively. The detection limits (DLs) defined as the sample concentrations producing signal to noise ratios of 3 were 0.20 and 0.040 ng mL 1 for RAC and SAL, respectively. As seen in Table 1, the DLs of this method are lower than those of the most reported approaches. The relative standard deviations (RSDs) for 1.0 and 10 ng mL 1 RAC were 6.0% and 6.2%, respectively, while the RSDs for 1.0 and 10 ng mL 1 SAL were 5.9% and 5.6%, respectively,

SAL

2

3

RAC

SAL

RAC

SAL

0.500

20.0

20.0

5.00

5.00

0.464 6.7 92.8

18.6 4.2 93.0

17.1 3.9 85.5

4.38 5.1 87.6

4.22 3.6 84.4

RAC 1.00 0.926 4.6 92.6

SAL 0.500 0.431 5.8 86.2

showing acceptable reproducibility. Furthermore, the storage durability of the immunochromatographic test strip was also studied by periodical testing. Only less than 10% of the initial response was lost after storage at 4 C (sealed) for 2 months. Therefore, the stability of the test strip was acceptable. 3.5. Application in spiked swine urine samples Three blank swine urines were spiked with standard RAC and SAL at different known amounts, and detected using the proposed method to assess its application potential. As seen in Table 2, the recoveries for the spiked RAC and SAL were 89–109% and 90–115%, respectively. All RSDs were less than 7.8%, suggesting acceptable accuracy of this proposed approach. Also, the recovery tests were carried out using LC–MS/MS with the standard reference method NYG 1063.3-2008 (Ministry of Agriculture of China) [36], to verify the reliability of the proposed method. The results presented in Table 2 showed that the agreement of the two methods was acceptable. 4. Conclusions In summary, an immunochromatographic test strip was developed for the rapid detection of RAC and SAL within 20 min. Compared with the conventional visual colloidal gold-based test strip for qualitative or semi-quantitative assay, this quantitative method shows much higher sensitivity due to the application of CL detection. Furthermore, fabrication of two parallel test zones on it allows multianalyte immunoassay of two b2-agonists using a single strip. This method also possesses attractive characteristics such as simple manipulation, long-term stability, low cost, and high assay efficiency. Therefore this test strip is a portable device suitable for rapid screening and field analysis, and shows a promising prospect in food safety. In the future this proof-ofprinciple device can be also developed for point-of-care test of biomarkers panel for clinical diagnostics purpose.



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Acknowledgements This work was financially supported by the Natural Science Foundation of China (21175111), Program for Innovative Research Team in University of Chongqing (2013), Natural Science Foundation of Chongqing (CSTC2013jjB0096), and the Fundamental Research Funds for the Central Universities (XDJK2013A025 and XDJK2012A002). References [1] E. Shishani, S.C. Chai, S. Jamokha, G. Aznar, M.K. Hoffman, Determination of ractopamine in animal tissues by liquid chromatography-fluorescence and liquid chromatography/tandem mass spectrometry, Anal. Chim. Acta 483 (2003) 137–145. [2] J.F. Martinez-Navarro, Food poisoning related to consumption of illicit b-agonist in liver, Lancet 336 (1990) 1311–1312. [3] G. Brambilla, T. Cenci, F. Franconi, R. Galarini, A. Macri, F. Rodoni, M. Strozzi, A. Loizzo, Clinical and pharmacological profile in a clenbuterol epidemic poisoning of contaminated beef meat in Italy, Toxicol. Lett. 114 (2000) 47–53. [4] W. Du, G. Zhao, Q. Fu, M. Sun, H.Y. Zhou, C. Chang, Combined microextraction by packed sorbent and high-performance liquid chromatography-ultraviolet detection for rapid analysis of ractopamine in porcine muscle and urine samples, Food Chem. 145 (2014) 789–795. [5] Y.T. Zhang, Z.J. Zhang, Y.H. Sun, Y. Wei, Development of an analytical method for the determination of b2-agonist residues in animal tissues by highperformance liquid chromatography with on-line electrogenerated [Cu (HIO6)2]5 -luminol chemiluminescence detection, J. Agric. Food Chem. 55 (2007) 4949–4956. [6] M. Caban, P. Stepnowski, M. Kwiatkowski, N. Migowska, J. Kumirska, Determination of b-blockers and b-agonists using gas chromatography and gas chromatography–mass spectrometry – a comparative study of the derivatization step, J. Chromatogr. A 1218 (2011) 8110–8122. [7] P. Gallo, G. Brambilla, B. Neri, M. Fiori, C. Testa, L. Serpe, Purification of clenbuterol-like b2-agonist drugs of new generation from bovine urine and hair by a1-acid glycoprotein affinity chromatography and determination by gas chromatography–mass spectrometry, Anal. Chim. Acta 587 (2007) 67–74. [8] X.J. Wang, F. Zhang, F. Ding, W.Q. Li, Q.Y. Chen, X.G. Chu, C.B. Cheng, Simultaneous determination of 12 b-agonists in feeds by ultra-highperformance liquid chromatography–quadrupole-time-of-flight mass spectrometry, J. Chromatogr. A 1278 (2013) 82–88. [9] C. Li, Y.L. Wu, T. Yang, Y. Zhang, W.G. Huang-Fu, Simultaneous determination of clenbuterol, salbutamol and ractopamine in milk by reversed-phase liquid chromatography tandem mass spectrometry with isotope dilution, J. Chromatogr. A 1217 (2010) 7873–7877. [10] L.B. Li, H.W. Du, H. Yu, L. Xu, T.Y. You, Application of ionic liquid as additive in determination of three b-agonists by capillary electrophoresis with amperometric detection, Electrophoresis 34 (2013) 277–283. [11] H. Lodén, C. Pettersson, T. Arvidsson, A. Amini, Quantitative determination of salbutamol in tablets by multiple-injection capillary zone electrophoresis, J. Chromatogr. A 1207 (2008) 181–185. [12] O. Anurukvorakun, W. Buchberger, M. Himmelsbach, C.W. Klampel, L. Suntornsuk, A sensitive non-aqueous capillary electrophoresis–mass spectrometric method for multiresidue analyses of b-agonists in pork, Biomed. Chromatogr. 24 (2010) 588–599. [13] S.Y. Sheu, Y.C. Lei, Y.T. Tai, T.H. Chang, T.F. Kuo, Screening of salbutamol residues in swine meat and animal feed by an enzyme immunoassay in Taiwan, Anal. Chim. Acta 654 (2009) 148–153. [14] B.Y. Cao, G.Z. He, H. Yang, H.F. Chang, S.Q. Li, A.P. Deng, Development of a highly sensitive and specific enzyme-linked immunosorbent assay (ELISA) for the detection of phenylethanolamine A in tissue and feed samples and confirmed by liquid chromatography tandem mass spectrometry (LC–MS/MS), Talanta 115 (2013) 624–630. [15] G. Liu, H.D. Chen, H.Z. Peng, S.P. Song, J.M. Gao, J.X. Lu, M. Ding, L.Y. Li, S.Z. Ren, Z.Y. Zou, C.H. Fan, A carbon nanotube-based high-sensitivity electrochemical immunosensor for rapid and portable detection of clenbuterol, Biosens. Bioelectron. 28 (2011) 308–313. [16] S. Liu, Q. Lin, X.M. Zhang, X.R. He, X.R. Xing, W.J. Lian, J.D. Huang, Electrochemical immunosensor for salbutamol detection based on CSFe3O4-PAMAM-GNPs nanocomposites and HRP-MWCNTs-Ab bioconjugates for signal amplification, Sens. Actuators B 156 (2011) 71–78.

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Development of an ELISA and immunochromatographic strip for highly sensitive detection of microcystin-LR.

A highly sensitive ELISA and immunochromatographic strip for the detection of Salmonella typhimurium in milk samples.

Development of an Immunochromatographic Strip Test for the Rapid Detection of Alternariol Monomethyl Ether in Fruit.

Development of an immunochromatographic strip test for the rapid detection of zearalenone in corn.

Development of an immunochromatographic strip test for rapid detection of ciprofloxacin in milk samples.

Development of an immunochromatographic strip test for rapid detection of lily symptomless virus.

A rapid one-step immunochromatographic test strip for rabies detection using canine serum samples.

Development of immunochromatographic strip test using fluorescent, micellar silica nanosensors for rapid detection of B. abortus antibodies in milk samples.

Development of an immunochromatographic strip for rapid detection of Pantoea stewartii subsp. stewartii.

Development of an immunochromatographic test strip for detection of cholera toxin.

Rapid and quantitative detection of zoonotic influenza A virus infection utilizing coumarin-derived dendrimer-based fluorescent immunochromatographic strip test (FICT).

Development of a Calibration Strip for Immunochromatographic Assay Detection Systems.

Rapid diagnosis of cryptococcosis using an antigen detection immunochromatographic test.

Detection of Staphylococcus Enterotoxin B (SEB) Using an Immunochromatographic Test Strip.

Development of a Colloidal Gold-Based Immunochromatographic Strip for Rapid Detection of Klebsiella pneumoniae Serotypes K1 and K2.

Development of an ultrasensitive immunochromatographic assay (ICA) strip for the rapid detection of phenylethanolamine A in urine and pork samples.

Rapid penicillinase paper strip test for detection of beta-lactamase-producing Haemophilus influenzae and Neisseria gonorrhoeae.

Development of a colloidal gold immunochromatographic strip for the rapid detection of soft-shelled turtle systemic septicemia spherical virus.

Correction for Okabayashi et al., Detection of Chikungunya Virus Antigen by a Novel Rapid Immunochromatographic Test.

Development of a test strip for rapid detection of lactoperoxidase in raw milk.

Rapid and quantitative detection of C-reactive protein using quantum dots and immunochromatographic test strips.

Efficiency assessment of immunochromatographic strip test for the diagnosis of alpha-thalassemia-1 carriers.

Utility of a rapid immunochromatographic strip test in detecting canine parvovirus infection compared with polymerase chain reaction.

Preparation of colloidal gold immunochromatographic strip for detection of Paragonimiasis skrjabini.