Skip to main content
SpringerLink
Log in

Enhancing the immunofluorescent sensitivity for detection of Acidovorax citrulli using fluorescein isothiocyanate labeled antigen and antibody

  • Research Paper
  • Published:
Analytical and Bioanalytical Chemistry Aims and scope Submit manuscript

Abstract

A rapid lateral flow immunochromatographic strip (ICS) using fluorescein isothiocyanate (FITC) labeled antigen and antibody was developed for the detection of Acidovorax citrulli (Ac) in melons and vegetable samples. In the ICS, signal amplification was realized based on antigen Ac and anti-Ac monoclonal antibody (McAb) 4F conjugated with FITC, respectively, which were forming two probes. The control line and the test line were obtained by immobilizing the goat anti-mouse IgG antibody and anti-Ac McAb 6D on both sides of the nitrocellulose membrane. The visual detection limit of the strip was 105 CFU/mL, which was 10-fold sensitive compared to the strip of FITC only labeling antigen or antibody. Signal amplification ICS was successfully applied to the detection of Ac in melon and vegetable samples with less detection time and operation procedures compared to the traditional enzyme-linked immunosorbent assay (ELISA) and PCR methods. This is the first report of using FITC labeled antigen and McAb as dual fluorescent probes to develop a direct-type immunofluorescence strip for the rapid and sensitive detection of Ac, which demonstrates a powerful tool for rapidly screening Ac in plant materials and other samples.

The schematic presentation of the test strip (a) and the positive result (b) or negative result (c) of the test strip

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  1. Feng JJ, Li JQ, Walcott RR, Zhang GM, Luo LX, Kang L, et al. Advances in detection of Acidovorax citrulli, the causal agent of bacterial fruit blotch of cucurbits. Seed Sci Technol. 2013;41(1):1–15.

    Article  CAS  Google Scholar 

  2. Rane KK, Latin RX. Bacterial fruit blotch of watermelon: association of the pathogen with seed. Plant Dis. 1992;76(5):761–5.

    Article  Google Scholar 

  3. Webb RE, Goth RW. A seedborne bacterium isolated from watermelon. Plant Dis Rep. 1965;49:818–21.

    Google Scholar 

  4. Bahar O, Kritzman G, Burdman S. Bacterial fruit blotch of melon: screens for disease tolerance and role of seed transmission in pathogenicity. Eur J Plant Pathol. 2009;123(1):71–83.

    Article  Google Scholar 

  5. Singh J, Sharma S, Nara S. Evaluation of gold nanoparticle based lateral flow assays for diagnosis of enterobacteriaceae members in food and water. Food Chem. 2015;170:470–83.

    Article  CAS  Google Scholar 

  6. Ren W, Huang Z, Xu Y, Li Y, Ji Y, Su B. Urchin-like gold nanoparticle-based immunochromatographic strip test for rapid detection of fumonisin B1 in grains. Anal Bioanal Chem. 2015;407(24):7341–8.

    Article  CAS  Google Scholar 

  7. Liu Y, Wu A, Hu J, Lin M, Wen M, Zhang X, et al. Detection of 3-phenoxybenzoic acid in river water with a colloidal gold based lateral flow immunoassay. Anal Biochem. 2015;483(1):7–11.

    Article  CAS  Google Scholar 

  8. Zhou Y, Li YS, Meng XY, Zhang YY, Yang L, Li ZH, et al. Production of a monoclonal antibody and development of an immunoassay for detection of Cr(III) in water samples. Chemosphere. 2013;93(10):2467–72.

    Article  CAS  Google Scholar 

  9. Zhang MZ, Wang MZ, Chen ZL, Fang JH, Fang MM, Liu J, et al. Development of a colloidal gold-based lateral-flow immunoassay for the rapid simultaneous detection of clenbuterol and ractopamine in swine urine. Anal Bioanal Chem. 2009;395(8):2591–9.

    Article  CAS  Google Scholar 

  10. Byzova NA, Smirnova NI, Zherdev AV, Eremin SA, Shanin IA, Lei HT, et al. Rapid immunochromatographic assay for ofloxacin in animal original foodstuffs using native antisera labeled by colloidal gold. Talanta. 2014;119:125–1232.

    Article  CAS  Google Scholar 

  11. Di Nardo F, Anfossi L, Giovannoli C, Passini C, Goftman VV, Goryacheva IY, et al. A fluorescent immunochromatographic strip test using quantum dots for fumonisins detection. Talanta. 2016;150:463–8.

    Article  Google Scholar 

  12. Sirkka N, Lyytikäinen A, Savukoski T, Soukka T. Upconverting nanophosphors as reporters in a highly sensitive heterogeneous immunoassay for cardiac troponin I. Anal Chim Acta. 2016;925:82–7.

    Article  CAS  Google Scholar 

  13. Anfossi L, Di Nardo F, Giovannoli C, Passini C, Baggiani C. Increased sensitivity of lateral flow immunoassay for ochratoxin A through silver enhancement. Anal Bioanal Chem. 2013;405(30):9859–67.

    Article  CAS  Google Scholar 

  14. Liu D, Huang Y, Wang S, Liu K, Chen M, Xiong Y, et al. A modified lateral flow immunoassay for the detection of trace aflatoxin M1 based on immunomagnetic nanobeads with different antibody concentrations. Food Control. 2015;51:218–24.

    Article  Google Scholar 

  15. Nagatani N, Yamanaka K, Ushijima H, Koketsu R, Sasaki T, Ikuta K, et al. Detection of influenza virus using a lateral flow immunoassay for amplified DNA by a microfluidic RT-PCR chip. Analyst. 2012;137(137):3422–6.

    Article  CAS  Google Scholar 

  16. Liu Q, Liu XP, Wei YP, Mao CJ, Niu HL, Song JM, et al. Electrochemiluminescence immunoassay for the carcinoembryonic antigen using CdSe:Eu nanocrystals. Microchim Acta. 2017;184(5):1353–60.

    Article  CAS  Google Scholar 

  17. Wang C, Wu J, Zong C, Ju H, Yan F. Highly sensitive rapid chemiluminescent immunoassay using the DNAzyme label for signal amplification. Analyst. 2011;136(20):4295–300.

    Article  CAS  Google Scholar 

  18. Saniee P, Siavoshi F, Nikbakht BG, Khormali M, Sarrafnejad A, Malekzadeh R. Localization of H.pylori within the vacuole of Candida yeast by direct immunofluorescence technique. Arch Iran Med. 2013;16(12):705–10.

    Google Scholar 

  19. Yeo KB, Kim HB, Choi YS, Pack SP. Highly effective detection of inflamed cells using a modified bradykinin ligand labeled with FITC fluorescence. Enzyme Microb Technol. 2016;82:191–6.

    Article  CAS  Google Scholar 

  20. Kim BS, Oh JM, Kim KS, Seo KS, Cho JS, Khang G, et al. BSA-FITC-loaded microcapsules for in vivo delivery. Biomaterials. 2009;30(5):902–9.

    Article  CAS  Google Scholar 

  21. Murányi J, Gyulavári P, Varga A, Bökönyi G, Tanai H, Vántus T, et al. Synthesis, characterization, and systematic comparison of FITC-labelled GnRH-I, -II and -III analogues on various tumour cells. J Peptide Sci. 2016;22(8):552–60.

    Article  Google Scholar 

  22. Blajman JE, Astesana DM, Zimmermann JA, Rossler E, Scharpen AR, Berisvil AP, et al. Quantification of FITC-labelled probiotic Lactobacillus salivarius DSPV 001P during gastrointestinal transit in broilers. Beneficial Microbes. 2017;8(1):55–64.

    Article  CAS  Google Scholar 

  23. Zeng H, Guo W, Liang B, Li J, Zhai X, Song C, et al. Self-paired monoclonal antibody lateral flow immunoassay strip for rapid detection of Acidovorax avenae subsp. citrulli. Anal Bioanal Chem. 2016;408(22):6071–8.

    Article  CAS  Google Scholar 

  24. Hu C, Chiang C, Hong P, Yeh M. Influence of charge on FITC-BSA-loaded chondroitin sulfate-chitosan nanoparticles upon cell uptake in human Caco-2 cell monolayers. Int J Nanomed. 2012;7(8):4861–72.

    CAS  Google Scholar 

  25. Himananto O, Luxananil P, Kumpoosiri P. Novel and highly specific monoclonal antibody to Acidovorax citrulli and development of ELISA-based detection in cucurbit leaves and seed. Plant Dis. 2011;95(95):1172–8.

    Article  CAS  Google Scholar 

  26. Charlermroj R, Himananto O, Seepiban C, Kumpoosiri M, Warin N, Oplatowska M, et al. Multiplex detection of plant pathogens using a microsphere immunoassay technology. Plos One. 2013;8(4):e62344.

    Article  CAS  Google Scholar 

  27. Bahar O, Efrat M, Hadar E, Dutta B, Walcott RR, Burdman S. New subspecies-specific polymerase chain reaction-based assay for the detection of Acidovorax avenae subsp. citrulli. Plant Pathol. 2008;57(4):754–63.

    Article  CAS  Google Scholar 

  28. Ha Y, Fessehaie A, Ling KS, Wechter WP, Keinath AP, Walcott RR. Simultaneous detection of Acidovorax avenae subsp. citrulli and Didymella bryoniae in cucurbit seedlots using magnetic capture hybridization and real-time polymerase chain reaction. Phytopathology. 2009;99(6):666–78.

    Article  CAS  Google Scholar 

  29. Zhao W, Lu J, Ma W, Xu C, Kuang H, Zhu S. Rapid on-site detection of Acidovorax avenae subsp. citrulli by gold-labeled DNA strip sensor. Biosens Bioelectron. 2011;26(10):4241–4.

    Article  CAS  Google Scholar 

  30. Qu H, Zhang Y, Qu B, Kong H, Qin G, Liu S, et al. Rapid lateral-flow immunoassay for the quantum dot-based detection of puerarin. Biosens Bioelectron. 2016;81:358–62.

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This research is supported by the Natural Science Foundation of China (No. 31371776), Science and Technology Innovation Plan of Shanghai: Yangtze River Delta joint research (No. 15395810900), Minimally Motivated Innovation Fund (YS30809102). and Graduate Education Innovation Program of Shanghai.

Author information

Authors and Affiliations

  1. School of Medical Instrument and Food Engineering, University of Shanghai for Science and Technology, Shanghai, 200093, China

    Haijuan Zeng, Xuzhao Zhai, Shujuan Wang & Qing Liu

  2. Hepatobiliary Surgery Department, Ningxia People’s Hospital, Yinchuan, 750002, China

    Duoqiang Zhang

Authors
  1. Haijuan Zeng
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Duoqiang Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Xuzhao Zhai
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Shujuan Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Qing Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Qing Liu.

Ethics declarations

Conflict of Interest

The authors confirm that no conflict of interest exists for this manuscript and all of the authors agree to the submission of this paper.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zeng, H., Zhang, D., Zhai, X. et al. Enhancing the immunofluorescent sensitivity for detection of Acidovorax citrulli using fluorescein isothiocyanate labeled antigen and antibody. Anal Bioanal Chem 410, 71–77 (2018). https://doi.org/10.1007/s00216-017-0690-2

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00216-017-0690-2

Keywords

Search

Navigation