Abstract
Microcystis species identification is essential for ecological studies and water bloom control. Immunoassays are more specific and convenient and several approaches have been used to develop for diagnosing harmful red tide algae. However, investigations on Microcystis identification using immunological approaches are still in the initial stage. In this study, Microcystis aeruginosa PCC7806 lysates were utilized as coated antigens to enrich and screen specific Microcystis nanobodies from a human domain antibody display library. After three rounds of enrichment, 10 positive monoclonal particles were isolated from the library and the most two positive nanobodies (DAb2 and Dab3) were effectively produced in Escherichia coli BL21. Finally, the DAb2 showed specific immune binding to different Microcystis by the immuno-dot blot assay. This antibody could be used to establish an immunological method to identify Microcystis.
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Data Availability Statement
The data generated in this study are available from the corresponding author or the first author upon reasonable request.
References
Adkins J A, Boehle K, Friend C et al. 2017. Colorimetric and electrochemical bacteria detection using printed paperand transparency-based analytic devices. Analytical Chemistry, 89(6): 3613–3621, https://doi.org/10.1021/acs.analchem.6b05009.
Bhardwaj J, Kim M W, Jang J. 2020. Rapid airborne influenza virus quantification using an antibody-based electrochemical paper sensor and electrostatic particle concentrator. Environmental Science & Technology, 54(17): 10700–10712, https://doi.org/10.1021/acs.est.0c00441.
Blanco Y, Moreno-Paz M, Parro V. 2017. Experimental protocol for detecting Cyanobacteria in liquid and solid samples with an antibody microarray chip. Journal of Visualized Experiments, 120: e54994, https://doi.org/10.3791/54994.
Blanco Y, Quesada A, Gallardo-Carreño I et al. 2015. CYANOCHIP: an antibody microarray for high-taxonomical-resolution cyanobacterial monitoring. Environmental Science & Technology, 49(3): 1611–1620, https://doi.org/10.1021/es5051106.
Bradford M M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72(1–2): 248–254, https://doi.org/10.1016/0003-2697(76)90527-3.
Carrera M, Garet E, Barreiro A et al. 2010. Generation of monoclonal antibodies for the specific immunodetection of the toxic dinoflagellate Alexandrium minutum Halim from Spanish waters. Harmful Algae, 9(3): 272–280, https://doi.org/10.1016/j.hal.2009.11.004.
Folorunsho O G, Oloketuyi S F, Mazzega E et al. 2021. Nanobody-dependent detection of Microcystis aeruginosa by ELISA and thermal lens spectrometry. Applied Biochemistry and Biotechnology, 193(9): 2729–2741, https://doi.org/10.1007/s12010-021-03552-6.
García-García A, Madrid R, González I et al. 2020. A novel approach to produce phage single domain antibody fragments for the detection of gluten in foods. Food Chemistry, 321: 126685, https://doi.org/10.1016/j.foodchem.2020.126685.
Gas F, Pinto L, Baus B et al. 2009. Monoclonal antibody against the surface of Alexandrium minutum used in a whole-cell ELISA. Harmful Algae, 8(3): 538–545, https://doi.org/10.1016/j.hal.2008.08.027.
Guiry M D, Guiry G M. 2019. AlgaeBase. World-wide electronic publication, National University of Ireland, Galway. http://www.algaebase.org. Accessed on 2019-12-1
Jiang W Z, Cossey S, Rosenberg J N et al. 2014. A rapid live-cell ELISA for characterizing antibodies against cell surface antigens of Chlamydomonas reinhardtii and its use in isolating algae from natural environments with related cell wall components. BMC Plant Biology, 14(1): 244, https://doi.org/10.1186/s12870-014-0244-0.
Jiang W Z, Rosenberg J N, Wauchope A D et al. 2013. Generation of a phage-display library of single-domain camelid VHH antibodies directed against Chlamydomonas reinhardtii antigens, and characterization of VHHs binding cell-surface antigens. The Plant Journal, 76(4): 709–717, https://doi.org/10.1111/tpj.12316.
Komárek J, Komárková J. 2002. Review of the European Microcystis-morphospecies (Cyanoprokaryotes) from nature. Czech Phycology, 2: 1–24.
Lee J, Kim J H, Kim B N et al. 2020. Identification of novel paraben-binding peptides using phage display. Environmental Pollution, 267: 115479, https://doi.org/10.1016/j.envpol.2020.115479.
Li M, Zhu W, Gao L et al. 2013. Changes in extracellular polysaccharide content and morphology of Microcystis aeruginosa at different specific growth rates. Journal of Applied Phycology, 25(4): 1023–1030, https://doi.org/10.1007/s10811-012-9937-7.
Mazzega E, Beran A, Cabrini M et al. 2019. In vitro isolation of nanobodies for selective Alexandrium minutum recognition: a model for convenient development of dedicated immuno-reagents to study and diagnostic toxic unicellular algae. Harmful Algae, 82: 44–51, https://doi.org/10.1016/j.hal.2019.01.002.
Mendoza H, López-Rodas V, González-Gil S et al. 1995. The use of polyclonal antisera and blocking of antibodies in the identification of marine dinoflagellates: species-specific and clone-specific antisera against Gymnodinium and Alexandrium. Journal of Experimental Marine Biology and Ecology, 186(1): 103–115, https://doi.org/10.1016/0022-0981(94)00160-F.
Mitra M, Dewez D, García-Cerdán J G et al. 2012. Polyclonal antibodies against the TLA1 protein also recognize with high specificity the D2 reaction center protein of PSII in the green alga Chlamydomonas reinhardtii. Photosynthesis Research, 112(1): 39–47, https://doi.org/10.1007/s11120-012-9733-x.
Niu W M, He E Q, Wu Q G et al. 2012. Use of fluorescent europium chelates as labels for detection of microcystin-LR in Taihu Lake, China. Journal of Rare Earths, 30(9): 941–946, https://doi.org/10.1016/S1002-0721(12)60158-6.
Otsuka S, Suda S, Li R H et al. 2000. Morphological variability of colonies of Microcystis morphospecies in culture. The Journal of General and Applied Microbiology, 46(1): 39–50, https://doi.org/10.2323/jgam.46.39.
Qiu Y L, Li P, Dong S et al. 2018. Phage-mediated competitive chemiluminescent immunoassay for detecting Cry1Ab toxin by using an anti-idiotypic camel nanobody. Journal of Agricultural and Food Chemistry, 66(4): 950–956, https://doi.org/10.1021/acs.jafc.7b04923.
Sun Q Q, Zhu W, Li M et al. 2016. Morphological changes of Microcystis aeruginosa colonies in culture. Journal of Limnology, 75(1): 14–23, https://doi.org/10.4081/jlimnol.2015.1225.
Szkola A, Linares E M, Worbs S et al. 2014. Rapid and simultaneous detection of ricin, staphylococcal enterotoxin B and saxitoxin by chemiluminescence-based microarray immunoassay. Analyst, 139(22): 5885–5892, https://doi.org/10.1039/C4AN00345D.
Tan W H, Liu Y, Wu Z X et al. 2010. CpcBA-IGS as an effective marker to characterize Microcystis wesenbergii (Komárek) Komárek in Kondrateva (cyanobacteria). Harmful Algae, 9(6): 607–612, https://doi.org/10.1016/j.hal.2010.04.011.
Thiruppathiraja C, Kamatchiammal S, Adaikkappan P et al. 2011. An advanced dual labeled gold nanoparticles probe to detect Cryptosporidium parvum using rapid immuno-dot blot assay. Biosensors and Bioelectronics, 26(11): 4624–4627, https://doi.org/10.1016/j.bios.2011.05.006.
Wang Y, Zhang X, Zhang C Z et al. 2012. Isolation of single chain variable fragment (scFv) specific for Cry1C toxin from human single fold scFv libraries. Toxicon, 60(7): 1290–1297, https://doi.org/10.1016/j.toxicon.2012.08.014.
Waterbury J B. 2006. The cyanobacteria-isolation, purification and identification. In: Dworkin M, Falkow S, Rosenberg E et al eds. The Prokaryotes. Springer, New York. p.1053–1073, https://doi.org/10.1007/0-387-30744-3_38.
West N J, Bacchieri R, Hansen G et al. 2006. Rapid quantification of the toxic alga Prymnesium parvum in natural samples by use of a specific monoclonal antibody and solid-phase cytometry. Applied and Environmental Microbiology, 72(1): 860–868, https://doi.org/10.1128/AEM.72.1.860-868.2006.
Xiao M, Li M, Reynolds C S. 2018. Colony formation in the cyanobacterium Microcystis. Biological Reviews, 93(3): 1399–1420, https://doi.org/10.1111/brv.12401.
Xu C X, He D, Zu Y et al. 2021. Microcystin-LR heterologous genetically engineered antibody recombinant and its binding activity improvement and application in immunoassay. Journal of Hazardous Materials, 406: 124596, https://doi.org/10.1016/j.jhazmat.2020.124596.
Xu C X, Liu X Q, Liu Y et al. 2019a. High sensitive single chain variable fragment screening from a microcystin-LR immunized mouse phage antibody library and its application in immunoassay. Talanta, 197: 397–405, https://doi.org/10.1016/j.talanta.2019.01.064.
Xu C X, Miao W J, He Y et al. 2019b. Construction of an immunized rabbit phage display antibody library for screening microcystin-LR high sensitive single-chain antibody. International Journal of Biological Macromolecules, 123: 369–378, https://doi.org/10.1016/j.ijbiomac.2018.11.122.
Xu C X, Yang Y, Liu L W et al. 2018. Microcystin-LR nanobody screening from an alpaca phage display nanobody library and its expression and application. Ecotoxicology and Environmental Safety, 151: 220–227, https://doi.org/10.1016/j.ecoenv.2018.01.003.
Yu H W, Jang A, Kim L H et al. 2011. Bead-based competitive fluorescence immunoassay for sensitive and rapid diagnosis of cyanotoxin risk in drinking water. Environmental Science & Technology, 45(18): 7804–7811, https://doi.org/10.1021/es201333f.
Zhang W Q, Lin M Q, Yan Q et al. 2021. An intracellular nanobody targeting T4SS effector inhibits Ehrlichia infection. Proceedings of the National Academy of Sciences, 118(18): e2024102118, https://doi.org/10.1073/pnas.2024102118.
Zhang X, Liu Y, Zhang C Z et al. 2012. Rapid isolation of single-chain antibodies from a human synthetic phage display library for detection of Bacillus thuringiensis (Bt) Cry1B toxin. Ecotoxicology and Environmental Safety, 81: 84–90, https://doi.org/10.1016/j.ecoenv.2012.04.021.
Zhao A Z, Tohidkia M R, Siegel D L et al. 2016. Phage antibody display libraries: a powerful antibody discovery platform for immunotherapy. Critical Reviews in Biotechnology, 36(2): 276–289, https://doi.org/10.3109/07388551.2014.958978.
Zhu Y Y, Wu J J X, Han L J et al. 2020. Nanozyme sensor arrays based on heteroatom-doped graphene for detecting pesticides. Analytical Chemistry, 92(11): 7444–7452, https://doi.org/10.1021/acs.analchem.9b05110.
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Supported by the National Natural Science Foundation of China (Nos. 31971561, 31370217, 31701724) and the Research Program of Jiangsu Province of China
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Zu, Y., Miao, W., Luo, Y. et al. Screening of nanobody against Microcystis from a human phage display nanobody library. J. Ocean. Limnol. 40, 1696–1705 (2022). https://doi.org/10.1007/s00343-022-1361-5
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DOI: https://doi.org/10.1007/s00343-022-1361-5