POLÁCHOVÁ, Veronika, Matěj PASTUCHA, Zuzana MIKUŠOVÁ, Matthias Jürgen MICKERT, Antonín HLAVÁČEK, Hans-Heiner GORRIS, Petr SKLÁDAL and Zdeněk FARKA. Click-conjugated photon-upconversion nanoparticles in an immunoassay for honeybee pathogen Melissococcus plutonius. Nanoscale. Cambridge: Royal Society of Chemistry, 2019, vol. 11, No 17, p. 8343-8351. ISSN 2040-3364. Available from: https://dx.doi.org/10.1039/C9NR01246J.
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Basic information
Original name Click-conjugated photon-upconversion nanoparticles in an immunoassay for honeybee pathogen Melissococcus plutonius
Authors POLÁCHOVÁ, Veronika (203 Czech Republic, belonging to the institution), Matěj PASTUCHA (203 Czech Republic, belonging to the institution), Zuzana MIKUŠOVÁ (703 Slovakia, belonging to the institution), Matthias Jürgen MICKERT (276 Germany), Antonín HLAVÁČEK (203 Czech Republic), Hans-Heiner GORRIS (276 Germany), Petr SKLÁDAL (203 Czech Republic, belonging to the institution) and Zdeněk FARKA (203 Czech Republic, guarantor, belonging to the institution).
Edition Nanoscale, Cambridge, Royal Society of Chemistry, 2019, 2040-3364.
Other information
Original language English
Type of outcome Article in a journal
Field of Study 10406 Analytical chemistry
Country of publisher Czech Republic
Confidentiality degree is not subject to a state or trade secret
WWW URL
Impact factor Impact factor: 6.895
RIV identification code RIV/00216224:14740/19:00108343
Organization unit Central European Institute of Technology
Doi http://dx.doi.org/10.1039/C9NR01246J
UT WoS 000466774500026
Keywords in English Bacteria detection; European foulbrood diagnosis; Upconversion-linked immunosorbent assay; ULISA; Antibody development; Apis mellifera
Tags CF CRYO, CF NANO, CF PROT, rivok
Tags International impact, Reviewed
Changed by Changed by: Mgr. Pavla Foltynová, Ph.D., učo 106624. Changed: 31/3/2020 21:25.
Abstract
European foulbrood (EFB) is an infectious disease affecting honeybee larvae caused by the bacterium Melissococcus plutonius. The enzyme-linked immunosorbent assay (ELISA) is the gold standard for antibody-based bacteria detection, however, its sensitivity is not high enough to reveal early-stage EFB infection. Photon-upconversion nanoparticles (UCNPs) are lanthanide-doped nanomaterials that emit light of shorter wavelength under near-infrared (NIR) excitation and thus avoid optical background interference. After conjugation with specific biorecognition molecules, UCNPs can be used as ultrasensitive labels in immunoassays. Here, we introduce a method for conjugation of UCNPs with streptavidin based on copper-free click chemistry, which involves surface modification of UCNPs with alkyne-modified bovine serum albumin (BSA) that prevents the non-specific binding and provides reactive groups for conjugation with streptavidin-azide. To develop a sandwich upconversion-linked immunosorbent assay (ULISA) for M. plutonius detection, we have prepared a rabbit polyclonal anti-Melissococcus antibody. The specific capture of the bacteria was followed by binding of biotinylated antibody and UCNP–BSA–streptavidin conjugate for a highly sensitive upconversion readout. The assay yielded an LOD of 340 CFU mL-1 with a wide working range up to 10^9 CFU mL-1, which is 400 times better than the LOD of the conventional ELISA. The practical applicability of the ULISA was successfully demonstrated by detecting M. plutonius in spiked real samples of bees, larvae and bottom hive debris. These results show a great potential of the assay for early diagnosis of EFB, which can prevent uncontrolled spreading of the infection and losses of honeybee colonies.
Links
LM2015043, research and development projectName: Česká infrastruktura pro integrativní strukturní biologii (Acronym: CIISB)
Investor: Ministry of Education, Youth and Sports of the CR
LQ1601, research and development projectName: CEITEC 2020 (Acronym: CEITEC2020)
Investor: Ministry of Education, Youth and Sports of the CR
MUNI/A/1575/2018, interní kód MUName: Podpora biochemického výzkumu v roce 2019
Investor: Masaryk University, Category A
TJ01000386, research and development projectName: Imunostanovení pro diagnostiku hniloby včelího plodu
Investor: Technology Agency of the Czech Republic
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