Single-molecule upconversion-linked digital immunosensing of
cancer biomarkers

a 2018

Single-molecule upconversion-linked digital immunosensing of cancer biomarkers

FARKA, Zdeněk, Matthias J. MICKERT, Antonín HLAVÁČEK, Hans H. GORRIS, Petr SKLÁDAL et. al.

Základní údaje

Originální název

Single-molecule upconversion-linked digital immunosensing of cancer biomarkers

Autoři

FARKA, Zdeněk, Matthias J. MICKERT, Antonín HLAVÁČEK, Hans H. GORRIS a Petr SKLÁDAL

Vydání

70. sjezd českých a slovenských chemických společností, 2018

Další údaje

Jazyk

angličtina

Typ výsledku

Konferenční abstrakt

Obor

10406 Analytical chemistry

Stát vydavatele

Česká republika

Utajení

není předmětem státního či obchodního tajemství

Organizační jednotka

Středoevropský technologický institut

ISSN

Klíčová slova anglicky

single-molecule analysis; upconversion-linked immunosorbent assay; cancer biomarker; prostate specific antigen; photon-upconversion nanoparticle

Změněno: 22. 3. 2019 15:14, Mgr. Pavla Foltynová, Ph.D.

Anotace

V originále

The ability to detect cancer biomarkers at the single-molecule level promises the ultimate sensitivity in clinical diagnosis. Fluorescence-based single-molecule analysis, however, is limited by matrix interferences, and therefore is typically not suitable for real world analytical applications. This optical background can be elegantly avoided by using photon-upconversion nanoparticles (UCNPs) that emit shorter-wavelength light under near-infrared excitation (anti-Stokes emission). Conjugates of UCNPs with antibody can be used as a background-free luminescent tracer in upconversion-linked immunoassays. We have developed an optical approach for visualizing individual UCNPs and applied it for the sensitive detection of the cancer biomarker prostate specific antigen (PSA). An inverted wide-field microscope was equipped with a fiber-coupled 980 nm continuous-wave laser diode, 100× heat-resistant objective and a sensitive sCMOS camera. Silica-coated UCNPs with carboxyl groups on the surface were synthesized and conjugated with anti-PSA antibody. The structure and monodispersity of the nanoconjugates was studied in detail by transmission electron microscopy and agarose gel electrophoresis. Individual sandwich immunocomplexes consisting of (1) immobilized anti-PSA antibody, (2) PSA, and (3) anti-PSA antibody-UCNP conjugate were counted under an upconversion wide-field microscope equipped with a 980 nm laser excitation source. The noise-surpassing digital readout provided single particle resolution with almost no instrumental background. This allowed to reach a limit of detection (LOD) of 1.2 pg mL−1 (42 fM) of PSA in 25 % blood serum which is about ten times more sensitive than a classical (analog) readout of luminescence intensity. The digital signal readout is a completely new approach with the potential to pave the way for a new generation of immunoassays.

Návaznosti

LQ1601, projekt VaV

Název: CEITEC 2020 (Akronym: CEITEC2020)

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, CEITEC 2020

Citovat

FARKA, Zdeněk, Matthias J. MICKERT, Antonín HLAVÁČEK, Hans H. GORRIS a Petr SKLÁDAL. Single-molecule upconversion-linked digital immunosensing of cancer biomarkers. In 70. sjezd českých a slovenských chemických společností. 2018. ISSN 2336-7202.

@proceedings{1438256,
   author = {Farka, Zdeněk and Mickert, Matthias J. and Hlaváček, Antonín and Gorris, Hans H. and Skládal, Petr},
   booktitle = {70. sjezd českých a slovenských chemických společností},
   keywords = {single-molecule analysis; upconversion-linked immunosorbent assay; cancer biomarker; prostate specific antigen; photon-upconversion nanoparticle},
   language = {eng},
   title = {Single-molecule upconversion-linked digital immunosensing of cancer biomarkers},
   year = {2018}
}

TY  - CONF
ID  - 1438256
AU  - Farka, Zdeněk - Mickert, Matthias J. - Hlaváček, Antonín - Gorris, Hans H. - Skládal, Petr
PY  - 2018
TI  - Single-molecule upconversion-linked digital immunosensing of cancer biomarkers
KW  - single-molecule analysis
KW  - upconversion-linked immunosorbent assay
KW  - cancer biomarker
KW  - prostate specific antigen
KW  - photon-upconversion nanoparticle
N2  - The ability to detect cancer biomarkers at the single-molecule level promises the ultimate sensitivity in clinical diagnosis. Fluorescence-based single-molecule analysis, however, is limited by matrix interferences, and therefore is typically not suitable for real world analytical applications. This optical background can be elegantly avoided by using photon-upconversion nanoparticles (UCNPs) that emit shorter-wavelength light under near-infrared excitation (anti-Stokes emission). Conjugates of UCNPs with antibody can be used as a background-free luminescent tracer in upconversion-linked immunoassays. We have developed an optical approach for visualizing individual UCNPs and applied it for the sensitive detection of the cancer biomarker prostate specific antigen (PSA). An inverted wide-field microscope was equipped with a fiber-coupled 980 nm continuous-wave laser diode, 100× heat-resistant objective and a sensitive sCMOS camera. Silica-coated UCNPs with carboxyl groups on the surface were synthesized and conjugated with anti-PSA antibody. The structure and monodispersity of the nanoconjugates was studied in detail by transmission electron microscopy and agarose gel electrophoresis. Individual sandwich immunocomplexes consisting of (1) immobilized anti-PSA antibody, (2) PSA, and (3) anti-PSA antibody-UCNP conjugate were counted under an upconversion wide-field microscope equipped with a 980 nm laser excitation source. The noise-surpassing digital readout provided single particle resolution with almost no instrumental background. This allowed to reach a limit of detection (LOD) of 1.2 pg mL−1 (42 fM) of PSA in 25 % blood serum which is about ten times more sensitive than a classical (analog) readout of luminescence intensity. The digital signal readout is a completely new approach with the potential to pave the way for a new generation of immunoassays.
ER  -

FARKA, Zdeněk, Matthias J. MICKERT, Antonín HLAVÁČEK, Hans H. GORRIS a Petr SKLÁDAL. Single-molecule upconversion-linked digital immunosensing of cancer biomarkers. In \textit{70. sjezd českých a slovenských chemických společností}. 2018. ISSN~2336-7202.

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