Digital detection of subfemtomolar concentrations of
prostate-specific antigen by single molecule immunosensing

a 2021

Digital detection of subfemtomolar concentrations of prostate-specific antigen by single molecule immunosensing

GORRIS, Hans-Heiner, Matthias Jürgen MICKERT, Zdeněk FARKA, Uliana KOSTIV, Antonín HLAVÁČEK et. al.

Basic information

Original name

Digital detection of subfemtomolar concentrations of prostate-specific antigen by single molecule immunosensing

Authors

GORRIS, Hans-Heiner, Matthias Jürgen MICKERT, Zdeněk FARKA, Uliana KOSTIV, Antonín HLAVÁČEK, Daniel HORÁK and Petr SKLÁDAL

Edition

31st Anniversary World Congress on Biosensors, 2021

Other information

Language

English

Type of outcome

Konferenční abstrakt

Field of Study

10406 Analytical chemistry

Country of publisher

Czech Republic

Confidentiality degree

není předmětem státního či obchodního tajemství

Organization unit

Faculty of Science

Abstract

V originále

The ability to detect single molecules holds the promise to reach the ultimate sensitivity in analytical chemistry. Several problems, however, must be addressed before single-molecule techniques can be employed for routine analytical applications. Photon-upconversion nanoparticles (UCNPs) are excellent labels for single molecule immunoassays because they emit short-wavelength light under near-infrared (NIR, 980 nm) excitation (anti-Stokes emission), which avoids autofluorescence and light scattering. These unique photoluminescent features enable the detection of UCNPs at the single nanoparticle level by conventional wide-field epiluminescence microscopy. Furthermore, very homogeneous UCNP labels were prepared by gel electrophoretic separation. For the upconversion-linked immunoassay (ULISA), microtiter plates were first coated with anti-prostate-specific antigen (PSA) antibodies to capture PSA. PSA was then detected either by an anti-PSA antibody-UCNP conjugate or a biotinylated anti-PSA antibody in combination with a streptavidin-PEG UCNP conjugate. The PSA concentration was determined on the same microtiter plate either (1) by using an upconversion microtiter plate reader (analog mode) or (2) by counting individual immunocomplexes under an upconversion wide-field microscope (digital mode). The lowest limit of detection was achieved by using the streptavidin-PEG-neridronate conjugate as a detection label. Counting single analyte molecules improved the limit of detection (LOD) (23 fg mL-1, 800 aM) by more than one order of magnitude compared to the analogue readout.

Links

GA21-03156S, interní kód MU

Name: Foton-upkonverzní značky pro mikrofluidní jednomolekulové imunostanovení proteinových biomarkerů

Investor: Czech Science Foundation

LQ1601, research and development project

Name: CEITEC 2020 (Acronym: CEITEC2020)

Investor: Ministry of Education, Youth and Sports of the CR

LTAB19011, research and development project

Name: Luminiscenční imunostanovení jako citlivý nástroj pro diagnostiku nemocí včel

Investor: Ministry of Education, Youth and Sports of the CR, Sensitive bioanalytical tools for the surveillance of honey bee diseases, INTER-ACTION

Citovat

GORRIS, Hans-Heiner, Matthias Jürgen MICKERT, Zdeněk FARKA, Uliana KOSTIV, Antonín HLAVÁČEK, Daniel HORÁK and Petr SKLÁDAL. Digital detection of subfemtomolar concentrations of prostate-specific antigen by single molecule immunosensing. In 31st Anniversary World Congress on Biosensors. 2021.

@proceedings{1784817,
   author = {Gorris, HansandHeiner and Mickert, Matthias Jürgen and Farka, Zdeněk and Kostiv, Uliana and Hlaváček, Antonín and Horák, Daniel and Skládal, Petr},
   booktitle = {31st Anniversary World Congress on Biosensors},
   language = {eng},
   title = {Digital detection of subfemtomolar concentrations of prostate-specific antigen by single molecule immunosensing},
   year = {2021}
}

TY  - CONF
ID  - 1784817
AU  - Gorris, Hans-Heiner - Mickert, Matthias Jürgen - Farka, Zdeněk - Kostiv, Uliana - Hlaváček, Antonín - Horák, Daniel - Skládal, Petr
PY  - 2021
TI  - Digital detection of subfemtomolar concentrations of prostate-specific antigen by single molecule immunosensing
N2  - The ability to detect single molecules holds the promise to reach the ultimate sensitivity in analytical chemistry. Several problems, however, must be addressed before single-molecule techniques can be employed for routine analytical applications. Photon-upconversion nanoparticles (UCNPs) are excellent labels for single molecule immunoassays because they emit short-wavelength light under near-infrared (NIR, 980 nm) excitation (anti-Stokes emission), which avoids autofluorescence and light scattering. These unique photoluminescent features enable the detection of UCNPs at the single nanoparticle level by conventional wide-field epiluminescence microscopy. Furthermore, very homogeneous UCNP labels were prepared by gel electrophoretic separation. For the upconversion-linked immunoassay (ULISA), microtiter plates were first coated with anti-prostate-specific antigen (PSA) antibodies to capture PSA. PSA was then detected either by an anti-PSA antibody-UCNP conjugate or a biotinylated anti-PSA antibody in combination with a streptavidin-PEG UCNP conjugate. The PSA concentration was determined on the same microtiter plate either (1) by using an upconversion microtiter plate reader (analog mode) or (2) by counting individual immunocomplexes under an upconversion wide-field microscope (digital mode). The lowest limit of detection was achieved by using the streptavidin-PEG-neridronate conjugate as a detection label. Counting single analyte molecules improved the limit of detection (LOD) (23 fg mL-1, 800 aM) by more than one order of magnitude compared to the analogue readout.
ER  -

GORRIS, Hans-Heiner, Matthias Jürgen MICKERT, Zdeněk FARKA, Uliana KOSTIV, Antonín HLAVÁČEK, Daniel HORÁK and Petr SKLÁDAL. Digital detection of subfemtomolar concentrations of prostate-specific antigen by single molecule immunosensing. In \textit{31st Anniversary World Congress on Biosensors}. 2021.

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