Publication details
Single molecule upconversion-linked immunosorbent assay with extended dynamic range for the sensitive detection of diagnostic biomarkers
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| Year of publication | 2018 |
| Type | Conference abstract |
| MU Faculty or unit | |
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| Description | Detecting disease markers at the single molecule level has the potential to drastically improve the sensitivity in clinical diagnosis. Prostate specific antigen (PSA) is a reliable protein marker for prostate cancer, the most common form of cancer among men. After surgeries like radical prostatectomy PSA levels are very low. Detecting small increases in PSA concentration shortly after surgery helps to take the necessary steps to save the patient’s life. High photostability, near infrared excitation and biocompatibility after silanization make upconverting nanoparticles (UCNPs) a powerful tool for single molecule immunoassays. A conventional epifluorescence microscope (Nikon Eclipse Ti) was equipped with a 980 nm continuous wave laser diode (4 W) and a 100x objective with a NA of 1.49. Immunocomplexes consisting of (1) an anti-PSA capture antibody immobilized on the bottom of a microwell, (2) PSA, and (3) a silanized UCNP (NaYF4:Yb3+,Er3+) conjugated with a second anti-PSA antibody were counted under the microscope and additionally scanned in a microplate reader equipped with a 980 nm laser. The single molecule (digital) upconversion-linked immunosorbent assay (ULISA) reached a limit of detection of 1.2 pg ml-1 (42 fM) in 25% blood serum, which is about ten times more sensitive than the analog readout and commercial ELISAs. The dynamic range covered three orders of magnitude. The simple design and high sensitivity make this assay a pioneer for a new generation of digital immunoassays. |
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