Homogeneous Immunosorbent Assay Based on Single-Particle Enumeration Using Upconversion Nanoparticles for the Sensitive Detection of Cancer Biomarkers

Abstract

Prostate-specific antigen (PSA) is an intercellular glycoprotein produced primarily by the prostate gland, which is commonly chosen as the initial target for the early diagnosis of prostate cancer. In this work, we demonstrate a simple yet sensitive sandwich-type single-particle enumeration (SPE) immunoassay for the quantitative detection of PSA in a flow chamber. The design is based on the luminescence resonance energy transfer (LRET) between upconversion nanoparticles (UCNPs) and gold nanoparticles (GNPs). The carboxyl group-functionalized UCNPs are conjugated with anti-PSA detection antibodies (Ab) and serve as the luminescence energy donor, while GNPs are modified with anti-PSA capture antibodies (Ab) and act as the energy acceptor. In the presence of target antigen (i.e., PSA), the specific immnuoreaction brings the donor and acceptor into close proximity, resulting in quenched luminescence. Through statistical counting of the target-dependent fluorescent particles on the glass slide surface, the quantity of antigens in the solution is accurately determined. The dynamic range for PSA detection in Tris-buffered saline (TBS) is 0-500 pM and the limit-of-detection (LOD) is 1.0 pM, which is much lower than the cutoff level in patients' serum samples. In the serum sample assay, comparable LOD was also achieved (i.e., 2.3 pM). As a consequence, this method will find promising applications for the selective detection of cancer biomarkers in clinical diagnosis.

Nanotechnology: a promising method for oral cancer detection and diagnosis

“…Routine measurement methods-including enzyme-linked immunosorbent assay (ELISA), immunohistochemistry, Western Blot, and polymerase chain reaction-still bear a limited detection sensitivity ranging from pM to fM (10 −12 to 10 −15 M) concentration levels [ 22 , 23 , 35 ]. The application of nanotechnology may enhance the detection sensitivity for biomarkers with low concentrations in the tissue samples or body fluids [ 122 , 123 ].…”

Detection and monitoring prostate specific antigen using nanotechnology approaches to biosensing

“…A strategy in a similar fashion but relying on the quenching of up-conversion nanoparticles functionalized with anti-PSA antibodies in a sandwich type immunoassay has been reported. The single particle counting in a microscope setup allowed for PSA limits of detection as low as 1 pmol$L -1 [72].…”

Advances in Optical Single‐Molecule Detection: En Route to Supersensitive Bioaffinity Assays

“…Recently,wehave prepared ad etection label based on ac onjugate of PEGcoated UCNPs with streptavidin, which allowed to decrease the label concentration and further improved the LOD by afactor of 50 to % 20 fg mL À1 . [22b] Li et al [101] designed ad igital homogeneous sandwich immunosorbent assay based on UCNPs (NaYF 4 :Yb 3+ ,Er 3+ , 42 nm in diameter) for PSA. An antibody-coated UCNP, PSA, and an antibody-coated Au NP formed an immunocomplex.…”