Catalytic Gold Deposition for Ultrasensitive Optical Immunosensing of Prostate Specific Antigen

Abstract

A major challenge in the development of bioanalytical methods is to achieve a rapid and robust quantification of disease biomarkers present at very low concentration levels in complex biological samples. An immunoassay platform is presented herein for ultrasensitive and fast detection of the prostate-specific antigen (PSA), a well-recognized cancer biomarker. A sandwich type immunosensor has been developed employing a detection antibody labeled with inorganic nanoparticles acting as tags for further indirect quantification of the analyte. The required high sensitivity is then achieved through a controlled gold deposition on the nanoparticle surface, carried out after completing the recognition step of the immunoassay, thus effectively amplifying the size of the nanoparticles from nm to µm range. Due to such an amplification procedure, quantification of the biomolecule could be carried out directly on the immunoassay plates using confocal microscopy for measurement of the reflected light produced by gold-enlarged nanostructures. The high specificity of the immunoassay was demonstrated with the addition of a major abundant protein in serum (albumin) at much higher concentrations. An extremely low detection limit for PSA quantification (LOD of 1.1 fg·mL−1 PSA) has been achieved. Such excellent LOD is 2–3 orders of magnitude lower than the clinically relevant PSA levels present in biological samples (4–10 ng·mL−1) and even to monitor eventual recurrence after clinical treatment of a prostate tumor (0.1 ng·mL−1). In fact, the broad dynamic range obtained (4 orders of magnitude) would allow the PSA quantification of diverse samples at very different relevant levels.

Post-Assay Chemical Enhancement for Highly Sensitive Lateral Flow Immunoassays: A Critical Review

“…Gold enhancement was initially developed to increase small GNPs’ visibility in immunohistochemistry [ 47 ]. Since then, gold enhancement has been widely utilized in various bioanalytical techniques [ 48 , 49 , 50 ]. Focusing on LFIA, gold enhancement is used to increase the detectability of GNP in the test zone by increasing their size.…”

Bio-Tailored Sensing at the Nanoscale: Biochemical Aspects and Applications

“…Further examples must be recalled that take advantage of using proteins and nanomaterials coupled to optical methods and or characteristics to reveal analytes, as for instance those reported for a multiplex lateral flow lateral flow assay device onto nitrocellulose membrane and antibodies-functionalized AuNSs for detection of metalloproteinase-9, S100 calcium-binding protein B and neuro-specific enolase in blood [ 117 ], antibodies-functionalized magnetic Fe 3 O 4 NPs capped with a gold shell to detect the human growth hormone in plasma [ 118 ] and antibodies-conjugated Mn-ZnS QDs for analyte-induced gold amplification for detection of prostate-specific antigen in serum [ 119 ].…”

Optical Microscopy Systems for the Detection of Unlabeled Nanoparticles

“…Finally, Cid-Barrio et al used CRM for ultrasensitive quantification of disease biomarkers in complex biological samples. 56 To this end, they developed a sandwich immunoassay platform for the detection of prostate specific antigen (PSA), a cancer biomarker. PSA was bound by capture antibodies and its detection was realized by antibodies coupled to Mn-ZnS quantum dots (QDs) or gold nanoparticles.…”