Surface-enhanced Raman scattering (SERS)-based immunosystem for ultrasensitive detection of the 90K biomarker

Abstract

The research and the individuation of tumour markers in biological fluids are currently one of the main tools to support diagnosis, prognosis, and monitoring of the therapeutic response in oncology. Although the identification of tumour markers in asymptomatic patients is crucial for early diagnosis, its application is still limited by the relatively low sensitivity and the complexity of existing methods (i.e. ELISA, mass spectrometry). We developed an easy, fast, and ultrasensitive surface-enhanced Raman scattering (SERS)-based system, for the detection and quantitation of the LGALS3BP (90K) biomarker that was used as a model, based on the development of antibody-functionalized nanostructured gold surfaces. The detection system was effective for the ultrasensitive detection and characterization of samples of different biochemical compositions. In conclusion, this work could provide the foundation for the development of a medical diagnostic device with the highest predictive power when compared with the methods currently used in cancer diagnostics.

Keywords: Bioanalytical methods; Biomarkers; LGAL3BP; SERS substrates; Surface-enhanced Raman spectroscopy (SERS).

Fig. 1

Two-step functionalization of SERS substrates. a A heterobifunctional linker mediates the covalent binding of anti-90K capture antibody to the gold SERS active surface of nanostructured substrates; the active SERS surface (in orange) is made of stochastic nanostructure feature fragments from tens of nanometres to few microns nanostructured gold. b Schematic representation of Raman scattering after the interaction of the system with the 90K antigen

Fig. 2

SEM images show the different levels of aggregation of AuNPs depending on the functionalization state. a Non-functionalized AuNPs. b LA(PEG)3Mal AuNPs. c, d Anti-90K capture (1959cr) antibody-functionalized AuNPs

Fig. 3

PCA of Raman spectra from substrates analyzed before and after antibody functionalization and in presence of the 90K antigen. PCA data shows a clear separation between empty-Matos, 1959Cr-MatoS, and 90K-1959Cr-MatoS

Fig. 4

PCA of Raman spectra at diverse time of 90K antigen exposure. There is a clear separation between 1959Cr-Matos and 90K-1959Cr-Matos already after 5 min of incubation with the antigen

Fig. 5

PCA of Raman spectra of oncologic serum samples. Data refers to 1919Cr-MatoS analyzed before the interaction with oncologic sera (M1 and M2) and after the incubation with two diverse serum dilutions (M1; 10⁻⁶) and (M2; 10⁻⁹)

Fig. 6

PCA of Raman spectra of healthy serum samples. Data shows the lack of a clear separation between 1959Cr-MatoS before and after the incubation with two diverse healthy serum dilutions, indicating that the presence of other serum components does not overlap with 90K detection