| Lung cancer is the highest numbers of deaths which can reach a quarter of all cancers. Early clinical diagnosis is very significant and essential in improving the survival rate and evaluating the recovery of cancer patients. Many technologies are developed for lung cancer detection, however they suffer the disadvantage of invading, expensive, time-consuming and low sensitivity. Recently, a new immunoassay technique based on the surface-enhanced Raman scattering (SERS) by antibody conjugated metal nanoparticles has attracted much attention from many groups due to its rapid and sensitive sensing capability for biological analysis. The most critical aspect of performing the SERS-based immunoassay is fabrication of SERS tags. In our work, the novel monodispersed HGNs were embedded with 4-mercapobenzonic acid to form SERS tags and magnetic microspheres were synthesized as enrichment substrates. Based the sandwich immunoassay strategy, CEA in the level of 10 pg/mL-100 ng/mL have been detected with a highly specific, sensitive.The details are as follows:(1) A novel monodispersed HGNs are fabricated by seed-mediated methods. In the seed-mediated method, small gold nanoparticles are pre-formed on the surface of the monodispersed SiO2 to form SiO2/GNPs-nanocomposite particles (were obtained from Nissian Chemical,～110 nm with a coefficient of variation of 2%) as deposition sites for gold produced by reduction. As more gold is reduced by H2O2, the GNPs grow larger and formation of a continuous shell layer. By HF corroding the silica core of GNSs, monodispersed and high SERS enhance HGNs are obtained. Although this method suffers the disadvantage of complex process and time-consuming, the obtained GNSs are much smoother with better controllability and reproducibility. Compare the SERS spectrum for 4-MBA-labeled GNSs with HGNs, the result displays that the Raman intensity of 4-MBA labeled HGNs at 1080 cm-1 is 10 times higher than that of GNSs, which indicating the excellent SERS enhancement effect of HGNs.(2) HGNs were obtained by seed-mediated methods with better controllability, monodispersity and repeatability. Firstly,4-MBA was used as a Raman reporter, also a donor of carboxyl group for conjugating detection antibody by active ester method to structure SERS tags. Secondly, 2-Mercaptoethanol was used as protector to reduce nonspecific binding and aggregation. Finally, detection antibody were conjugated to the SERS tags. On the other hand, about 1 μm of magnetic microspheres were fabricated and coated with silica to conjugate capture antibody as capture and separation substrates. The FT-IR spectra provided evidence that McAb-conjugated HGNs tags and CEA PcAb-conjugated magnetic microspheres have been successfully prepared.(3) Based on this surface enhanced raman immunoassays, CEA McAb-conjugated HGNs-based SERS tags and CEA PcAb-conjugated magnetic microspheres were used to detect CEA up to 100 ng/mL with a detection limit of 10 pg/mL, which was two orders of magnitudes lower than that of GNSs-based SERS tags. The results show that a very good linear response in the lower concentration range between detection concentration and Raman signal intensity. The monodispersed HGNs were desirable SERS tags for quantitative sandwich immunoassay assay, showing strong potential for the clinical diagnosis of cancer biomarkers. |
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