The Detection And Analysis Of Lung Cancer Cells Based On Gold Nanostar SERS Substrates

As a booming analytical technology with great potential, surface-enhanced Raman scattering (SERS) spectroscopy has been well applied in in biology, chemistry and medicine, mainly attributing to its fingerprint characteristic, stability, resistance to photo bleaching, noninvasive detection and the Raman signal of water is very weak. The application of SERS mainly depends on SERS active substrates, and gold nanoparticles are usually used to act as SERS substrates because of their easy synthesis, relatively low toxicity and good stability. Based on SERS active substrates, SERS can highly enhance the Raman signals of target species as much as 14 orders of magnitude and even allow the signal detection at single-molecule level. Among all gold nanoparticles, gold nanostars with tunable NIR plasmon and multiple sharp branches have been reported to show stronger SERS enhancement than gold nanorods and gold nanospheres under similar experimental conditions. To achieve higher enhancement effect, gold nanostars can be assembled by taking advantage of the electrostatically assisted APTES-functionalized surface-assembly method. Because of high SERS enhancement effect, gold nanostars has attracted great attention. This paper reports the preparation of gold nanostars via seed-mediated growth approach and then preparation of gold nanostars substrates with higher enhancement effect, homogeneous SERS ctivity and high reproducibility via electrostatic self-assembly method. Based on the obtained gold nanostar substrates, a sensitive and efficient SERS immunoassy system for carcinoembryonic antigen (CEA) detection has been presented. Beside, the obtained substrates also enable us to successfully compare and distinguish two lung cancer cell lines [human lung adenocarcinoma epithelial cell line (A549), human non-small cell lung cancer cell line (H1229)] and one normal lung cell line [human type Ⅱ alveolar epithelial cell line (AT Ⅱ)] through analysis and comparison of SERS spectra.The details are as follows:1. The preparation and characterization of gold nanostar SERS substratesBased on a seed-mediated growth approach, gold nanostars with uniform size and shape have been prepared, and related SERS spectra shows their high enhancement effect. Then, taking advantage of electrostatically assisted APTES-functionalized surface-assembly method, gold nanostars were assembled to get active SERS substrates, and different assembly time for substrates was studied. Besides, we also studied some characters of substrates, including uniformity, stability and reproducibility, and the results show gold nanostar SERS substrates were prepared with higher enhancement effect, homogeneous SERS activity and high reproducibility, which indicates the suitable application of substrates in detection of proteins and living cells.2. The detection of carcinoembryonic antigen based on gold nanostar SERS substratesBased on the obtained substrates, a sensitive and efficient SERS immunoassy system for carcinoembryonic antigen (CEA) detection has been presented. On the one hand, the CEA antibody molecules are linked to the gold nanostars labeled with 4-mercaptobenzoic acid (MBA). On the other hand, the CEA antigen molecules with different concentrations are respectively immobilized on gold nanostar substrates modified with dimercaptosuccinic acid (DMSA). When the gold nanostar SERS probes (Gold nanostar@MBA@CEA antibody) are present, they can conjugate with the CEA antigen molecules of gold nanostar immunoassay substrates (Gold nanostar substrate@DMSA@CEA). As a result, the Raman signal of MBA used as Raman reporter and linking group is greatly amplified due to the double SERS enhancement of gold nanostar SERS probes and gold nanostar immunoassay substrates, which is also influenced by the concentration of CEA antigen molecules. Based on the SERS immunoassay, a broad linear range(10 pg/mL to 1μg/mL) with a correlation coefficient (R2) of 0.994 is exhibited.3. The detection and dfferentiation of lung cancer cell lines and normal cell line based on gold nanostar SERS substratesAfter the cleaning and asepsis processing of gold nanostar SERS substrates, we successfully compare and distinguish two lung cancer cell lines [human lung adenocarcinoma epithelial cell line (A549), human non-small cell lung cancer cell line (H1229)] and one normal lung cell line [human type II alveolar epithelial cell line (AT II)] through SERS spectra. The difference spectra (A549-H1229, A549-AT II and H1229-AT II) among the three cell lines are studied to better interpret the spectral differences and gain insight into the biochemical variation. Furthermore, principal component analysis (PCA) score plots of the SERS spectra are also used to better view the differences among these three cell lines. This sensitive, efficient and non-invasive detection method, based on gold nanostar substrates, will have great potential for the differentiation of living cells and play an important role in the early diagnosis and treatment of cancers.