Bo Yin,Sha Sha Li,Shao Hua Lu,Jia Ying Mi,Hong Lin Zhai
Although a large number of fluorescent probes have been developed, the simultaneous quantitative analysis of intracellular thiols is still difficult due to the spectral overlap and the complexity of the intracellular environment. In this study, a multi-signal fluorescent probe was employed for the simultaneous quantification of intracellular glutathione (GSH), cysteine (Cys) and homocysteine (Hcy). As the feature variables of the target components, the Tchebichef image moments (TMs) calculated from the grayscale images of the 3D fluorescence spectra were used to establish the quantitative linear models by stepwise regression. The intra-day and inter-day precisions of the proposed method were less than 5.6% and 8.7%, respectively. The recoveries ranged from 97.0% to 105.9%. In addition, the proposed approach was applied to the simultaneous quantitative determination of Cys, GSH and Hcy in the MCF-10A cell (a type of normal cell) and MDA-MB-231 cancer cell. The obtained results indicated that the concentrations of the three thiols in the cancer cell were higher than those in the normal cell. This study not only provides an effective approach for the quantification of multi-target bio-molecules in complicated intracellular environments, but also further extends the applications of multi-signal fluorescent probes, which will promote the design of new multi-signal fluorescent probes.
Chunrong Li,Yuzhi Xu,Si-Yang Liu,Yanfei Zhang,Wen Yin,Zong Dai,Xiaoyong Zou
Ascorbic acid (AA) and glutathione (GSH), the most abundant intracellular reductive substances, have been widely used as biomarkers for cancer cells identification. The current methods relying on imaging of AA or GSH alone to identify cancer cells may cause systematic errors, since a mutual conversion relationship exists between AA and GSH. In this work, we propose a fluorescent nanosensor for the simultaneous imaging of intracellular reductive substances including AA and GSH. Biocompatible fluorescent silicon nanoparticles (SiNPs) with rich surface amine and carboxyl groups were synthesized. The fluorescence of the SiNP was initially quenched by chelation of Fe 3+ ions, forming SiNP/Fe 3+ complex as the fluorescent nanosensor. Upon the redox reaction with reductive substances, the nanosensor showed sensitively fluorescent recovery. Moreover, benefited from the efficient cellular uptake of the SiNP/Fe 3+ and the overexpressed intracellular reductive substances in cancer cells, the fluorescent nanosensor was used to accurately identify the human breast carcinoma (MCF-7) cells from normal mammary epithelial (MCF-10A) cells by imaging of intracellular AA and GSH simultaneously. This strategy would be promising in imaging-guided precision cancer diagnosis.
Accompanying the occurrence of inflammatory reaction to release cytokines, pyroptosis can activate an immune response for resistance against cancer. Consequently, elevated levels of cytokines released by cancer cells are highly correlated with the effectiveness of cancer treatment. Herein, a noninvasive surface-enhanced Raman spectroscopy (SERS) immunosensor was developed to sensitively and specifically measure the tumor necrosis factor-α (TNF-α), a proinflammatory cytokine, during the cell pyroptosis process. The sandwiched structure of the sensor is functionalized with a TNF-α binding antibody for detecting TNF-α at concentrations as low as 1 pg/mL. Importantly, electrical stimulation (ES) can fleetly trigger cancer cell pyroptosis to induce the overexpression of receptor interacting protein 3 (RIP3), which is a significant protein that regulates the inflammatory response. The overexpression of RIP3 can activate caspase-1 to promote the upregulation of cytokine levels. Notably, the cytokine levels of TNF-α released from cancer cells (MCF-7 cells) were apparently higher than those of normal cells (MCF-10A cells) during pyroptosis detected by the SERS immunosensors. Due to its obvious superiorities of simple fabrication and fast readout without sample pretreatment, the developed SERS platform has a potential application value for diagnosis and treatment of cancer.
危险品化学品经营许可证（带存储）