研究目的
To develop a high spatial resolution sensor for measuring extracellular pH in proximity of the cell membrane using gold nanoparticles (AuNP) and surface enhanced Raman spectroscopy (SERS), and to test the sensor on HepG2 liver cancer cells and MKN28 gastric cancer cells before and after inhibition of Na+/H+ exchanger.
研究成果
The study successfully developed a method for highly localized pH bio-sensing using SERS and AuNP. The sensor was able to visualize highly localized variations of pH induced by H+ extrusion, particularly in cancer cells. The method demonstrated sensitivity to dynamic variations of proton trafficking, as shown by the increase in cell surface pH after inhibition of Na+/H+ exchanger.
研究不足
The study's limitations include the potential for micro-aggregations of AuNP at the edge of cell clusters, which could affect pH measurements, and the need for further optimization of the protocol for different cell types and conditions.
1:Experimental Design and Method Selection:
The study utilized gold nanoparticles (AuNP) conjugated with 4-mercaptobenzoic acid (4-MBA) and a pyridyldithiol-biotin compound (HPDP-B) for pH sensing. Surface enhanced Raman spectroscopy (SERS) was employed to measure the pH-dependent concentration of deprotonated 4-MBA.
2:Sample Selection and Data Sources:
HepG2 liver cancer cells and MKN28 gastric cancer cells were used.
3:List of Experimental Equipment and Materials:
Gold nanourchins, 4-MBA, HPDP-B, streptavidin, sulfo-NHS-ester-biotin compound (NHS-B), and isotonic buffer solutions at different pH levels.
4:Experimental Procedures and Operational Workflow:
Cells were labelled with NHS-B, treated with streptavidin, and then incubated with conjugated AuNP. SERS and confocal fluorescence microscopy (CFM) were used for analysis.
5:Data Analysis Methods:
SERS spectra were analyzed and deconvoluted using Gaussian-Lorentzian functions after baseline subtraction.
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