Detection of Cytochrome c in a Single Cell Using an Optical Nanobiosensor

Abstract

In this work, the intracellular measurement of cytochrome c using an optical nanobiosensor is demonstrated. The nanobiosensor is a unique fiberoptics-based tool which allows the minimally invasive analysis of intracellular components. Cytochrome c is a very important protein to the process which produces cellular energy. In addition, cytochrome c is well-known as the protein involved in apoptosis, or programmed cell death. delta-Aminolevulinic acid (5-ALA) was used to induce apoptosis in MCF-7 human breast carcinoma cells. 5-ALA, a photodynamic therapy (PDT) drug in cells was activated by a HeNe laser beam. After the PDT photoactivation, the release of cytochrome c from the mitochondria to the cytoplasm in a MCF-7 cell was monitored by the optical nanobiosensor inserted inside the single cell and followed by an enzyme-linked immunosorbent assay (ELISA) outside the cell. The combination of the nanobiosensor with the ELISA immunoassay improved the detection sensitivity of the nanobiosensor due to enzymatic amplification. Our results lead to the investigation of an apoptotic pathway at the single cell level.

Plasmonic gold mushroom arrays with refractive index sensing figures of merit approaching the theoretical limit

“…Cyt c is an electron-carrying protein and has a central role in cellular oxidation reactions in animals and plants. It has been extensively studied as a model system for biosensors 39 . The representative reflectance spectra measured when the array was exposed to Cyt c solutions at varying concentrations are shown in Fig.…”

Biomedical applications of nanobiosensors: the state-of-the-art

“…In both cases, the biorecognition process was completed by adding a primary antibody and a secondary antibody with horse radish peroxidase (HRP) as enzyme label; methylene blue was the redox mediator added to the electrolyte solution. 41 Song et al 42 reported that the detection of cytochrome c probed in a single cell using the optical nanobiosensor having mouse anticytochrome c antibodies. They proved that the combination of nanobiosensor with the ELISA immunoassay (enzyme-linked immunosorbent assay) improved the detection sensitivity of the nanobiosensor due to enzymatic amplification.…”

“…The authors suggested that an intimate link between nanoscale sensor structure and biodetection sensitivity will aid the development of high performance diagnostic tools diseases diagnosis. 129 Song et al 42 demonstrated the intracellular measurement of cytochrome c using an optical nanobiosensor. Cytochrome c is an important protein in the process which produces cellular energy.…”

Microfluidic and bio-applications of optical microfibres

“…The sensor response time was of the order of τ~100ms. Biochemical compounds measured in-vivo, in live cells include benzopyrene tetrol and benzo[a]pyrene [18], cytochrome c [19], Caspase-9 [20], a telomerase [21] and DNA sequences [22]. The small sensor size allow for in-situ detection of specific chemicals without visible damage to the cell.…”

“…Ion concentrations were measured by functionalizing the sensor apex with plasticized polymeric membranes embedding ion complexation agents and fluorescent dyes. Potassium [20] and Calcium [23] concentrations were detected using valinomycin and Fura-2/AM or fluo-3/AM calcium-dyes as highly-selective ligands. Other compounds measured in live cells include oxygen [24], nitrites [25], chloride ion [25] nitric oxide [26] and glutamate [27].…”