An electrochemiluminescence imaging device and its application

A technology of electrochemistry and luminescence imaging, which is applied in the direction of chemiluminescence/bioluminescence, measuring devices, and analysis by making materials undergo chemical reactions, which can solve the problems of detection flux limitation and impossibility, and achieve simple composition , increase the flux, promote the effect of luminous efficiency

Active Publication Date: 2016-08-31
NANJING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The disadvantage of this technology is that each cell surface must be detected independently, so the detection throughput is severely limited
Although in the presence of luminol, these released molecules are converted into electrochemiluminescent hydrogen peroxide by its oxidase, but it must be realized in the submicron region, and the sampling time of the second level (time In the range of resolution), it is still impossible to obtain a detectable optical signal for such a low concentration of biomolecules

Method used

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  • An electrochemiluminescence imaging device and its application
  • An electrochemiluminescence imaging device and its application
  • An electrochemiluminescence imaging device and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1 Visual detection of hydrogen peroxide in solution.

[0036] (1) Determination of detection conditions.

[0037] In the experiment, an O-ring with a diameter of 1 cm was pasted on an indium tin oxide (ITO) glass sheet as the solution chamber, and the ITO glass sheet was used as the working electrode, and L012 was selected as the luminescent substance under positive voltage. L012 is a luminol analog, which has a higher luminescence effect than luminol in the presence of hydrogen peroxide, so it was selected as the luminescence substance under positive voltage. 200 μM of L012 and 100 μM of hydrogen peroxide were added dropwise to the O-ring for a total of 600 μL. In view of the fact that the luminescence peak voltage of L012 and hydrogen peroxide on the ITO glass electrode is 1.0V (wherein, 1.0V is the peak voltage of the electrolysis of hydrogen peroxide on the surface of the ITO electrode, and then the generated oxygen radicals make L012 emit light, so for dif...

Embodiment 2

[0043] Example 2 Visual detection of hydrogen peroxide leakage from the surface of single cells.

[0044] Single-cell leakage of hydrogen peroxide imaging was performed by culturing 100-5000 HeLa cells in solution chambers on ITO glass slides and then treated with 10 μl of 20 ng / ml crotyl alcohol-12-tetradecanoate-13-acetate (PMA, Phorbol12 -myristate-13-acetate, referred to as phorbol ester, purchased from sigma-aldrich, item number is P8139) to stimulate the production of hydrogen peroxide. It has been reported that PMA stimulates intracellular NADPA oxidase to accumulate hydrogen peroxide, resulting in the leakage of hydrogen peroxide. According to the same imaging procedure, brightfield and background images in the presence of L012 were obtained ( Figure 5 A and 5B). In the luminescence image, it appears as a darker intensity due to the adhesion of cells to the electrode surface, which hinders the diffusion of L012 to the electrode surface. After the cells released hyd...

Embodiment 3

[0047] Example 3 Electrochemiluminescence imaging of single cell surface molecules.

[0048]In addition to imaging the hydrogen peroxide released by the cells, the electrochemiluminescence imaging method of the present invention can also utilize the oxidase corresponding to the molecules on the cell surface to convert them into hydrogen peroxide, thereby realizing the imaging of the cell surface molecules. To demonstrate that cell surface molecules can be imaged, we chose cholesterol as a template molecule to detect the distribution of activated cholesterol molecules in the cell membrane. Activated cholesterol has a higher chemical potential (tendency to escape), which is important for cholesterol transport across cell membranes. Although cell membrane cholesterol has been imaged using fluorescent cholesterol analogs or flipins, cell-surface activated cholesterol has never been successfully imaged because these fluorescent probes cannot differentiate between activated and non-...

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PUM

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Abstract

The invention discloses an electrochemiluminescent imaging device which comprises an object lens, a light transmission channel, a single-photon detection electron multiplication CCD (charge coupled device) and an electrochemiluminescent generator, wherein the electrochemiluminescent generator comprises an ITO (indium tin oxide) electrode, a silver electrode and a voltage generator; the object lens, the light transmission channel and the single-photon detection electron multiplication CCD are connected in sequence; the ITO electrode is connected with the positive electrode of the voltage generator; the silver electrode is connected with the negative electrode of the voltage generator. The invention further discloses applications of the electrochemiluminescent imaging device to visual detection on hydrogen peroxide, hydrogen peroxide leaking from the surface of single cells and cell surface molecules. The electrochemiluminescent imaging device is simple in composition, high in detection flux, easy to operate and high in luminous efficiency, and can perform the parallel quick detection.

Description

technical field [0001] The invention belongs to the technical field of biochemical industry, and in particular relates to an electrochemiluminescence imaging device and its application. Background technique [0002] The use of electrochemical techniques to image single cells can obtain important biological information such as cell surface chemical components and local activities. Currently, the most common microscopy instrument is electrochemical scanning microscope. The microscope uses a microelectrode to scan the surface of a single living cell to detect electroactive chemical species. By reducing the electrode size to the nanometer level, the spatial resolution of the electrochemical scanning microscope can be achieved to the nanometer level. The disadvantage of this technique is that each cell surface must be detected individually and thus the throughput of detection is severely limited. As an alternative to electrochemical scanning microscopy, recently developed surfa...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/76G01N21/69G01N21/01
Inventor 方丹君江德臣周俊宇
Owner NANJING MEDICAL UNIV
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