Nano probe for long-time parallel detection of activities of multiple proteinases

A protease activity and nanoprobe technology, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of not being able to monitor changes in protease activity in real time for a long time, and not be able to detect protease activity in living cells in parallel. Strong photobleaching effect

Active Publication Date: 2015-07-15
TSINGHUA UNIV
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Problems solved by technology

[0008] In order to overcome the problem that the activity of proteases in living cells cannot be detected in parallel at present, and the protease activity changes cannot be monitored in real time for a long time, the present invention provides a nanoprobe for long-term real-time parallel detection of various protease activities

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  • Nano probe for long-time parallel detection of activities of multiple proteinases
  • Nano probe for long-time parallel detection of activities of multiple proteinases
  • Nano probe for long-time parallel detection of activities of multiple proteinases

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Embodiment Construction

[0022] The invention can be used for long-term real-time parallel detection of various protease activities. For example, when cells undergo apoptosis, caspase (1, 3, 9, etc.) series proteases and CtrB enzymes are activated, and the corresponding probes in the present invention can be designed to perform parallel proteases on various proteases involved in apoptosis. detection. The specific embodiment of the present invention will be further described below in conjunction with the example of detecting intracellular Caspase-3 and Caspase-9 enzyme activities.

[0023] Take 1pmol quantum dot QD605 (CdSe / ZnS quantum dot, emission wavelength is 605nm), each quantum dot is decorated with about 4-8 streptavidin; after connecting with quantum dot, one streptavidin can also Linked with 3 biotins. That is, each quantum dot can be connected with 12-24 biotins.

[0024]In the test, the number of gold nanoparticles is 5-10 times the number of total binding sites of streptavidin; one strep...

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Abstract

The invention belongs to the field of protein detection, and particularly relates to a nano probe capable of long-time parallel detection of activities of multiple proteinases in living cells on the basis of energy transfer. The probe has a penetrating peptide-quantum dot-substrate polypeptide-nano gold particle structure, and can efficiently enter cells in a way which is harmless to the cells. The probe has high photobleaching resistance, and can detect activities of proteinases in the living cells for a long time (up to one month). The probe can reflect the distribution of the proteinases in the cells, and can reflect the activation time of the proteinases to be detected and the activation sequence of the proteinases to be detected in real time. The probe can gather the characteristics of enzymes in the cells and the mutual influence among the enzymes more comprehensively.

Description

technical field [0001] The invention belongs to the field of protein detection, in particular to a nanoprobe based on energy transfer that can be used for long-term real-time parallel detection of various protease activities in living cells. Background technique [0002] In the history of protein development, the study of protease occupies a very important position. It has abundant sources, easy purification, small molecular weight and simple function. All stages of the cell life cycle are closely related to proteases. For example, in the process of cell fusion and differentiation and macromolecular assembly, there are corresponding specific proteases involved, and they play a very important role in biological regulation. At present, the research on the primary structure to the advanced structure of protein and the mechanism of action of enzyme has been at the forefront of the biomedical field. [0003] In 2005, Emmanue et al. from Rice University in the United States devel...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/37
Inventor 任大海尤政王俊王斌
Owner TSINGHUA UNIV
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