Multi-fluorescence channel detection system for real-time fluorescence quantitative PCR

Abstract

The invention discloses a multi-fluorescence channel detection system for a real-time fluorescence quantitative PCR. The multi-fluorescence channel detection system comprises fluorescence detection units, an optical fiber disc and a turntable, wherein each fluorescence detection unit comprises a light source, an excitation light filter, a dichroscope, optical fiber coupling lenses, optical fibers, a detection light filter and a photoelectric sensor, the dichroscopes enable existing excitation units and detection units to be combined to form a whole, the light emitted by the light source is sequentially subjected to filtration of the excitation light filters and coupling of the optical fiber coupling lenses and finally is emitted into a test tube through the optical fibers to excite the fluorescent substances of samples in the test tubes so as to produce fluorescent light, a part of the fluorescent light returns to the optical fiber coupling lenses through the optical fibers sequentially to be collimated, the detection light filters filter out pure fluorescent light, and the fluorescent light is finally emitted to the photoelectric sensors to perform photoelectric conversion. Multiple optical fibers are inserted into the optical fiber disc, multiple fluorescence detection units are distributed on the turntable, and fluorescence signals of multiple fluorescence channels at multiple test tube hole positions can be sequentially detected through the turntable rotating around the circle center of the optical fiber disc for one circle.

Description

technical field [0001] The invention relates to a real-time fluorescence quantitative PCR detection system, in particular to a multi-fluorescence channel detection system which adopts a fluorescence detection unit combined with an excitation unit and a detection unit for real-time fluorescence quantitative PCR detection. Background technique [0002] In 1996, the AppliedBiosystem company of the United States proposed real-time fluorescent quantitative PCR (real-timeqPCR) on the basis of polymerase chain reaction (Polymerase Chain Reaction, referred to as PCR). In the method, a specific DNA fluorescent probe is added to a PCR reaction system, and the amplification of the target DNA is monitored in real time by collecting the fluorescence intensity of the reaction solution during each temperature cycle. Later, multiplex PCR (MultiplexPCR) was proposed, that is, using the specificity of primers and fluorescent probes binding to target DNA, multiple pairs of primers and multiple...

AI Technical Summary

Problems solved by technology

[0011] 1. Each test tube hole needs 2 optical fibers, and the cost of the optical system is relatively high

[0012] 2. Each test tube hole on the temperature block needs to process 2 holes to insert the optical fiber. When the temperature block has more test tube holes (such as 96 holes), the mechanical structure design and processing of the temperature block are very difficult.

It is difficult to design a temperature block that is small in size and can process so many optical fiber jacks

[0013] 3. If it is expanded to multi-fluorescence channels, twice as many fluorescence excitation and detection units are required to be arranged on the turntable. The volume limitation of the excitation and detection units makes the diameter of the excitation circle and detection circle of the turntable must be increased to accommodate more unit

Then the effective cross-sectional area of ​​the fiber at the excitation end and the emission end becomes half of the existing scheme, and the coupling efficiency of the excitation light and the receiving efficiency of the fluorescence will also decrease by half, resulting in the final fluorescence signal intensity being reduced to four times that of the existing scheme. 1 / 1, which would seriously reduce the signal-to-noise ratio of the fluorescent signal

[0019] 4. It is unavoidable that the expansion of existing solutions to multiple fluorescence channels will increase the diameter of the turntable and increase the fluorescence detection cycle.

[0020] 5. It is impossible to avoid the problem of poor consistency of fluorescence signals in different test tube well positions in the existing scheme

[0021] In summary, the above two existing solutions still cannot fundamentally solve the following problems:

[0022] 1. The contradiction between the number of optical fibers corresponding to a single test tube hole and the signal-to-noise ratio of fluorescence signals

[0023] 2. The contradiction between the number of fluorescent channels and the detection speed

[0024] 3. The problem of poor consistency of fluorescent signals

[0025] These shortcomings make the application of existing technical solutions in the field of real-time fluorescent quantitative PCR greatly limited

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