Large-area digital PCR microsphere fluorescence high-flux detection device and method

Abstract

The invention discloses a large-area digital PCR microsphere fluorescence high-flux detection device, which comprises a fluorescence exciting light path and a fluorescence detection light path, wherein along the fluorescence exciting light path, exciting light emitted by a light source device sequentially passes through a focusing device and an objective lens device to be emitted onto a digital PCR chip of an objective table; along the fluorescence detection light path, the fluorescence generated by the digital PCR chip being excited by the exciting light sequentially passes through the objective lens device, the focusing device and an imaging device to be imaged and is then transmitted to an image collection sensor; the objective table is also provided with a temperature control device; the temperature control device controls the circulation temperature of the reaction of the digital PCR chip in real time. The invention discloses a large-area digital PCR microsphere fluorescence high-flux detection method. The higher detection efficiency and the large-area high-flux detection accuracy can be obtained. The method and the device can realize the real-time digital qPCR detection with high flux, high sensitivity and high specificity; the important significance is realized on improving the performance, the precision and the flux of a digital PCR detection instrument.

Description

technical field [0001] The invention relates to the field of nucleic acid detection in the field of biomedical testing, in particular to a device and method capable of realizing large-area digital PCR droplet fluorescence high-throughput detection. Background technique [0002] In fields such as early diagnosis and screening of tumors, blood virus typing and load analysis, and noninvasive prenatal diagnosis, high-speed, high-sensitivity, and high-specificity detection of ultra-low-concentration nucleic acids is required, and conventional PCR is increasingly unable to meet the requirements. need. In order to achieve this function, people have developed a digital PCR technology on the basis of PCR, which improves the detection sensitivity by 1 to 2 orders of magnitude. [0003] The "divide and conquer" detection strategy adopted by droplet digital PCR uses the sample and PCR reagent mixture as the dispersed phase, oil as the continuous phase, and uses microfluidic technology ...

AI Technical Summary

Problems solved by technology

[0005] (1) The flow detection speed is slow, the detection speed is 200-500 pieces / s, the detection process takes a long time, and the throughput is low;

[0006] (2) End-point detection can only be performed on a single droplet, and real-time qPCR is not possible, which reduces specificity and sensitivity;

[0007] (3) The optical path hardware is complicated, and it is difficult to realize multiple digital PCR detection (more than 4);

[0008] (4) The transfer process of the two droplets will result in the loss of droplets and possible external pollution;

[0009] (5) The thermal inertia of the heating block of the conventional PCR amplification instrument is large, and the thermal cycle time is long (about 2 hours)

However, existing detection devices such as fluorescence microscopes cannot meet the above detection requirements.

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