An all-in-one device for rapid Raman measurement and laser ejection sorting of single cells

A single-cell, laser technology, applied in Raman scattering, material excitation analysis, sorting, etc., can solve the problems of exogenous DNA contamination, limiting sorting flux and efficiency, etc., to avoid external exposure, improve efficiency, reduce effect of possibility

Active Publication Date: 2021-08-24
QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] (1) Cells on the surface of the substrate need to perform Raman measurement in the forward direction, and then flip the substrate for reverse ejection, resulting in the need to re-address and position the corresponding measurement cells before sorting, which severely limits the throughput and efficiency of sorting ;
[0004] (2) The cells in the measurement and sorting are exposed to the external environment, which is very easy to introduce foreign DNA contamination in the subsequent genome amplification operation;
[0005] (3) At present, there is no integrated device that can connect Raman measurement-cell sorting-cell lysis-genome amplification without barriers

Method used

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  • An all-in-one device for rapid Raman measurement and laser ejection sorting of single cells
  • An all-in-one device for rapid Raman measurement and laser ejection sorting of single cells
  • An all-in-one device for rapid Raman measurement and laser ejection sorting of single cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1: Production and effect of laser ejection substrates.

[0049] The substrate of the laser ejection substrate uses a light transparent material including, but not limited to, silicate glass, quartz glass, calcium fluoride glass; the bubble plating film material is a transparent metal composite oxide film material, including but not limited to ITO, FTO, AZO , A plating film is formed on the surface of the base material in the surface of the base material, the thickness of the metal composite oxide film is not less than 70% by deposition of spraying, evaporation, sputtering and other deposition. Figure 1 The light-transmitted comparison of the quartz glass substrate is shown, and the light-transmitting comparison of the ITO film, 1a is a quartz glass substrate, 1B is a laser ejection substrate plated with an ITO film). The laser ejection substrate (1b) plated with a plated film compared to the substrate (1a), the light transmittance is almost no different, and the sin...

Embodiment 2

[0050] Example 2: Sample chip, production and integrated device combination of the receiving chip

[0051] Figure 4 A single-cell laser echo sorting integrated device is schematically illustrated, and the single cell laser echo sorting sorting integrated device includes a laser ejection substrate 1, a point-like chip 2, and a receiving chip 3.

[0052] The point-like chip 2 employs a transparent soft polymer material including, but not limited to, PDMS, silicone rubber, PVC, thickness is made of 0.1-2 mm, and the chip contains an array microchannel 2-1, and the microchannel diameter is 0.5-2.5mm. .

[0053] The receiving chip 3 employs a transparent material including, but not limited to, one or several combinations of silicate glass, PDMS, PMMA, and the chip contains a certain depth of array micro-well 3-1, and the micro-well size is 1-5mm. Its position corresponds to the micropolor position in the above-described point-like chip.

[0054] The monocyte laser echo sorting integra...

Embodiment 3

[0055] Example 3: Single-cell fast Raman measurement and sorting by integrated devices

[0056] Taking marine blue bacteria, a single cell laser echo sorting integrated device is used to perform single-cell fast Raman measurement and sorting method, including the following procedure:

[0057] The spotty chip 2 is brought into laser ejection substrate 1 plating film 1-2 front, laboratory pure culture marine blue bacteria, straw over water after centrifugation and pure water, resuspended in water, take 0.5 μL of cell suspension drip Dotted in the point, the room temperature is dry. The receiving chip 3 is then bonded to the point-like chip 2, and the micrometer 3-1 is integrally formed with the point-like hole one by one. The above single cell laser echo sorting integrated device is placed on the stage of the front-grade co-equivalent, the laser ejection substrate 1 is on, the receiving chip 3 is under, passed through the 50X microscope objective, passed through the laser ejection s...

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Abstract

The invention relates to a laser ejection substrate and an integrated device comprising the substrate for single cell rapid measurement and laser ejection sorting. The integrated device includes a laser ejection substrate, a spotting chip and a receiving chip. The laser ejection substrate, the spotting chip and the receiving chip can be stacked in sequence and bonded reversibly to form an integrated corresponding structure. The invention adopts a fully transparent substrate, which avoids substrate flipping and cell re-addressing, and realizes "immediately measuring and popping" a single cell; adopts a three-layer closed system, avoids external exposure during operation, and reduces external exposure. The possibility of source contamination; the array microwell receiving device is used to improve the efficiency of single cell acquisition and directly connect with downstream operations.

Description

Technical field [0001] The present invention relates to a single cell analysis and a sorting technique, and more particularly to an integrated combination device for realizing rapid measurement of single cells and laser echo sorting. Background technique [0002] Single-cell Raman spectroscopy techniques developed in recent years have a method of reducing FACS technology to a certain extent, to a certain extent, to some extent. Single-cell Raman ejection technology is based on laser-inducing forward transfer (LIFT) principle, and the cells are placed on the ejection substrate, and the surface of the substrate is plated with a layer of diamond material, which can be rapidly peeled or melted under the action of pulsed laser. Thereby, the cells attached to the surface are separated from the substrate. Since the cell position is fixed, a single cell can be observed or Raman measurement for a single cell, and the correctness of the screening of the target cells can be improved. But th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/65B07C5/34B07C5/36
CPCB07C5/34B07C5/36G01N21/65
Inventor 勾洪磊荆晓艳苏晓璐籍月彤徐健
Owner QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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