Cultivation device and method for adherent cell thin membrane of laboratory

Abstract

The invention relates to a laboratory adherent cell thin film culture device and the method of the field of biological engineering technology, and the device includes a flat dish, an organic polymer thin film layer and an adhesive liquid layer. The organic polymer thin film layer is adhered at the inner bottom surface of the flat dish. The polymer material thin film is wetted with the adhesive liquid to be adhered at the bottom surface of a culture dish as an adherent cell attachment surface, so as to facilitate the rapid separation of the cells during the passage, the cells are rapidly put into a volumetric culture liquid and then are exfoliated by swinging method, further to carry out counting and passage based on this, the accurate component ensures the parallel initial inoculation conditions of the next generation; compared with the original passage method, the operation is more simple, and the bacteria contamination opportunities are fewer. The different polymer thin films can beattached for matching different needs, so as to improve digestive efficiency or survival rate of the adherent culture.

Description

technical field [0001] The invention relates to a device and a method in the technical field of bioengineering, in particular to a device and a method for culturing adherent cell membranes in a laboratory. Background technique [0002] The routine adherent cell culture and subculture method in the laboratory is roughly as follows. Take the Vero cell line inoculated on a 65mm plate as an example, add a small amount of cell suspension inoculated to about 2ml of culture medium, shake it gently and place it in a CO2 incubator at 37°C When the cells grow to a certain density, the cells are subcultured. When the cells are subcultured, first suck out the medium, wash it twice with 1-2ml of PBS, add 1ml of trypsin or collagenase, incubate in the incubator for 5-10min, then add serum-containing medium to stop the digestion, gently pipette repeatedly , until the cells on the plate fall off completely, and the bottom of the plate becomes smooth and translucent, use a pipette to absorb...

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Problems solved by technology

[0003] From the above steps, it can be seen that there are still many disadvantages for experiments that strictly require a low probability of bacterial infection, parallel culture of the next generation, and batch passage: (1) There are certain technical requirements for the pipetting operation in the process of cell digestion, which is likely to cause late The experimental error is not easy to predict

(2) The efficiency of pipetting at different positions of the plate is different, and the result of subculture is related to the difference caused by the uneven spreading of the cells at the time of inoculation, and the cells may not fall off evenly due to the different emphasis of random pipetting

(4) When there is a large concentration difference, cell diffusion is easier to proceed, and repeated blowing and blowing generally use high-concentration cell suspensions, which will cause greater damage to the cells. If the suspension is transferred after each blowing and blowing, and then fresh culture medium, the cost increases many times, it takes many times more sterile pipettes, and the chance of contamination and contamination of pure medium is doubled

(5) Although the operation is carried out in a sterile ultra-clean bench, the traditional pipetting method causes the cells to be exposed for a long time, especially for cells with good adherence. Sometimes it takes about ten pipetting times per square centimeter to rinse the plate until it is smooth. The effect of cleanness and complete detachment of cells is needed, so effective methods are needed to reduce the exposure of cells and increase the safety of the culture and passage process

At present, there is a lack of the least link and the most direct measurement method

(7) If it is a glass plate, the cleaning work is slightly difficult

[0004] After searching the existing technical literature, it was found that "Preparation of chitosan / polyethylene glycol succinate film and its relationship with "Compatibility of Myofibroblasts" has been mentioned, but film culture has not been systematically proposed as a new and commonly used cell culture device or method in the laboratory. In addition, Zhao Dicheng et al. People's "Separation of adhering cell colonies with a direct digestion method" article, cells are seeded on the fibrin membrane, this idea and its benefits (facilitating direct digestion and effectively obtaining single cell clones) are very consistent with the present invention, but The material is not conducive to popularization and production, and cannot be used as a conventional new device in the laboratory

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