Preparation method of cartilage cell primer layer, and cartilage tissue
A chondrocyte and cartilage tissue technology, applied in the field of cartilage tissue engineering and the preparation of chondrocyte primer layers, can solve the problems of high price, restrict the development and application of tissue engineered cartilage, etc., and achieve good adaptability and good biocompatibility. Effect
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
preparation example Construction
[0023] A method for preparing a chondrocyte primer layer, which is obtained by carrying out a three-dimensional dynamic co-culture of a carrier and an autologous chondrocyte; the culture conditions of the three-dimensional dynamic co-culture include: 36.5-37.5°C, 4.5-5.5% CO 2 , 9~11%O 2 The culture system of the three-dimensional dynamic co-culture includes 10ng / mL basic fibroblast growth factor; before the three-dimensional dynamic co-culture, the density of the autologous chondrocytes in the carrier is 1.6~2.0×10 5 / cm 2 The carrier includes bone cement; the composition of the three-dimensional dynamic co-culture culture system includes: DMEM medium, 4-5mM L-glutamyl, serum with a volume fraction of 9-11%, and double antibiotics.
[0024] In some embodiments, the three-dimensional dynamic co-cultivation is performed in a bioreactor.
[0025] In some embodiments, the culture conditions of the three-dimensional dynamic co-culture can also be: 37°C, 5% CO 2 , 10%O 2 .
[...
Embodiment
[0078] 1. Chondrocyte Biopsy Sampling
[0079] 1.1 Under "absolute" aseptic conditions, 4-5 mm of cartilage tissue was sampled from the lateral patellar groove biopsy, and placed in culture medium I.
[0080] 1.2 Cut the cartilage tissue into small pieces in a petri dish, 1x2mm in size, put it into a 250mL empty flask, and wash it twice with PBS.
[0081] 1.3 Add 40mL pronase solution, and gently stir the cartilage tissue for 30 minutes at 37°C (110rpm).
[0082] 1.4 Take it out from the pronase, place it in 40mL 200U / mL collagenase at 37 degrees for 18-24 hours, and stir gently (110rpm).
[0083] 1.5 Filter the suspension with a 70μm sieve and put it into a 50mL Falcon tube. Centrifuge (300xg) for 10 minutes at room temperature.
[0084] 1.6 Discard the supernatant, and wash the cell ball twice with PBS.
[0085] 1.7 Resuspend the cells in an appropriate amount (20-50mL) of culture medium II.
[0086] 1.8 Take 10 μL of cell suspension, dilute and stain with 90 μL trypan ...
experiment example
[0109] Immunofluorescence staining experiment of cartilage tissue.
[0110] The cartilage tissue prepared by the present invention and the source cartilage were sliced separately, and the densities of the two were observed and compared after immunofluorescent staining. Visible, the prepared cartilage tissue density of the present invention ( figure 2 ) and source cartilage ( figure 1 ) are very close in density.
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More