An in vitro screening method for transplantation hydrogel microspheres
A technology of hydrogel microspheres and screening methods, which is applied in the field of biomedicine, can solve problems such as poor stability and repeatability, inconsistent conclusions, and high experimental costs, and achieve high repeatability, low experimental cost, and high controllability. Effect
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Embodiment 1
[0021] 1. Add 0.8ml of autoclaved 2% agarose solution to a 12-well plate (22.1 mm in diameter), let it stand still, and automatically solidify at the bottom of the orifice plate after cooling.
[0022] 2. Add the prepared sodium alginate-chitosan microspheres with a particle size of 400 μm into the well plate, and add 50 μL to each well.
[0023] 3. After the mouse fibroblasts with a confluence of 80% in the culture flask were digested and centrifuged, the medium (1640 medium, wherein 20 mM hepes was used as a buffer system) was resuspended to 1×10 6 cells / ml, add 1ml cell suspension to each well.
[0024] 4. Place the orifice plate at 37°C, 5% (V / V) CO 2 On a two-dimensional shaker in an incubator with / 95% (V / V) air, vibrate at a speed of 160 rpm for 10 min every 30 min 4 h before culturing. After static culture, observation under the microscope after 12 hours found that there were more inflammatory cell adhesions on the surface of the microspheres, and the cells all showe...
Embodiment 2
[0028] 1. Add 1ml of autoclaved 1.5% agarose solution to a 12-well plate (diameter 22.1mm), let it stand still, and automatically solidify at the bottom of the well plate after cooling.
[0029] 2. Add the prepared sodium alginate-chitosan microspheres with a particle size of 400 μm into the well plate, and add 50 μL to each well.
[0030] 3. Digest and centrifuge the mouse fibroblasts with a confluence of 100% in the culture flask, and resuspend the culture medium to 1×10 6 cells / ml, add 1ml cell suspension to each well.
[0031] 4. Place the orifice plate at 37°C, 5% (V / V) CO 2 On a two-dimensional shaker in an incubator with / 95% (V / V) air, vibrate at a speed of 160 rpm for 10 min every 30 min 4 h before culturing. After static culture, observation under the microscope after 12 hours found that there were more inflammatory cell adhesions on the surface of the microspheres, and the cells all showed a stretched adhesion state (such as figure 1 B).
[0032] 5. According to...
Embodiment 3
[0035] 1. Add 1ml of autoclaved 1.5% agarose solution to a 12-well plate (diameter 22.1mm), let it stand still, and automatically solidify at the bottom of the well plate after cooling.
[0036] 2. Add the prepared sodium alginate-chitosan-sodium alginate gel microspheres with a particle size of 400 μm into the well plate, and add 50 μL to each well.
[0037] 3. Digest and centrifuge the mouse fibroblasts with a confluence of 100% in the culture flask, and resuspend the culture medium to 1×10 6 cells / ml, add 1ml cell suspension to each well.
[0038] 4. Place the orifice plate at 37°C, 5% (V / V) CO 2 On a two-dimensional shaker in an incubator with / 95% (V / V) air, vibrate at a speed of 160 rpm for 10 min every 30 min 4 h before culturing. After static culture, observation under a microscope after 12 hours found that there was basically no inflammatory cell adhesion on the surface of the microspheres (such as figure 1 C).
[0039] 5. According to the cell adhesion scoring ru...
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