文档介绍：PRIMARY CULTURE OF MOUSE SPLEEN CELLS & IDENTIFICATION OF LIVE OR DEAD CELLS
BY DIFFERENTIAL STAINING
Qianqian Chen(1e muscle layer open in a V shape and lift up to expose peritoneal cavity.
4. Push aside the stomach in the upper-left side of the abdomen to expose the sack-shaped, dark-red spleen. Cut off the spleen with scissors and put it into the glass dish.
5. Inject PBS with the L-shape needle into the spleen along the longer axis. Spleen will bloat and become pale. * Not to penetrate the spleen.
6. Punch some holes with the needle on the spleen and scratch it so that spleen cells will flow out to the PBS. At the end, spleen goes from red to yellow and PBS will turn into red. Discard the remaining of the spleen.
7. Blow the remaining PBS with spleen cells with pipette several times to dispense cell clumps into individual cells.
8. Transfer cell suspension to a microcentrifuge tube. Add PBS to 1ml total volume and spin at 500g for 5 min.
9. Discard supernatant. Resuspend cell pellet with 1ml cell culture medium and transfer to the plastic culture dish, which has already 2 ml .
Rock the fish to separate cells evenly.
Culture in the incubator at 37°C for one to two weeks.
Observe cultured cells by naked eyes and under the Inverted Microscope .
Tap the dish to release attached cells and swirl to the dish to resuspend all cells .
Take ml of