Blood cell separation method for portunus trituberculatus
A technology of portunus trituratus and a separation method, which is applied to cell dissociation methods, animal cells, invertebrate cells, etc., can solve the problem of not developing crab blood cell separation technology, etc., and achieves simple operation, high separation efficiency, and good activity. Effect
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Embodiment 1
[0035] A kind of Portunus trituberculatus blood cell separation method that present embodiment proposes, it comprises the following steps:
[0036] Step 1, preparation of cell separation solution:
[0037] 1_1. Prepare a PBS buffer solution with a volume molar concentration of 0.1M.
[0038] 1_2. Using a sodium chloride solution with a volume molar concentration of 3M to adjust the osmotic pressure of the PBS buffer solution to 900 mOs·mol / kg to obtain an isotonic solution. Among them, 10ml of PBS buffer solution and 1.2ml of sodium chloride solution can be taken.
[0039] 1-3, the isotonic solution is used as diluent, and the volume percent concentration of dilution is 60% iodixanol (OptiPrep TM ), three kinds of iodixanol solutions with volume percentage concentration of 10%, 15% and 20% were configured in gradient.
[0040] 1_4. Using the existing cushion technology, add three kinds of iodixanol solutions with volume percentage concentrations of 10%, 15% and 20% respecti...
Embodiment 2
[0054] This embodiment carries out further processing on the basis of the first embodiment. That is: after step 3_2, the clear cell solution in the separated clear cell layer, the small granular cell solution in the small granular cell layer, and the large granular cell solution in the large granular cell layer are washed with cell separation liquid to keep more Good activity, the specific process is as follows: absorb the clear cell solution from the clear cell layer, add isotonic solution whose volume is 3 times the volume of the absorbed clear cell solution, and centrifuge at 1200g for 2.5 minutes at 4°C , to remove the supernatant to obtain a precipitate; then add an isotonic solution with a volume 3 times the volume of the precipitate to the precipitate, perform centrifugation at 1200g for 2.5min at 4°C, remove the supernatant, and obtain an elution The clear cell solution after the cell separation medium; in the same manner, the small granular cell solution after the cel...
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