A kind of application of insect cell immunosuppressant
An immunosuppressant and insect cell technology, which is applied in the field of natural microorganism-derived active product compounds, can solve the problems of no insecticidal activity of the second instar armyworm and no significant impact on the immune system, and achieves enhanced infectious death and good pest control. and pest physiological adaptive regulation, the effect of good application prospects
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Embodiment 1
[0029] Example 1 Spiral ketal compound 5'-epi-SPA-6952A inhibits insect cell viability
[0030] Experimental Materials:
[0031]Diptera Drosophila melanogaster (Drosophila melanogaster) Schneider's S2 cells were purchased from the Institute of Cells, Chinese Academy of Sciences. The spiro ketal compound 5'-epi-SPA-6952A was prepared by referring to the method disclosed in the patent document CN101684448A. TNM-FH insect cell culture medium was purchased from HyClone Company. Trypsin and penicillin (100UI) / streptomycin (100mg / mL) mixed antibiotics were purchased from Gibico. Thiazolium blue (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide, MTT) was purchased from Sigma.
[0032] experimental method:
[0033] (1) Take Tn-5B1-4 cells in the logarithmic growth phase and adjust the cell density to 5×10 4 Cells / mL were seeded in 96-well culture plates until the cell monolayer grew to 80% confluence.
[0034] (2) Add 100 μL of the test compound solution to the expe...
Embodiment 2
[0041] Example 2 Spiral ketal compound 5'-epi-SPA-6952A induces apoptosis in insect cells
[0042] Experimental Materials:
[0043] Lepidoptera insect Trichoplusia ni Tn-5B1-4 cells were purchased from the Institute of Cells, Chinese Academy of Sciences. Annexin V-FITC Apoptosis Detection Kit and propidium iodide (PI) were purchased from Shanghai Beyotime Institute of Biotechnology. FACS Calibur flow cytometer is a product of Becton Dickinson Company of the United States.
[0044] experimental method:
[0045] (1) Inoculate 1×10 5 cell / mL Tn-5B1-4 cells in the logarithmic growth phase were placed in a 6-well plate until the cell monolayer reached 80% confluence.
[0046] (2) In the experimental group, 4 mL of the test compound solution was added to the final concentrations of 0.008, 0.016, and 0.032 μg / mL. In the control group, 4 mL of TNM-FH cell culture medium was added. Set more than 3 parallel wells in each group, place at 37°C, 5% CO 2 and saturated humidity incuba...
Embodiment 3
[0053] Example 3 Spiro ketal compound 5'-epi-SPA-6952A induces morphological changes in insect hemocytes
[0054] Experimental Materials:
[0055] TNM-FH insect cell culture medium was purchased from HyClone Company. Giemsa stain was purchased from Sigma.
[0056] experimental method:
[0057] (1) Collection and in vitro culture of blood cells. After the body surface of the 5th instar Spodoptera litura larvae was washed with distilled water, it was disinfected with 75% alcohol. The gastropod was punctured with a sterile needle, and the hemolymph was collected in an Eppendorf centrifuge tube containing a small amount of phenylthiourea crystals, mixed gently, and kept in an ice bath for later use.
[0058] (2) In vitro culture of blood cells. Take 20 μL of hemolymph in a 24-well cell culture plate, add TNM-FH insect medium, and then place it in a constant temperature incubator at 28°C for cultivation.
[0059] (3) In the experimental group, 20 μL of the test compound solut...
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