EdU Labeling and Immunofluorescent Staining of Drosophila Larval Wing Imaginal Disc

材料与试剂

1. 各种规格枪头 (配套移液枪)
2. 各种规格EP管
3. 载破片
4. 盖破片
5. 三龄果蝇幼虫
6. NaCl (国药沪试，catalog number: 10019318)
7. KCl (生工，catalog number: PB0440)
8. Na₂HPO₄ (生工，catalog number: S0404)
9. KH₂PO₄ (生工，catalog number: PB0445)
10. Schneider昆虫培养基 (Sigma, catalog number: S9895)
11. 8% PFA溶液 (8% Paraformaldehyde aqueous solution) (Electron MicroscopySciences, catalog number: 157-8)
12. Triton X-100 (生工，catalog number: A110694-0100)
13. Bovine serum albumin (BSA) (生工，catalog number: A600332-0100)
14. 一抗
15. 荧光二抗
16. DAPI (Sigma, catalog number: D9542)
17. 抗淬灭剂：VECTASHIELD^® Antifade Mounting Medium (VECTOR, catalog number: H-1000)
18. ddH₂O
19. Click-iT^TM EdU Alexa Fluor^TM 555 Imaging Kit (Invitrogen, catalog number: C10338)
20. 1× PBS, pH 7.4 (见溶液配方)
21. SD溶液 (见溶液配方)
22. 4% PFA溶液 (见溶液配方)
23. 0.1% PBT (见溶液配方)
24. 0.5% PBT (见溶液配方)
25. 3% BSA blocking buffer (见溶液配方)
26. Click-iT^TM EdU Alexa Fluor^TM 555 Imaging Kit (见溶液配方)
27. DAPI溶液 (见溶液配方)
    

仪器设备

1. 剪刀
2. 体式显微镜 (Nikon)
3. 精细解剖镊× 2
4. 解剖板
5. 各量程移液枪
6. 滚轴 (海门市其林贝尔仪器制造公司，SRT-202滚轴式混合器)
7. 激光共聚焦显微镜 (Olympus, FV1000)
   

实验步骤