ABDP 493/503  别名：中性脂滴荧光探针

ABDP 493/503 (同BODIPY 493/503) 是一种亲脂性荧光探针，定位在极性脂上，能用于标记细胞中性脂内容物，特别定位在活细胞和固定细胞的脂滴上。ABDP 493/503 与落射荧光显微镜、共聚焦荧光显微镜和双质子荧光显微镜，以及流式细胞仪兼容。最大激发和发射波长分别是493/503nm，适用于活细胞和固定细胞标记。

储存液的制备和保存

1）将低温保存的 ABDP 493/503 置于室温回温约20min，低速离心后加入一定量的无水乙醇或无水DMSO配制成适量浓度的母液，比如5mM（充分溶解即可）。根据单次用量将储存液分装，≤-20℃避光干燥保存。需注意溶液内湿度的逐渐积累会随着时间引起探针聚集，从而务必干燥保存储存液。

2）如需长期保存，可以用无水乙醇溶解粉末，之后分装到小量，使用真空泵来挥发掉乙醇。置于≤-20℃避光干燥保存。

BODIPY染色用于流式细胞仪（摘自公开文献，仅供参考）

1. Grow cells under culture conditions relevant for the study. A 35 mm dish/well is sufficient for the cell numbers required in this assay. For our assays, 50,000 A498 cells in 35 mm well were sufficient. Overnight incubation of cells with 30 μM oleic acid can serve as a positive control for increased neutral lipid content, as oleic acid is a potent inducer of triglyceride synthesis and storage. Fatty acid free BSA serves as a control.

2. At the time-point of interest, prepare 2 μM BODIPY staining solution in PBS. The volume of staining solution required for each sample corresponds to the volume of media used for culturing cells.

3. Wash cells with a quick rinse using 3 ml PBS to remove media/serum.

4. Incubate on BODIPY staining solution in the dark for 15 min at 37 °C. Include an unstained control for flow cytometry.

Note: From this point, protect samples from light as much as possible.

5. Wash cells with a quick rinse using 3 ml PBS to remove staining solution.

6. Trypsinize cells to generate a single cell suspension. For the A498 cell line used in this protocol, cells were incubated with Trypsin-EDTA (0.25%) for 5 min at 37 °C.

7. Add 5 ml of PBS and transfer cell suspension to a 15 ml conical tube.

8. Pellet cells at 250 × g, 5 min, 4 °C.

9. Aspirate supernatant, wash the cell pellet with a quick rinse using 3 ml PBS, and pellet cells at 250 × g, 5 min, 4 °C.

10. Carefully aspirate the supernatant and resuspend cells in 300 μl 1× flow cytometry buffer.

11. Pass cell suspension through a 35 μm filter into a FACS tube.

12. Perform flow cytometry. Obtain a minimum of 10,000 events per condition.

13. The investigator can analyze data as mean fluorescence or display the data as a histogram.