Function of lipid droplets
脂滴 (LD) 由中性脂质组成，例如三酰基甘油和胆固醇酯，它们被磷脂单层包围，并且无处不在，不仅在脂肪细胞中 1)。 虽然 LDs 被简单地认为是一种脂质储存单元，但最近的一项研究表明，LDs 在调节脂质代谢、自噬2) 和细胞衰老3) 方面发挥着重要作用。因此，作为阐明其机制的重要工具，LDs 受到了极大的关注 LD的形成、生长、融合和收缩。
1) T. Fujimoto et al., “Lipid droplets: a classic organelle with new outfits.” Histochem Cell Biol., 2008, 130(2), 263.2) R. Singh et al., “Autophagy regulates lipid metabolism.” Nature, 2009, 458(7242), 1131.3) M. Yokoyama et al., “Inhibition of endothelial p53 improves metabolic abnormalities related to dietary obesity.” Cell Reports, 2014, 7(5), 1691.
Only by the addition of a reagent, the imaging of lipid droplets (LDs) or the quantitative variation of LDs in live and fixed cells becomes quantifiable.
Lipid Droplet Assay Kit 大大缩短了整个过程，可用于活细胞。Lipid Droplet Assay Kit 中提供的荧光染料可用于活细胞和固定细胞。 与使用比色试剂的方法相比，使用Lipid Droplet Assay Kit的方法可以缩短测量时间。 此外，使用 Lipid Droplet Assay Kit 可以提高实验的重复性，因为染料不会沉积在板中。
Experimental example of plate assayChanges in lipid droplets were examined after the addition of oleic acid or Triacsin C (acyl-CoA synthetase inhibitor) to the A549 cell culture medium.
As a result, we confirmed that the oleic acid-treated cells show an increase in the number of LDs, compared to control and Triacsin C-treated cells.
Blue ：Ex. 376 – 386 nm / Em 435 – 455 nmDeep Red ：Ex. 623 – 633 nm / Em 649 – 669 nm
Reagent ComparisonChanges in lipid droplets were examined after the addition of oleic acid or Triacsin C (acyl-CoA synthetase inhibitor) to the HeLa cell culture medium.As a result, we confirmed that the oleic acid-treated cells show an increase in the number of LDs, compared to control and Triacsin C-treated cells.
<Detection Condition>Blue ：Ex. 405 nm/ Em 425 – 475 nmDeep Red ：Ex. 640 nm/ Em 650 – 670 nm
Related Product Information
| Function | Product Code | Product | Size |
| Imaging | LD01 | Lipi-Blue | 10 nmol |
| LD02 | Lipi-Green | 10 nmol | |
| LD03 | Lipi-Red | 100 nmol | |
| LD04 | Lipi-Deep Red | 10 nmol | |
| Quantification (Plate Reader, FCM) | LD05 | Lipi Droplet Assay Kit-Blue | 1 set |
| LD06 | Lipi Droplet Assay Kit-Deep Red | 1 set |
Yes, Lipi-dye can be used for fixed cells.* Please use paraformaldehyde (PFA) for fixation. Alcohol fixation is not recommended because it may affect the structure of lipid droplets.* Depending on the cell, it may not be stained or weakened in sensitivity due to fixed conditions before and after staining. In that case, please consider fixed conditions.○ Fix cells after staining1. Remove the medium and wash twice with PBS.2. Add Lipi series Working solution (in PBS) in the cells and incubated at 37 ℃ for 30 minutes.3. Remove the supernatant and wash twice with PBS.4. Add 4% paraformaldehyde (PFA) /PBS solution to the cells and incubate at room temperature for 5 minutes.5. Remove the supernatant and wash with PBS.○ Fix cells before staining1. Remove the medium and wash twice with PBS.2. Add 4% paraformaldehyde (PFA) /PBS solution to the cells and incubate at room temperature for 5 minutes.3. Remove the supernatant and wash twice with PBS.4. Add Lipi-dye Working solution(in PBS) in the cells and incubated at 37 ℃ for 15 minutes.5. Remove the supernatant and wash with PBS.
Preparing a stock solution of oleic acidRequired Reagents:・BSA (bovine serum albumin)・Oleic acid・0.1 mol/L Tris-HCl (pH 8.0)Procedure:(1) Dissolve 0.14 g/mL BSA in 0.1 mol/L Tris-HCl (pH 8.0).(2) Add 4 mmol/L oleic acid to a disposable centrifuge tube.(3) Add BSA solution (prepared in step 1).(4) Cap the tube and mix on a rotary shaker (Be sure the solution is transparent, indicating that oleic acid has been conjugated to BSA).(4) Filter the solution prepared above (step 4) using 0.22μm filter membranes.(5) Store oleic acid stock solution at 4˚C.*Use the appropriate amount of oleic acid stock solution for culture medium to prepare working solution.*Oleic acid working solution cannot be stored. Please prepare the working solution immediately before usage.
Inducing lipid droplets(1) Incubate cells for 24 hours at 37˚C in a 5% CO2 atmosphere.(2) Add 200 µmol/L working solution (prepared from oleic acid stock solution) to culture medium and incubate for further 24 hours.