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Cell Navigator® Fluorimetric Lipid Droplet Assay Kit *Green Fluorescence*

Lipid droplets, also referred to as lipid bodies, oil bodies or adiposomes, are lipid-rich cellular organelles that regulate the storage and hydrolysis of neutral lipids. They also serve as a reservoir of lipid source for many important biological processes such as fatty acid and cellular cholesterol for energy and membrane formation and maintenance. Abnormal accumulation of the cytoplasmic lipid droplets occurs in a variety of pathological conditions and can be an indicator of metabolic deficiency or pathogenesis. AAT Bioquest's Cell Navigator® Fluorimetric Lipid Droplets Assay Kit is a simple assay that could quantitatively measure lipid droplet accumulation. Nile Green™ is used in the kit for lipophilic stain. Nile Green™ is intensely fluorescent in a lipid-rich environment while it has minimal fluorescence in aqueous media. It is an excellent vital stain for the detection of intracellular lipid droplets with fluorescence microscopy, flow cytometry or fluorescence microplate reader. Unlike Nile Red which has broad range of fluorescence spectrum, Nile Green™ stains intracellular lipid droplets with green fluorescence only. It can be used with other fluorescence dyes for multicolor staining. The green fluorescence signal could be observed using the filter set of FITC.

Example protocol

AT A GLANCE

Protocol summary

  1. Prepare cells with test compounds
  2. Add Nile Green™ working solution
  3. Incubate at room temperature or 37°C for 10 to 30 min
  4. Read fluorescence intensity with fluorescence microscope using FITC filter

Important notes
Following is our recommended protocol for live cells. This protocol only provides a guideline, and should be modified according to your specific needs. Since Nile Green™ has minimal fluorescence in aqueous media, aspiration of the growth medium and removal of Nile Green™ staining solution after staining is optional. Stained cells can be fixed with 3 - 4% formaldehyde. In addition, prefixed cells (3 - 4% formaldehyde fixation) can be stained with Nile Green™ staining solution.

PREPARATION OF WORKING SOLUTION

Prepare Nile Green™ working solution by diluting 5 µL of 200X Nile Green™ (Component A) to 1 mL of Staining Buffer (Component B). Note: 50 µL of Nile Green™ (Component A) is enough for one 96-well plate. Protect from light. The optimal concentration of the Nile Green™ varies depending on specific applications. The staining conditions may be modified according to a particular cell type and the permeability of the cells or tissues to the probe.

For guidelines on cell sample preparation, please visit
https://www.aatbio.com/resources/guides/cell-sample-preparation.html

SAMPLE EXPERIMENTAL PROTOCOL

For adherent cells:

  1. Grow cells either in a 96-well black wall/clear bottom plate (100 µL/well/96-well) or on cover-slips inside a petri dish filled with the appropriate culture medium.

  2. Gently aspirate the culture medium and add equal volume (such as 100 µL/well/96-well plate) of the Nile Green™ staining solution.

  3. Incubate the cells in a 37°C, 5% CO2 incubator for 10 - 30 minutes.

  4. Remove Nile Green™ working solution (Optional).

  5. Read Fluorescence at 485/520 nm with a microplate reader or observe the cells using a fluorescence microscope with a FITC filter set.

For suspension cells:

  1. Centrifuge the cells at 1000 rpm for 5 minutes to get 1 - 5 × 105 cells per tube.

  2. Resuspend cells in 500 µL of Nile Green™ working solution.

  3. Incubate at room temperature or 37°C for 10 to 30 min, protected from light.

  4. Centrifuge to remove the Nile Green™ working solution, and resuspend cells in 500 µL of pre-warmed medium or buffer of your choice to get 1 - 5 × 105 cells per tube (Optional).

  5. Monitor the fluorescence increase using fluorescence microscope with a FITC filter set.

Spectrum

Product family

NameExcitation (nm)Emission (nm)Extinction coefficient (cm -1 M -1)Quantum yield
Cell Navigator® Fluorimetric Lipid Droplet Assay Kit *Red Fluorescence*559635380000.70001

Citations

View all 4 citations: Citation Explorer
Pathological analysis of Prader-Willi syndrome using adipocytes
Authors: Kishimura, Urara and Soeda, Shuhei and Ito, Daiki and Ueta, Yoko and Harada, Maki and Tanaka, Mai and Taniura, Hideo
Journal: Biochemical and Biophysical Research Communications (2024): 150124
Exploration and Validation of Ferroptosis-Associated Genes in ADAR1 Deletion-Induced NAFLD through RNA-seq Analysis
Authors: Yin, Xuecui and Mi, Yang and Wang, Xiaohan and Li, Ya and Zhu, Xiaohui and Bukhari, Ihtisham and Wang, Qingde and Zheng, Pengyuan and Xue, Xia and Tang, Youcai
Journal: International Immunopharmacology (2024): 112177
ABHD5-CPT1B: An Important Way of Regulating Placental Lipid Metabolism in Gestational Diabetes Mellitus
Authors: Yang, Yuqi and Peng, Yue and Yu, Bin and Wang, Huiyan
Journal: Archives of Medical Research (2024): 102925

References

View all 26 references: Citation Explorer
Live cell imaging and analysis of lipid droplets biogenesis in HCV infected cells
Authors: Nevo-Yassaf, I.; Lovelle, M.; Nahmias, Y.; Hirschberg, K.; Sklan, E. H.
Journal: Methods (2017)
Lipid droplets and associated proteins in sebocytes
Authors: Schneider, M. R.
Journal: Exp Cell Res (2016): 205-8
and beyond
Authors: Wang, C. W., Lipid droplets, lipophagy
Journal: Biochim Biophys Acta (2016): 793-805
Acute accumulation of free cholesterol induces the degradation of perilipin 2 and Rab18-dependent fusion of ER and lipid droplets in cultured human hepatocytes
Authors: Makino, A.; Hullin-Matsuda, F.; Murate, M.; Abe, M.; Tomishige, N.; Fukuda, M.; Yamashita, S.; Fujimoto, T.; Vidal, H.; Lagarde, M.; Delton, I.; Kobayashi, T.
Journal: Mol Biol Cell (2016): 3293-3304
Expression of hepatic lipid droplets is decreased in the nitrofen model of congenital diaphragmatic hernia
Authors: Takahashi, H.; Kutasy, B.; Friedmacher, F.; Takahashi, T.; Puri, P.
Journal: Pediatr Surg Int (2016): 155-60
Page updated on October 6, 2024

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Catalog Number22730
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Spectral properties

Correction Factor (260 nm)

0.015

Correction Factor (280 nm)

0.018

Extinction coefficient (cm -1 M -1)

81000

Excitation (nm)

504

Emission (nm)

510

Storage, safety and handling

H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
UNSPSC12352200

Platform

Fluorescence microscope

ExcitationFITC filter set
EmissionFITC filter set
Recommended plateBlack wall, clear bottom

Fluorescence microplate reader

Excitation485 nm
Emission520 nm
Cutoff510 nm
Recommended plateBlack wall, clear bottom
Instrument specification(s)Bottom read mode

Components

Fluorescence images of intracellular lipid droplets in control (Left) and Oleic Acid treated HeLa cells (Right) using Cell Navigator® Lipid Droplets Fluorescence Assay Kit. HeLa cells were incubated with 300 uM of Oleic Acid for 24 hours to induce intracellular lipid droplets formation. After washing with PBS, the cells were labeled with 1X Nile Green™ and Hoechst 33342  (Cat#17533).
Fluorescence images of intracellular lipid droplets in control (Left) and Oleic Acid treated HeLa cells (Right) using Cell Navigator® Lipid Droplets Fluorescence Assay Kit. HeLa cells were incubated with 300 uM of Oleic Acid for 24 hours to induce intracellular lipid droplets formation. After washing with PBS, the cells were labeled with 1X Nile Green™ and Hoechst 33342  (Cat#17533).
Fluorescence images of intracellular lipid droplets in control (Left) and Oleic Acid treated HeLa cells (Right) using Cell Navigator® Lipid Droplets Fluorescence Assay Kit. HeLa cells were incubated with 300 uM of Oleic Acid for 24 hours to induce intracellular lipid droplets formation. After washing with PBS, the cells were labeled with 1X Nile Green™ and Hoechst 33342  (Cat#17533).