Nile red (Nile blue oxazone) is a lipophilic stain. Nile red has environment-sensitive fluorescence. Nile red is intensely fluorescent in a lipid-rich environment while it has minimal fluorescence in aqueous media. Nile red is an excellent vital stain for the detection of intracellular lipid droplets by fluorescence microscopy and flow cytof uorometry. Nile red stains intracellular lipid droplets red. The fluorescence wavelength is 559/635 nm^[1].

1.1 Preparation of stock solution
Use DMSO to prepare 1 mM stock solution.
1.2 Preparation of working solution
Use preheated serum-free cell culture medium or PBS to dilute the stock solution to a final concentration of 200-1000 nM.
Note: Please adjust the concentration of Nile Red working solution according to the actual situation and prepare it before use.
2. Cell staining
2.1 Suspended cells: Collect cells by centrifugation and wash twice with PBS for 5 minutes each time.
Adherent cells: Discard the culture medium and add trypsin to digest the cells. After centrifugation and discarding the supernatant, wash twice with PBS for 5 minutes each time.
2.2 Add 1 mL Nile Red working solution and incubate at room temperature for 5-10 minutes.
2.3 Centrifuge at 400 g, 4°C for 3-4 minutes and discard the supernatant.
2.4 Wash cells twice with PBS for 5 minutes each time.
2.5 Resuspend the cells in 1 mL serum-free medium or PBS and observe under a fluorescence microscope.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

When Nile red-stained Caenorhabditis elegans is viewed for green fluorescence, discrete lipid bodies can be observed throughout the intestine and other tissues either in clusters or evenly dispersed, depending on the animal's genotype or experimental treatment^[3].

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

318.37

C₂₀H₁₈N₂O₂

7385-67-3

Solid

Green to dark green

598

543

O=C1C2=CC=CC=C2C3=NC4=CC=C(N(CC)CC)C=C4OC3=C1

Room temperature in continental US; may vary elsewhere.

4°C, protect from light

*In solvent : -80°C, 2 years; -20°C, 1 year (protect from light)

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 2 years; -20°C, 1 year (protect from light). When stored at -80°C, please use it within 2 years. When stored at -20°C, please use it within 1 year.

• [1]. Greenspan P, et al. Nile red: a selective fluorescent stain for intracellular lipid droplets. J Cell Biol. 1985 Mar;100(3):965-73.  [Content Brief]

  [2]. Gibrán S Alemán-Nava, et al. How to use Nile Red, a selective fluorescent stain for microalgal neutral lipids. J Microbiol Methods. 2016 Sep;128:74-79.  [Content Brief]

  [3]. Wilber Escorcia, et al. Quantification of Lipid Abundance and Evaluation of Lipid Distribution in Caenorhabditis elegans by Nile Red and Oil Red O Staining. J Vis Exp. 2018 Mar 5;(133):57352.  [Content Brief]

  [4]. Elizabeth C Pino, et al. Biochemical and high throughput microscopic assessment of fat mass in Caenorhabditis elegans. J Vis Exp. 2013 Mar 30;(73):50180.  [Content Brief]