Quantitative assessment of adipocyte differentiation in cell culture

作者：Nils Arne Kraus, Franziska Ehebauer, Benedikt Zapp, Bianca Rudolphi, Bettina Johanna Kraus, Daniel Kraus · 发表于：Adipocyte · 年份：2016 · DOI：10.1080/21623945.2016.1240137 · 被引用次数：225 · 研究领域：Adipose Tissue and Metabolism、Adipokines, Inflammation, and Metabolic Diseases、Viral Infectious Diseases and Gene Expression in Insects

Adipocyte cell culture is an important tool for mechanistic studies of energy metabolism. Many factors affect the differentiation of adipocytes in culture. Oil red O staining can be used to assess the degree of differentiation. However, the validity of this method for quantitative analysis has not yet been established. Here we show that a protocol with arbitrarily chosen parameters does not measure in the linear range and is not suitable for quantitative analysis (R2 = 0.077, p = 0.382), and develop and validate an optimized protocol for quantitative oil red O staining of cultured adipocytes. 3T3-L1 preadipocytes and adipocytes are fixed with 4% formaldehyde and stained with 0.2% oil red O solution in 40% 2-propanol for 30 minutes. Dye is eluted with 2-propanol, and absorption of the eluate is measured photometrically at 510 nm. This optimized protocol achieves excellent correlation between defined amounts of differentiated adipocytes on constant-size culture plates and photometric absorption (R2 = 0.972, p = 6.585E-14). The performance of the method is independent of the culture plates used. Thus, the optimized oil red O staining protocol can be universally employed to quantitatively assess adipocyte differentiation.