Silica gel tlc plates

A 13 mL aliquot of bacterial culture was pelleted, resuspended in DI water and extracted with 10 mL of chloroform/methanol mixture (2: 1, vol: vol) as described^{13 (link)}. A 5 mL volume from the (bottom) chloroform layer was transferred to a new glass vial, dried with an air stream. The dried lipid was resuspended in 100 μL of hexane.
Ten microliter (μL) of lipid samples (in hexane) and glyceryl trioleate (TL) standards (ranging from 1 to 100 μg) were applied to silica gel TLC plates (Product No. 4850–820, Whatman, Piscataway, NJ) and separated in a solvent system of hexane: diethyl ether: acetic acid (80: 20: 1, vol: vol)^{5 (link)}. After separation, the TLC plate was dried, rinsed with 1 M sodium chloride solution, stained in a 0.2% (wt/vol in 1 M sodium chloride) amido black solution followed by color development as described before^{13 (link)}.
The TAG content of each sample was determined by analyzing the image of the TLC plate using ImageJ software (National Institutes of Health, Bethesda, MD). A TAG standard curve was developed by correlating the loaded amount (μg) of TAG standards to “(area)/(mean gray value)” of the relevant spots developed on the TLC plate. The standard curve was then used to determine the TAG content of the each sample based on the corresponding spot on the TLC plate.

Lipids were extracted by bead beating cells in water:CHCl₃:methanol (0.3:1:1), collecting the single phase, and drying it under N₂ gas. Lipids were resuspended in chloroform, separated on silica gel TLC plates (Whatman or Analtech) using hexane:ethyl ether:acetic acid (80:20:1), and detected by charring with cuprous sulfate. Bands were identified by comparison to standards.