Hi Sofea Sazali ,

There is no single recommended final % for Tween 40 or Tween 80 in cells as the final concentration that could be tolerated by cells differ for each cell lines. However, i would not suggest concentration of >0.05% for these type of solvent.

You can refer to this articles: https://doi.org/10.1079/bjn20041094 / https://www.mdpi.com/1422-0067/22/23/13110.

Do a pilot test to check what's the max % your cells could tolerate at your experimental condition. Also, ensure you have a proper vehicles control when you run your actual mtt later on.

Hello Sofea Sazali

The nature of the oil sample itself influences the required surfactant amount for effective emulsification and solubilization. For cell-compatible use, the optimal non-ionic surfactant concentration of Tween 40 or Tween 80 for dissolving an oil sample is typically 0.01% to 0.1% (w/v or v/v), as this range provides sufficient solubilization without causing cell toxicity.

You should begin with a low concentration (e.g., 0.05%) and gradually increase until you achieve sufficient solubilization while maintaining cell viability. Add Tween 40 or Tween 80 to a small amount of your oil sample to see if it forms a clear solution or stable emulsion. Once you have a working stock solution, add it to your complete cell culture medium at a very low final concentration. Incubate your cells with the prepared medium containing the oil and Tween 40 or Tween 80. Observe for any signs of cell death or reduced viability over a defined period (e.g., 24 hours or more). If cell viability is high and the oil is not effectively solubilized, gradually increase the Tween 40 or Tween 80 concentration and repeat the cell culture assay until you find a concentration that effectively solubilizes the oil without harming the cells.

If your oil sample requires very high concentrations, consider alternative emulsifiers or co-surfactants like ethanol to achieve the desired solubilization at lower, more cell-friendly Tween levels. If you must make a choice between Tween 40 and Tween 80, Tween 80 is a better general choice for a broad range of oil samples, especially those rich in unsaturated oils, as it has a higher affinity for lipophilic compounds. Tween 40 is also a good emulsifier but may be more specific to oils rich in saturated fats. If your primary goal is to maximize the solubilization of a broad mix of compounds from your oil sample, you should choose Tween 80.

Good Luck!

Hi!

You must use polysorbates, keep Tween-40/80 as low as feasible (typically ≤0.01–0.05% v/v), because nonionic surfactants can perturb membranes and show cell-type-dependent cytotoxicity; always titrate a matched vehicle control. As co-solvent back-ups, ethanol ≤0.5–1% v/v is usually tolerated by RAW264.7, whereas higher levels blunt responses; avoid DMSO here not only for insolubility but also because it modulates macrophage signaling. Finally, verify that the vehicle and colored/oily analyte do not chemically interfere with MTT readout by running “compound-only” blanks and an orthogonal viability assay.

For your reference:

Article Considerations regarding use of solvents in in vitro cell based assays