Tween 80

Tween 80 in Plant Tissue Culture

Safety Note: Always consult the SDS for Tween 80 and follow institutional safety procedures; treat unknowns conservatively. Tween 80 is generally considered non-toxic at low concentrations used in plant tissue culture, but skin and eye irritation are possible. Avoid ingestion.

Overview and Identity

Tween 80, also known as polysorbate 80, is a nonionic surfactant frequently employed in plant tissue culture media. Its inclusion enhances the efficacy of other components and improves culture performance.

Common Names, Synonyms, and Abbreviations

Tween 80

Polysorbate 80

Polyoxyethylene (20) sorbitan monooleate

Chemical Identity

Tween 80 is a complex mixture of polyoxyethylene sorbitan monooleate esters. The exact composition varies slightly between manufacturers. Tissue culture-grade Tween 80 is generally preferred, ensuring sterility and reduced levels of impurities that might interfere with plant growth. It’s typically supplied as a viscous, yellowish-brown liquid. No specific hydrate state is typically specified.

Functional Role(s) in Plant Tissue Culture

Primarily, Tween 80 functions as a surfactant in plant tissue culture. It reduces surface tension, aiding in the even distribution of other media components, promoting better wetting of explants, and improving the uptake of growth regulators and other nutrients. It is not a macronutrient, micronutrient, vitamin, PGR, buffer, chelator, gelling agent, sterilant, or mutagen.

Mechanism and Rationale in Vitro

Tween 80’s surfactant properties are key to its function. By lowering the surface tension of the medium, it facilitates better contact between the explant and the culture medium, supporting nutrient and gas exchange. This can lead to improved hydration, nutrient absorption, and overall explant health.

Stage-Specific Relevance

Tween 80 can be beneficial throughout various stages of plant tissue culture. It’s often used in callus induction, shoot proliferation, rooting, and somatic embryogenesis protocols. Its use with protoplasts is less common but could potentially improve cell wall formation. It is not directly involved in contamination control.

Interactions or Compatibility/Antagonism with Other Agents

Tween 80 generally exhibits good compatibility with most common plant tissue culture media components. However, interactions with certain gelling agents (e.g., reduced gelling strength at high concentrations) or potential to interfere with certain assays should be considered. It does not directly interact with auxin-cytokinin balance or chelators like EDTA. Photodegradation is not a significant concern at typical concentrations when exposure to light is minimized.

Preparation and Stock Solutions

Tween 80 is readily soluble in water, ethanol, and DMSO. Stock solutions are typically prepared by weighing the desired amount of Tween 80 and dissolving it in the chosen sterile solvent. pH adjustment is usually unnecessary, as Tween 80 itself has a neutral pH. Because it is heat-labile, sterilization is achieved by filtration through a 0.22µm filter after dissolving.

Example Stock Recipes:

10% (v/v) Tween 80 stock solution in water: Weigh 10g Tween 80 and add sterile distilled water to a final volume of 100mL in a sterile, amber glass bottle. Filter sterilize through 0.22µm filter.

5% (v/v) Tween 80 stock solution in ethanol: Weight 5g Tween 80 and add sterile absolute ethanol to a final volume of 100mL in a sterile, amber glass bottle. Filter sterilize through a 0.22µm filter.

Working Concentrations and Usage in Media

Working concentrations typically range from 0.01% to 0.1% (v/v), although precise concentrations will be highly dependent on the species and explant type. Some protocols add Tween 80 together with plant growth regulators (PGRs) to potentially improve absorption efficiency, but this is not generally accepted as a standard practice. Addition is usually done after autoclaving the basic medium while it is still warm (45-50°C) to ensure proper mixing.

Storage and Stability

Stock solutions should be stored in tightly closed, amber glass bottles at 4°C, protecting from direct light and avoiding excessive temperature fluctuations. Shelf life is typically 6-12 months, but it should be regularly checked for any signs of degradation (color change, precipitation).

Quality, Sourcing, and Compatibility

Use tissue-culture-grade Tween 80 to minimize the risk of contaminants that could affect plant growth. Lot-to-lot variability can occur; therefore, it is always recommended to run bioassays, checking for physical changes before use in a large batch of media.

Safety and Precautions

Tween 80 is generally considered non-toxic at standard tissue culture concentrations. However, skin and eye irritation can occur. Wear appropriate PPE (gloves, goggles, lab coat). Spills should be cleaned immediately with absorbent materials, following institutional protocols. Waste should be disposed of according to local regulations.

Troubleshooting and Optimization

Problems like reduced gelling strength, precipitation, or inconsistent regeneration can sometimes be linked to Tween 80. If issues arise, optimize empirically by adjusting concentration or switching to an alternative surfactant.

Example Protocols and Parameters

Callus Induction (Tobacco): Murashige and Skoog (MS) basal medium + 2,4-D (1 mg/L) + kinetin (0.5 mg/L) + 0.05% (v/v) Tween 80; pH 5.7; gellan gum: 2 g/L.

Shoot Proliferation (Carnation): MS basal medium + BA (1 mg/L) + GA3 (0.1 mg/L) + 0.02% (v/v) Tween 80; pH 5.8; agar: 6 g/L

Documentation and Labeling

Always clearly label stock solutions and working media with: chemical name (Tween 80), lot number, preparation date, stock concentration, solvent, pH, storage conditions, and expiry date. Cross-reference with media batch and treatment information.

Key Takeaways