Run a UV sterilization cycle before starting the liposome preparation workflow. This step helps prepare the chamber environment before liquid handling and formulation processing.

Record the initial chamber environment before the formulation run. This information may help compare batches prepared under different room or chamber conditions.

Turn on chamber illumination to support camera-based observation of the hydration vessel and liposome dispersion during the process.

Place the pre-prepared lipid film, lipid concentrate, or lipid stock dispersion into the hydration vessel before starting the automated run. If a thin-film method is used, solvent evaporation and drying should be completed externally before placing the vessel in the NSL system.

Dispense the hydration medium into the vessel containing the lipid phase or lipid film. This step initiates lipid hydration and supports vesicle formation.

Add a safe model payload solution to demonstrate liposome-based loading or carrier development. For general demonstration, use a coloured or fluorescent model compound instead of a therapeutic drug.

Start controlled stirring to support contact between the lipid phase and hydration medium. This helps disperse the lipid material and begins vesicle formation.

Apply mild heating to assist lipid hydration and vesicle formation. The selected temperature should be suitable for the lipid composition and should not degrade the model payload.

Hold the lipid mixture under controlled conditions to allow hydration, swelling of lipid layers, and preliminary vesicle formation.

Continue stirring after hydration to improve dispersion uniformity. This step supports the formation of a more homogeneous liposome suspension before sonication.

Add a stabilizer or cryoprotectant solution if required by the formulation design. This may help improve dispersion stability or prepare the liposome sample for later storage studies.

Mix gently after stabilizer addition to avoid excessive foam or bubble formation while maintaining uniform distribution of the added component.

Apply sonication to reduce larger vesicles and improve dispersion uniformity. Sonication duration should be optimized carefully because excessive sonication may cause heat generation, lipid degradation, or payload instability.

Use sonicator bath heating only when temperature control during sonication is required. Temperature should be selected based on lipid composition and payload stability.

Allow the sonicated liposome dispersion to stabilize after processing. This step helps reduce foam, thermal stress, and immediate post-sonication instability.

Use white chamber illumination and camera documentation to record the final appearance of the liposome dispersion. Observe cloudiness, uniformity, visible aggregation, settling, foam, and colour distribution if a model payload was used.

Use exhaust control when required during formulation handling, especially if lipid stock preparation involved residual volatile components. Any solvent-handling step should be performed externally with proper safety arrangements.

Remove the prepared nano-liposome dispersion from the system and transfer it into a clean labelled vial. Record batch ID, lipid composition, payload type, hydration temperature, stirring duration, sonication duration, and visual observations.

If required, remove unencapsulated payload using external purification methods such as centrifugation, dialysis, size-exclusion separation, or filtration. This step is not an NSL module and should be documented as an external process.

Characterize the prepared liposome dispersion using appropriate external methods such as particle size analysis, zeta potential measurement, encapsulation efficiency, microscopy, release study, and stability testing. Methodology The nano-liposome dispersion was prepared using a Protoly-managed workflow supported by selected NSL modules. Before the automated run, the lipid phase was prepared separately as a lipid film, lipid concentrate, or lipid stock dispersion. If the thin-film hydration approach was used, solvent evaporation and drying were completed externally before placing the lipid-containing vessel inside the NSL system. The NSL chamber was first subjected to a timed UV sterilization cycle. The initial chamber condition was recorded using the environment sensor module, and white LED illumination was activated to support visual monitoring. Hydration buffer or deionized water was dispensed into the vessel containing the lipid phase using the reservoir dispensing module. A safe model payload solution was then added through a separate reservoir channel to demonstrate the concept of liposome-based payload loading. Controlled stirring was initiated to support contact between the lipid material and the aqueous phase. Mild heating was applied using the heater module to improve lipid hydration and support vesicle formation. The mixture was then held for a defined hydration period, allowing lipid layers to swell and form vesicular structures. After hydration, stirring was continued to improve dispersion uniformity. If required, a stabilizer or cryoprotectant solution was added through another reservoir channel, followed by gentle stirring. The dispersion was then processed using the sonicator module to reduce larger vesicles and improve the uniformity of the liposome suspension. Sonicator bath heating was used only when temperature support was required during the sonication stage. After sonication, the dispersion was held for a short stabilization period to allow foam reduction and thermal relaxation. The final nano-liposome dispersion was visually documented using LED illumination and the camera module. The sample was manually collected into a clean labelled vial for offline purification and characterization. This workflow is intended for research-scale and educational nano-liposome preparation only. It does not include validated drug encapsulation, sterility assurance, biological safety testing, clinical evaluation, or regulatory approval studies.