nanoscale pharmaceutical formulations.

2. Scope

This SOP applies to the Analytical Method Development (AMD) department for characterizing nanoparticles, nanoemulsions, liposomes, polymeric micelles, and colloidal drug carriers in both R&D and QC environments.

3. Responsibilities

• Analytical Scientist: Designs DLS method, selects dispersants, and validates parameters.
• Instrument Analyst: Operates DLS equipment, conducts sample preparation and records results.
• QA Officer: Verifies compliance of DLS data and documentation.
• Head – AMD: Approves validated method and ensures readiness for regulatory inspection.

4. Accountability

The Head of AMD is accountable for ensuring that the DLS methods are accurate, reproducible, and aligned with regulatory requirements for nanoscale characterization.

5. Procedure

5.1 Instrument Preparation and Calibration

1. Verify calibration using NIST-traceable latex standards of known hydrodynamic diameter (e.g., 100 nm, 500 nm).
2. Calibrate temperature control, detector alignment, and baseline noise.
3. Document details in Annexure-1: DLS Calibration Log.

5.2 Sample Preparation

1. Use freshly prepared, well-dispersed nanoparticle suspensions.
2. Filter through 0.22 µm syringe filter to remove dust (optional, not for intact vesicles).
3. Degas dispersant medium (e.g., water, PBS) prior to use.
4. Dilute sample to achieve count rate within instrument’s optimal range (100–300 kcps).

5.3 Instrument Settings

1. Temperature: 25°C (unless stability study requires otherwise)
2. Measurement Angle: 90° (or 173° for backscatter configurations)
3. Measurement Duration: 10–20 runs, 10 seconds each
4. Viscosity & Refractive Index: Input according to dispersant medium

5.4 Method Development and Analysis

1. Define hydrodynamic diameter range to be analyzed (typically 1–1000 nm).
2. Ensure signal intensity and correlation function pass system checks.
3. Evaluate results for:
   • Z-average diameter (nm)
   • Polydispersity Index (PDI): Acceptable if ≤ 0.2 for monodisperse systems
   • Size distribution: By intensity, number, and volume
4. Log all results in Annexure-2: Particle Size Report.

5.5 Method Validation

1. Validate DLS method per ICH Q2(R2) and EU nanomedicine guidelines:
   • Precision: RSD ≤ 2% for Z-average
   • Accuracy: Verify against certified standards
   • Specificity: Evaluate interference from excipients
   • Robustness: Confirm stability under varying dilutions and storage times
2. Document in Annexure-3: Validation Summary Sheet.

5.6 Precautions and Troubleshooting

1. Avoid bubbles in cuvettes; use low-volume cuvettes if sample is limited.
2. Ensure no vibration or ambient light interference during measurement.
3. Reject measurements with poor correlation or multimodal peak inconsistencies unless justified.

6. Abbreviations

• DLS: Dynamic Light Scattering
• PDI: Polydispersity Index
• kcps: kilo counts per second
• nm: nanometer
• RSD: Relative Standard Deviation
• SOP: Standard Operating Procedure

7. Documents

1. DLS Calibration Log – Annexure-1
2. Particle Size Report – Annexure-2
3. Validation Summary Sheet – Annexure-3

8. References

• ICH Q2(R2) – Validation of Analytical Procedures
• USP <729> – Globule Size Distribution in Lipid-Based Formulations
• FDA Guidance on Nanotechnology Applications in Drug Products
• EU Nanomedicine Regulatory Guidelines

9. SOP Version

Version: 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name
Designation
Department

11. Annexures

Annexure-1: DLS Calibration Log

Date	Standard Size (nm)	Observed Size (nm)	PDI	Analyst
18/05/2025	100.0	99.8	0.08	Rajesh Kumar

Annexure-2: Particle Size Report

Sample ID	Z-Average (nm)	PDI	Peak Mode (nm)	Analyst
NANO/RS/031	125.6	0.14	124.2	Sunita Reddy

Annexure-3: Validation Summary Sheet

Parameter	Result	Criteria	Status
Precision	RSD = 1.8%	≤ 2%	Pass
Accuracy	101.2%	95%–105%	Pass

Revision History:

Revision Date	Revision No.	Details	Reason	Approved By
04/05/2025	2.0	Updated instrument settings and annexures; aligned with nanomedicine standards	Annual SOP Review