Standard Operating Procedure for Seed Train Validation in Biosimilar Manufacturing

Department	Biosimilars
SOP No.	SOP/BS/107/2025
Supersedes	SOP/BS/107/2022
Page No.	Page 1 of 13
Issue Date	04/05/2025
Effective Date	06/05/2025
Review Date	04/05/2026

1. Purpose

To define the procedure for validation of the seed train process used in upstream biosimilar production, ensuring consistency of cell growth and quality attributes from thaw to production-scale inoculum.

2. Scope

This SOP applies to all stages of the seed train from vial thawing, shake flask, spinner flask, and seed bioreactor expansions up to the point of inoculation into the main production bioreactor for biosimilar manufacturing.

3. Responsibilities

• Process Development: Define seed train parameters and validation protocol.
• Manufacturing: Execute seed train stages and record all critical data.
• QA: Review validation data, approve protocol and final report.

4. Accountability

The Head of Manufacturing Sciences is accountable for ensuring that seed train validation is performed, approved, and maintained in compliance with regulatory expectations.

5. Procedure

5.1 Preparation and Planning

1. Prepare Seed Train Validation Protocol including:
   • Acceptance criteria for growth, viability, and cell density
   • Scalability criteria (VCD growth factor, doubling time)
   • Sampling points and analytical methods
2. Obtain QA approval before execution.

5.2 Seed Train Execution

1. Start with a thawed vial from a qualified Master Cell Bank.
2. Expand through sequential vessels:
   • Shake flask (50–250 mL)
   • Spinner flask or Wave bag (500 mL–2 L)
   • Seed bioreactor (5 L–200 L)
3. Monitor and record:
   • Viable Cell Density (VCD)
   • Cell viability
   • pH, temperature, DO
   • Contamination (visual and microbial)

5.3 Sampling and Testing

1. Collect samples at each expansion stage:
   • Start and end of each vessel
   • Additional time points as required
2. Analyze for:
   • Growth rate and doubling time
   • Glucose consumption, lactate production
   • Viability ≥ 90% at end of each stage

5.4 Data Analysis and Documentation

1. Document all results in Annexure-1 and Annexure-2.
2. Evaluate data against pre-defined criteria.
3. Summarize in Seed Train Validation Report and submit to QA.

5.5 Revalidation Criteria

1. Revalidate seed train upon:
   • Change in media or supplements
   • Change in vessel configuration or scale
   • New cell bank introduction

6. Abbreviations

• VCD: Viable Cell Density
• QA: Quality Assurance
• MCC: Master Cell Bank
• DSP: Downstream Processing

7. Documents

1. Seed Train Execution Log – Annexure-1
2. Seed Train Validation Checklist – Annexure-2

8. References

• ICH Q8 – Pharmaceutical Development
• ICH Q9 – Quality Risk Management

9. SOP Version

Version: 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name
Designation
Department

11. Annexures

Annexure-1: Seed Train Execution Log

Date	Stage	VCD (10⁶ cells/mL)	Viability (%)	Glucose (g/L)	Remarks
04/05/2025	Shake Flask	1.2	94	2.5	Normal

Annexure-2: Seed Train Validation Checklist

Parameter	Target	Observed	Status	Reviewed By
Doubling Time	18–24 hrs	21 hrs	Pass	Ajay Verma

Revision History:

Revision Date	Revision No.	Revision Details	Reason for Revision	Approved By
04/05/2025	2.0	Updated validation metrics and added revalidation triggers	Process harmonization