Standard Operating Procedure for Creation of Working Cell Bank (WCB) for Biosimilars

Department	Biosimilars
SOP No.	SOP/BS/025/2025
Supersedes	SOP/BS/025/2022
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 the generation, labeling, freezing, testing, and storage of the Working Cell Bank (WCB) derived from a qualified Master Cell Bank (MCB) for biosimilar production use.

2. Scope

This SOP applies to Cell Line Development, Quality Control, and Quality Assurance teams involved in the preparation and release of the WCB for GMP manufacturing operations.

3. Responsibilities

• CLD Scientist: Expands MCB vial, prepares WCB batches under aseptic conditions, and oversees cryopreservation.
• QC Analyst: Performs viability, sterility, mycoplasma, and identity testing on WCB vials.
• QA Officer: Reviews batch records and approves WCB release for manufacturing use.

4. Accountability

The Head of CLD is accountable for ensuring the WCB is traceable, well-characterized, and GMP-compliant, with all activities documented and archived.

5. Procedure

5.1 MCB Revival and Expansion

1. Thaw a vial from the released MCB under aseptic conditions in a biosafety cabinet.
2. Expand cells in T-flasks and spinner flasks over 2–3 passages using production media.
3. Monitor cell density, morphology, and viability at each passage.

5.2 Harvest and Viability Check

1. Harvest cells when viable cell density reaches 2–5 × 10⁶ cells/mL with ≥95% viability.
2. Centrifuge at 300 × g for 5 minutes and resuspend in pre-chilled cryopreservation media (90% production medium + 10% DMSO).

5.3 Aliquoting and Labeling of WCB Vials

1. Dispense 1.0 mL per vial (target ≥ 100 vials) into sterile, labeled cryovials.
2. Label to include:
   • Cell Line ID
   • WCB Lot No.
   • Date of Freezing
   • Passage No.
   • Technician Initials

5.4 Controlled Rate Freezing

1. Freeze vials in a controlled-rate freezer at -1°C/min to -80°C.
2. Transfer to liquid nitrogen storage after 12–18 hours.

5.5 QC Testing of WCB

1. Send representative vials for:
   • Cell viability (Trypan Blue)
   • Sterility (USP <71>)
   • Mycoplasma testing (PCR and culture)
   • Identity confirmation (STR profiling)
2. Record results in WCB QC Log (Annexure-1).

5.6 Documentation and Release

1. Complete batch records for freezing, QC results, and inventory.
2. Prepare Certificate of Analysis and submit for QA review.
3. Release WCB after QA approval with full documentation.

6. Abbreviations

• WCB: Working Cell Bank
• MCB: Master Cell Bank
• DMSO: Dimethyl Sulfoxide
• STR: Short Tandem Repeat

7. Documents

1. WCB QC Log (Annexure-1)
2. WCB Freezing and Labeling Record (Annexure-2)

8. References

• ICH Q5D – Cell Substrate Derivation and Characterization
• WHO TRS 978 – GMP for Cell Banks
• CDSCO Biosimilar Quality Guidelines (India)

9. SOP Version

Version: 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name
Designation
Department

11. Annexures

Annexure-1: WCB QC Log

Test	Method	Result	Acceptance Criteria	Status
Viability	Trypan Blue	96%	≥ 85%	Pass
Sterility	USP <71>	Negative	No Growth	Pass
Mycoplasma	PCR	Negative	Absent	Pass
STR Profile	Capillary Electrophoresis	Matched	100% Match	Pass

Annexure-2: WCB Freezing and Labeling Record

Vial No.	WCB Lot No.	Cell Line ID	Freezing Date	Location	Operator
WCB001	WCB-BS-25A	CL-BS-025	03/05/2025	T2/R3/B5	Rajesh Kumar

Revision History:

Revision Date	Revision No.	Revision Details	Reason for Revision	Approved By
04/05/2025	2.0	Updated QC panel and added C of A preparation	Audit Recommendation