Standard Operating Procedure for Ultrafiltration/Diafiltration (UF/DF) Process in Biosimilar Manufacturing

Department	Biosimilars
SOP No.	SOP/BS/179/2025
Supersedes	SOP/BS/179/2022
Page No.	Page 1 of 10
Issue Date	04/05/2025
Effective Date	06/05/2025
Review Date	04/05/2026

1. Purpose

To establish a standardized procedure for performing Ultrafiltration/Diafiltration (UF/DF) of biosimilar drug substances for the purposes of concentration adjustment and buffer exchange using validated single-use or stainless steel TFF systems.

2. Scope

This SOP applies to all downstream processing operations where UF/DF is utilized for product concentration and formulation buffer exchange prior to final bulk storage of biosimilar proteins.

3. Responsibilities

• Production: Perform UF/DF operations, monitor transmembrane pressure (TMP), record processing parameters.
• Engineering: Maintain and calibrate UF/DF skids and pressure gauges.
• QA: Review batch records and verify integrity of UF membrane validation documentation.

4. Accountability

The Downstream Process Manager is accountable for ensuring proper execution, monitoring, and documentation of the UF/DF process in line with GMP standards.

5. Procedure

5.1 System Setup

1. Confirm that the UF/DF system has been cleaned, sanitized, and verified as per applicable cleaning SOP.
2. Install the ultrafiltration membrane (e.g., 10–30 kDa MWCO), ensuring correct flow orientation.
3. Perform system leak test with WFI at 1.5 bar for 10 minutes and document in Annexure-1.

5.2 Membrane Conditioning and Equilibration

1. Pre-rinse membrane with 2–3 volumes of WFI or equilibration buffer.
2. Equilibrate membrane with process buffer to match product matrix prior to sample loading.

5.3 Ultrafiltration Process (Concentration)

1. Load the clarified product into the feed tank.
2. Start recirculation with feed pressure ≤1.5 bar and TMP between 1.0–1.2 bar.
3. Monitor permeate flow and collect until target concentration is reached (e.g., 40–60 mg/mL).
4. Perform in-process protein measurement every 30–60 minutes and record in Annexure-2.

5.4 Diafiltration Process (Buffer Exchange)

1. Initiate diafiltration using the target buffer.
2. Perform 5–10 diavolumes of buffer exchange to ensure effective replacement.
3. Monitor conductivity or pH shift to confirm buffer transition.

5.5 Final Concentration Adjustment and Hold

1. Adjust final protein concentration and volume per batch specifications.
2. Collect retentate into sterile bulk hold bag or tank under aseptic conditions.

5.6 Post-Use Rinse and Integrity Check

1. Flush system with WFI to recover residual product (optional based on yield).
2. Perform post-use membrane integrity test if applicable (e.g., pressure hold).
3. Dispose of single-use components as per SOP for biowaste disposal.

6. Abbreviations

• UF: Ultrafiltration
• DF: Diafiltration
• TMP: Transmembrane Pressure
• MWCO: Molecular Weight Cut-Off

7. Documents

1. UF/DF System Setup Checklist – Annexure-1
2. Protein Concentration Log – Annexure-2

8. References

• ICH Q8 – Pharmaceutical Development
• FDA Guidance – Process Validation: General Principles and Practices
• WHO TRS 999 – Good Manufacturing Practice for Biotherapeutic Products

9. SOP Version

Version: 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name
Designation
Department

11. Annexures

Annexure-1: UF/DF System Setup Checklist

Activity	Status	Performed By	Date
Membrane Installed Correctly	Yes	Ajay Verma	04/05/2025
Leak Test Passed	Yes	Ajay Verma	04/05/2025
Rinse Completed	Yes	Ajay Verma	04/05/2025

Annexure-2: Protein Concentration Log

Time	Volume (L)	Protein Conc. (mg/mL)	TMP (bar)	Remarks
10:00	8.0	22.4	1.1	Stable
12:00	6.5	38.9	1.2	Approaching target

Revision History:

Revision Date	Revision No.	Details	Reason	Approved By
04/05/2025	2.0	Expanded diafiltration volume guidance and TMP monitoring	Process optimization