Standard Operating Procedure for Sterile Addition of Feeds and Supplements in Bioreactor Operations for Biosimilars

Department	Biosimilars
SOP No.	SOP/BS/097/2025
Supersedes	SOP/BS/097/2022
Page No.	Page 1 of 12
Issue Date	04/05/2025
Effective Date	06/05/2025
Review Date	04/05/2026

1. Purpose

To establish a validated, sterile method for the addition of feeds, buffers, and supplements to bioreactor systems during biosimilar production. This ensures aseptic conditions and prevents contamination that could compromise batch quality.

2. Scope

This SOP applies to all operations involving the transfer of nutrient feeds, glucose solutions, pH adjustment buffers, and other additives into both single-use and stainless-steel bioreactors in the biosimilars manufacturing unit.

3. Responsibilities

• Manufacturing Operator: Prepare, filter, and transfer solutions using sterile connections.
• QA Officer: Review batch records and ensure sterile practices are adhered to.
• Engineering: Ensure sterilization validation of equipment and connection lines.

4. Accountability

The Head of Manufacturing is accountable for ensuring sterile handling, equipment integrity, and operator compliance during feed addition processes.

5. Procedure

5.1 Preparation of Feed Solutions

1. Weigh and dissolve raw materials (glucose, amino acids, salts) in WFI using a validated mixing tank.
2. Filter final solution using a 0.22 μm sterile-grade filter into a pre-sterilized feed bag or bottle.
3. Label feed container with solution name, batch number, preparation date, and expiry (max 24 hours).

5.2 Bioreactor Port Preparation

1. Ensure that the bioreactor port for feed addition (e.g., Luer-lock, CPC) is sterile and capped.
2. Sanitize port exterior with 70% IPA prior to connection.

5.3 Sterile Connection Process

1. Use one of the following validated sterile connection methods:
   • Pre-sterilized tube welding (for single-use systems)
   • Steam-in-place (SIP) transfer lines for SS bioreactors
   • Aseptic quick-connects (e.g., Kleenpak, AseptiQuik)
2. Perform connections in a Grade A/B laminar flow or controlled area.

5.4 Feed Addition Execution

1. Open transfer line valves slowly to minimize pressure fluctuations.
2. Record the exact time, volume, and feed solution details in the batch record (Annexure-1).
3. Monitor bioreactor parameters (pH, DO, temperature) during and after addition.

5.5 Post-Addition Actions

1. Disconnect the transfer line using aseptic techniques and reseal the port if required.
2. Dispose of used tubing and feed containers as per biohazard protocols.
3. Retain a sample of the feed solution for QC analysis (sterility, osmolality, composition).

6. Abbreviations

• WFI: Water for Injection
• IPA: Isopropyl Alcohol
• SIP: Sterilize-In-Place
• QC: Quality Control

7. Documents

1. Feed Addition Log – Annexure-1
2. Sterile Connection Checklist – Annexure-2

8. References

• ICH Q7 – GMP for Active Pharmaceutical Ingredients
• WHO TRS 1010 Annex 4 – Good Manufacturing Practices for Biological 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: Feed Addition Log

Date	Batch No.	Feed Name	Volume (mL)	Added By	Time
04/05/2025	BS-215	Glucose 40%	500	Sunita Reddy	10:30 AM

Annexure-2: Sterile Connection Checklist

Connection Method	Probe Sterilized	Environment Grade	Integrity Test	Performed By	Date
AseptiQuik	Yes	Grade B	Pass	Rajesh Kumar	04/05/2025

Revision History:

Revision Date	Revision No.	Revision Details	Reason for Revision	Approved By
04/05/2025	2.0	Included sterile tubing methods and new annexures	Compliance enhancement