SOP-032: Establishment and Management of Master and Working Cell Banks
Purpose
To establish standardized procedures for creating, maintaining, and using Master Cell Banks (MCBs) and Working Cell Banks (WCBs) to preserve cell line integrity and ensure experimental reproducibility.
Scope
This SOP applies to all mammalian cell lines maintained for long-term laboratory use.
Responsibilities
The operator is responsible for:
- Establishing Master and Working Cell Banks.
- Ensuring cultures meet laboratory quality requirements before banking.
- Maintaining accurate inventory records.
- Preventing unnecessary passage of Master Cell Bank stocks.
Definitions
Master Cell Bank (MCB)
A collection of cryopreserved vials prepared from a single authenticated, contamination-free cell culture that serves as the primary long-term reference stock.
Working Cell Bank (WCB)
A collection of cryopreserved vials prepared from the Master Cell Bank and used for routine laboratory work.
Required Materials
Reagents
- Cryopreservation medium
- Complete culture medium
Equipment
- Cryovials
- Controlled-rate freezing container (if used)
- Liquid nitrogen storage system
- Class II Biosafety Cabinet
- Laboratory inventory records
Procedure
1. Verify Culture Quality
Before banking, confirm that the culture:
- Exhibits normal morphology.
- Is free from contamination.
- Meets laboratory quality requirements.
2. Establish the Master Cell Bank
Prepare multiple cryovials from a healthy, low-passage culture.
Assign each vial a unique identifier.
Store all MCB vials in designated long-term cryostorage.
3. Establish the Working Cell Bank
Recover a vial from the MCB.
Expand the culture under routine conditions.
Prepare multiple cryovials to create the WCB.
Store WCB vials separately from the MCB.
4. Routine Laboratory Use
Use only WCB vials for routine experiments whenever possible.
Reserve MCB vials for establishing new WCBs or replacing exhausted stocks.
5. Inventory Management
Maintain an inventory including:
- Cell line name
- Bank type (MCB or WCB)
- Cryovial identification
- Storage location
- Date of preparation
- Passage number
Acceptance Criteria
| Parameter | Acceptance Criterion |
|---|---|
| Cell morphology | Typical for the cell line |
| Contamination | None detected |
| Cell bank labeling | Complete and accurate |
| Inventory | Current and traceable |
Critical Control Points
- Establish the MCB from low-passage cultures.
- Authenticate the cell line before banking.
- Confirm negative mycoplasma status before cryopreservation.
- Never use MCB stocks for routine experiments.
- Maintain separate storage records for MCB and WCB.
Documentation
Record the following:
- Cell line
- Bank type (MCB/WCB)
- Passage number
- Number of cryovials
- Storage location
- Date of banking
- Operator initials
Quality Control
Verify:
- Cell identity has been authenticated.
- Mycoplasma testing has been completed.
- Cryovials are correctly labeled.
- Inventory records are current.
- Storage locations are accurate.
Troubleshooting
| Observation | Possible Cause | Corrective Action |
|---|---|---|
| Low recovery after thawing | Poor-quality source culture | Establish a new bank from a healthy culture |
| Mislabeled cryovials | Documentation error | Verify inventory and relabel according to laboratory policy |
| MCB depleted | Excessive routine use | Establish a new MCB from an authenticated early-passage culture |
| Inventory discrepancies | Incomplete record keeping | Audit storage records and update inventory |
Safety Considerations
- Handle cryogenic materials using appropriate PPE.
- Maintain aseptic technique during all banking procedures.
- Treat all cultures as potentially biohazardous.
- Dispose of defective cryovials according to laboratory biosafety procedures.