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View ProductsAre long timelines, inconsistent results, and low yields hindering your monoclonal antibody development? At ProteoGenix, we streamline your path to success with our cutting-edge Verified In-Situ Plate Seeding (VIPS™) technology, reducing development time by more than 50% and delivering clonality reports essential for successful IND submission. Our IP-free, royalty-free cell lines are engineered for stability and high productivity, offering yields exceeding 7g/L. With over 30 years of expertise and 100+ stable cell lines generated, we provide a platform focused on developability. Elevate your research and production with ProteoGenix’s proven solutions.
Why choose ProteoGenix’s stable cell line
generation services?
Advanced VIPS™ Technology
Accelerate custom cell line projects by over 50% with real-time, high-resolution imaging confirming monoclonality using state-of-the-art VIPS™ technology.
Yeilds Guarandeed
Plan for the long-term and secure your investments by choosing a stable cell line development service with yields exceeding 7g/L.
Diverse IP-Free Cell Lines
Benefit from our royalty-free cell lines including Proprietary CHO-1, CHO-S, CHO-DG44, HEK293 or even your preferred cell line… Get a service adapted to your requirements!
Emphasis on developability
Minimize production risks thanks to our extensive early testing of antibody candidates.
Ease of transference
Fast cell line and protocol transference to your CMO partners for cGMP bioproduction.
Protein production experts
500+ monoclonal antibodies developed, 100+ stable cell lines generated, 28+ years of experience in protein production. Choose a leading bioproduction company.
Our process of stable cell line generation
for monoclonal antibody production
Expression vector construction
- Gene synthesis optimized for the target expression system.
- Gene subcloning in a high-performing expression vector.
Developability Study
- Production testing in transient expression systems.
- Biochemical characterization properties and full reporting.
Host Cell Transfection and Characterization of Stable Transfected Pools
- Host cell stable transfection with recombinant plasmid.
- Generation and characterization of stable transfected pools.
- Selection and amplification of the most productive stable pools.
Best Monoclones Isolation and Selection by VIPS™ Technology
- Monoclone isolation using Verified In-Situ Plate Seeding (VIPS™).
- Screening and selection of highest producing clones via ELISA and other assays.
Characterization of the best monoclones
- Productivity evaluation in batch and/or fed-batch culture.
RCB QC and Stability Study
- QC analysis and custom cell line stability study.
- Transfer of RCB and optimized protocols.
Our recombinant antibody production
service content
| Step | Content | Timeline | Deliverables |
|---|---|---|---|
| Gene synthesis |
|
1 week |
|
| Transient Expression Evaluation |
|
2 weeks |
|
| Stable pool generation |
|
6 weeks |
|
| Single cell clone screening by VIPS™ |
|
~8-10 weeks |
|
| RCB Preparation and QC |
|
1 week |
|
| RCB Stability Study |
|
~ 4-5 weeks |
|
| Analytics |
|
~ 2-3 weeks |
|
Options:Further evaluation and characterization of the antibody can be done by
- Analysis of Antibody Aggregates (SEC-HPLC)
- Endotoxin detection test
- IgG Quantification (UV280)
- Activity analysis by ELISA against the antigen
- DSC Analysis (Thermostability Analysis by Differential Scanning Calorimetry)
- Affinity Determination (Kd) against soluble antigen via Biacore X100
- Affinity Determination (Kd) against antigen expressed on cell surface via Surface Plasmon
- Resonance imaging (SPRi)
- FACS analysis
- Biological potency (ADCC assay)
- Glycosylation profile (LC/MS)
VIPS™ Technical Specifications and Advantages
The use of Verified In-Situ Plate Seeding (VIPS™) technology in custom cell line development offers several significant technical advantages:
- High-Efficiency Cloning: VIPS™ technology enables high-efficiency single-cell seeding, crucial for generating monoclonal cell lines. This automated process significantly reduces the manual labor and potential for error associated with traditional methods like limiting dilution cloning, thus ensuring more reliable and faster cell line development.
- Assured Clonality: VIPS™ provides image-based evidence of clonality right from day zero. This “double-lock” approach ensures that each cell line starts from a single, visually confirmed cell, meeting stringent regulatory requirements for monoclonality. This is critical not only for scientific validation but also for regulatory approvals in biopharmaceutical production.
- Time and Cost Reduction: By automating the seeding and initial growth stages, VIPS™ cuts down the time required to establish stable cell lines. Traditional methods can be time-consuming and labor-intensive, but VIPS™ streamlines this process, leading to significant reductions in both time and operational costs associated with cell line development.
- Enhanced Outgrowth and Stability: The precise delivery and verification of single-cell seeding improve the outgrowth and stability of the clones. VIPS™ ensures that the cells grow from a verified single-cell origin, which enhances the uniformity and reproducibility of the cell lines developed. This uniformity is crucial when these cell lines are scaled up for therapeutic protein production, as it ensures consistent product quality.
- Regulatory Compliance: The detailed documentation and clonality reports generated by VIPS™ facilitate compliance with regulatory standards. These reports provide a reliable audit trail that can be critical during the review process by regulatory bodies, thus de-risking the submission and approval phases of biopharmaceutical development.
Custom Cell Line Development : Expression Systems and Formats
| Cell Lines | Amplification and Selection | Advantages | Applications |
|---|---|---|---|
| Proprietary CHO-K1 | Available with both MTX/DHFR-mediated or Methionine Sulfoximine (MSX)/GS-mediated amplification and selection. | Offers freedom to operate with a one-time fee payment, making it cost-effective for long-term use. | Suitable for the rapid and easy transfer for cGMP production, supporting scalable and flexible biomanufacturing processes. |
| CHO-STM | Utilizes MTX/DHFR-mediated or Methionine Sulfoximine (MSX)/GS-mediated amplification and selection. | No licence fee is required before moving towards commercial use, reducing initial investment costs. | Designed for straightforward scale-up and transfer, facilitating seamless progression from research to commercial production. |
| DG44 | Employs MTX/DHFR-mediated amplification and selection. | No license fees for early-stage use and is renowned for producing FDA-approved biotherapeutics. | Ideal for the bioproduction of therapeutic proteins under stringent regulatory requirements. |
| HEK293 Variants (HEK293, 293F, 293E) | These are human embryonic kidney cells that are versatile for transient and stable expression with high yield. | HEK293 cells are known for rapid growth and high protein yield, making them suitable for both research and commercial scale productions. | Commonly used in the production of viral vectors for gene therapy, vaccines, and recombinant protein products. |
| Customer-Specific Cell Lines | We offer the flexibility to develop and utilize custom cell lines as per client-specific requirements. | Tailored to meet unique project needs, ensuring optimal expression and productivity. | Custom cell lines are particularly useful for specialized therapeutic targets or when proprietary systems are needed for competitive differentiation. |
Case study: stable expression of an IgG1 monoclonal antibody
I. Gene Synthesis, Subcloning, And Early Testing
Antibody genes were optimized for expression in CHO cell lines, synthesized, and subcloned into our proprietary transient expression system.
CHO-K1 cells (30 ml) were transiently transfected, transient pools were grown in 30 ml, and antibody purification was performed using protein G resin.
| Yield: 18.2 mg/L | Quantity produced: 0.18 mg | Purity >90% |
|---|
| A final QC was performed by reduced and non-reduced PAGE analysis confirming the integrity of the antibody |
II. Development Of Stable-transfected Pools
- Construction of stable expression vectors containing:
- Heavy and light antibody chains
- CMV (human cytomegalovirus) promoters
- Polyadenylation signals for enhanced transgene expression
- Glutamine Synthetase (GS) – the selection marker
- Determination of natural resistance to Methionine Sulfoximine (MSX)
- Determination of natural resistance to Methionine Sulfoximine (MSX)
- Vector linearization
- Transfection of CHO-K1 suspension cultures
- Selection in the presence of MSX
3 positive stable pools were further characterized in fed-batch experiments and antibodies characterized by SEC-HPLC to confirm purity and detect potential aggregation
| Pool ID | Quantity (mg) | Yield (mg/L) | Purity |
|---|---|---|---|
| 1 | 37.3 | 1243 | >90% |
| 2 | 39.4 | 1313.3 | >90% |
| 3 | 43.6 | 1453.3 | >90% |
III. Production And Purification Scale-up
| 1L fed-batch production was carried out in 3L flasks and purification was performed using protein A resin |
|---|
| Final yield: 2015.5 mg/L | Final purity: >92% |
IV.isolation And Screening Of Stable Monoclones
Monoclones were isolated using the limiting dilution method in 96-well plates and
expression evaluation was performed by ELISA with anti-Fc antibodies
2 Rounds of limiting dilution
29 Best performing clones
Best monoclones were evaluated by SDS-PAGE and ELISA (Fc-antibody).
ELISA data (below) revealed 10 monoclones with high expression levels.
Bar length represents intensity obtained in ELISA, numbers represent the clone ID.
V. Evaluation Of Stable Monoclones
| Clone ID | 1 | 3 | 4 | 7 | 10 | 15 | 17 | 19 | 30 | 31 |
|---|---|---|---|---|---|---|---|---|---|---|
| Yield (g/L) | 2.05 | 2.86 | 1.07 | 1.68 | 4.34 | 2.57 | 3.21 | 2.60 | 1.45 | 1.25 |
Monoclones 3, 10, 17, and 19 maintained good stability and viability in
subsequent tests and were selected for further development
VI. Production Scale-up And Optimization
Monoclone 10 was used for initial scale-up with excellent results.
Why focus on monoclonal antibody developability?
95% of biologics under development fail to reach the market or clinic due to undetected developability issues. Early testing or screening has thus become crucial to overcome low success rates and minimize development risks. Producing monoclonal antibodies in vitro via transient systems is still the best way to produce small amounts of antibodies for measuring:
- Affinity/avidity
- Specificity/selectivity
- Aggregation profile
- Stability
- Ease of purification
- Ease of scalability
Detectable problems at this stage can be corrected by additional engineering of antibody leads. These engineering efforts, although laborious, can save considerable costs and time on the long run to license your therapeutic antibodies.
To learn more about stable cell line development, visit our frequently asked questions (FAQs) page. On this page, we cover all seminal principles and knowledge regarding monoclonal antibody production in stable cells and provide detailed insights into ProteoGenix’s unique platform for stable cell line development.