Antibody generation can be approached through two main methods: in vivo (within a living organism) andin vitro (in a laboratory setting). Below is a comparison of their advantages, drawbacks, and technical
differences.
Advantages
| Category | In Vivo | In Vitro |
| Antigen Format Compatibility | Broad, including native antigens and cDNA-encoded targets | Limited to purified proteins, peptides, or high-expressing cell lines |
| Specificity and Affinity | High specificity and affinity due to natural selection and affinity maturation | May require additional affinity maturation and optimization |
| Post-Translational Modifications | Incorporates mammalian modifications (e.g., glycosylation) | Lacks post-translational modifications, which can impact final antibody function |
| Humanization | Transgenic mice produce humanized antibodies, reducing immunogenicity risks | Direct screening of human libraries possible, minimizing immunogenicity concerns |
| Output Volume | High; large numbers of high-quality antibodies can be obtained from a single animal | Potentially higher output if large phage libraries are well-developed |
| Market Availability | 89% of approved antibodies are from in vivo methods | 11% of approved antibodies come from in vitro methods |
| Timeframe | Fast (2–3 months), though some protocols may take up to 8 months | Faster if libraries are already developed, but establishing libraries can take 6–7 months |
| Ease of Use | Technically easy to generate with established protocols | Technically challenging, requires advanced automation and expertise |
Drawbacks
| Category | In Vivo | In Vitro |
| Development Time | Can take up to 8 months (with rapid immunization, 1 month possible) | 6–7 months for initial library development |
| Antigen Limitations | Non-immunogenic or toxic antigens present challenges | Can handle non-immunogenic and toxic antigens effectively |
| Optimization Needs | May require humanization for therapeutic use | Requires optimization for affinity, specificity, and manufacturability |
| Post-Translational Modifications | Naturally incorporates modifications like glycosylation | Lacks mammalian modifications, leading to potential functional issues |
| Manufacturing Compatibility | Typically well-suited for large-scale manufacturing | Phage display antibodies may need further refinement for manufacturability |
| Ethical Concerns | Requires animal use and adherence to animal welfare regulations | No animal use in synthetic libraries (unless using immune/naive libraries) |
| Cost and Accessibility | Relatively cost-effective for both academia and industry | Higher costs, especially for small academic labs and start-ups |
Key Insight: 🟢 Market Dominance
Technical Complexity and Use Cases
| Category | In Vivo | In Vitro |
| Technical Complexity | Lower technical complexity, making it accessible to more labs | High technical complexity, requiring advanced phage libraries and automation |
| Antigen Screening | Broad antigen screening but limited with toxic/non-immunogenic antigens | Effective for screening non-immunogenic and toxic antigens |
| Automation Requirements | Less reliant on automation for antibody discovery | Requires extensive automation to screen the full diversity of large libraries |
| Target Discovery | Suitable for cDNA-encoded targets and complex antigens | Limited by antigen formats during panning, restricting discovery of certain targets |
| Time to Market | Faster process in most cases due to established protocols | Can be slower if libraries need to be built or optimized |
Key Insight: 🔍 Balancing Speed and Complexity
Timeframe and Post-Production Modifications
| Category | In Vivo | In Vitro |
| Timeframe | 2–3 months for rapid protocols, up to 8 months for complex cases | 6–7 months for library development, faster if libraries already exist |
| Affinity Maturation | Natural affinity maturation occurs during the immune response | May require laboratory affinity maturation and optimization |
Post-Translational Modifications
| Fully integrated post-translational modifications such as glycosylation | No post-translational modifications in phage display or panning processes
|
Cost and Accessibility Comparison
| Category | In Vivo | In Vitro |
| Cost | Generally affordable and widely accessible | Higher costs due to complex setups and automation |
| Accessibility for Small Labs | Well-established in academic research; low barriers to entry | Difficult and expensive to implement for small labs, start-ups, and academia |
| Scaling and Manufacturing | Easily scalable for large production and well-suited for clinical applications | Requires additional optimization for scaling and manufacturing |
Key Insight: 💡 In Vitro’s Potential Despite Complexity
While in vitro approaches can be more challenging and expensive to implement, they allow for the discovery
of antibodies against non-immunogenic or toxic antigens, a critical advantage in specific research areas.
Synergistic Potential: Combining In Vivo and In Vitro Approaches
| Process | In Vivo | In Vitro |
| Initial Antigen Exposure | Begin with immunization of animals | Directly screen from human libraries |
| Affinity Maturation | Utilize the natural affinity maturation process | Laboratory-based affinity maturation and optimization needed |
| Final Screening and Refinement | High-quality antibodies from animals refined with in vitro technologies | Further refinement can happen with post-production in vivo testing |
Biointron’s catalog products for in vivo research can be found at Abinvivo,
where we have a wide range of Benchmark Positive Antibodies, Isotype Negative Antibodies, Anti-Mouse Antibodies,
Bispecific Antibodies, and Antibody-Drug Conjugates. Contact us to find out more at info@biointron.com or +86 400-828-8830 / +1(732)790-8340.
