AtaGenix Laboratories
AtaGenix Laboratories
AtaGenix provides in vitro affinity maturation services to enhance antibody binding affinity by 10–100-fold through CDR-targeted mutagenesis and high-throughput screening. Whether your starting antibody comes from phage display, hybridoma, or single B cell discovery, affinity maturation transforms a moderate-affinity lead into a high-potency candidate ready for therapeutic, diagnostic, or research applications.
10–100x
KD Improvement
6–8 wks
Typical Turnaround
CDR 1–3
Full Saturation Mutagenesis
SPR/BLI
Kinetic Validation
Affinity maturation is particularly valuable when: naïve library hits have moderate affinity that needs improvement for therapeutic use; diagnostic antibodies require higher sensitivity; you need to rescue weak or borderline clones; or you want to engineer cross-reactivity to orthologs or mutated targets. We support all antibody formats including IgG, scFv, Fab, VHH, bispecifics, and Fc-fusions.
| Scenario | Starting Point | Goal |
| Naïve library hit → therapeutic | Naïve phage display hit with nM–low µM KD | Sub-nM affinity for therapeutic potency |
| Diagnostic sensitivity boost | Existing antibody with adequate specificity but low signal | Higher affinity = lower LOD in ELISA/IHC/FC |
| Weak clone rescue | Borderline hybridoma or scFv/VHH clone | Restore or improve affinity for functional use |
| Cross-reactivity engineering | Antibody that binds human target but not mouse/cyno ortholog | Engineered cross-species binding for preclinical studies |
| CAR-T / ADC / bispecific optimization | scFv or VHH with suboptimal affinity for payload delivery | Tuned affinity for optimal therapeutic index |
Full CDR Saturation Mutagenesis
We mutagenize CDR1, CDR2, and CDR3 regions individually or in combination using site-directed mutagenesis, error-prone PCR, or NNK codon scanning. This creates focused libraries that explore the full affinity landscape around your starting sequence.
Phage + Mammalian Display
Matured libraries are screened by phage display for high throughput, with top hits validated in mammalian display for expression and developability context. This dual-system approach avoids selecting variants that display well in E. coli but express poorly in CHO/HEK.
SPR/BLI Kinetic Validation
Every matured variant is characterized by SPR (Biacore) or BLI for kon, koff, and KD. We quantify the actual improvement in kinetics, not just ELISA signal. Hotspot mutations are then combined and re-screened to achieve maximum affinity gain.
AtaGenix's affinity maturation service takes your existing antibody sequence through CDR-targeted mutagenesis (error-prone PCR, site-directed mutagenesis, or NNK saturation), library construction (scFv/Fab/VHH format), phage or mammalian display screening, SPR/BLI kinetic validation, hotspot combination, and final expression in HEK293 or CHO. Typical turnaround: 6–8 weeks. Supported formats: IgG, scFv, Fab, VHH, bispecifics, Fc-fusions. Deliverables include affinity-matured sequences, SPR/BLI kinetic data, expression reports, and a technical report with improvement quantification.
A 5-step pipeline from your starting antibody sequence to affinity-improved, kinetically validated candidates. Typical timeline: 6–8 weeks.
01
Sequence Engineering
CDR1/2/3 mutagenesis
Error-prone PCR or NNK scanning
scFv / Fab / VHH library construction
02
Library Screening
Phage display panning
ELISA / FACS selection
Competitive elution for high-affinity enrichment
03
Hit Validation
SPR / BLI kinetic measurement
kon, koff, KD determination
Hotspot mutation identification
04
Combination & Re-screen
Beneficial mutations combined
Combinatorial library synthesis
Second-round screening for max gain
05
Expression & Delivery
HEK293 / CHO expression
Purification & QC
Matured sequences + kinetic data + report
Service Scope
| ✓ CDR saturation mutagenesis (CDR1, CDR2, CDR3) | ✓ Error-prone PCR & chain shuffling |
| ✓ Phage display + mammalian display screening | ✓ ELISA, FACS, competitive selection |
| ✓ SPR (Biacore) / BLI kinetic validation | ✓ Hotspot combination & re-screening |
| ✓ HEK293 / CHO recombinant expression | ✓ Formats: IgG, scFv, Fab, VHH, BsAb, Fc-fusion |
| ✓ Deliverables: matured sequences + kinetic data + report | ✓ Optional: thermostability (DSC), solubility profiling |
| Method | How It Works | Best For |
| CDR Saturation (NNK) | Systematically replaces each CDR residue with all 20 amino acids using degenerate codons | Comprehensive exploration when starting point is known |
| Error-Prone PCR | Random mutations introduced across the entire variable region at controlled frequency | Broad exploration when optimal positions are unknown |
| Chain Shuffling | One chain (VH or VL) is fixed while the other is replaced from a diverse repertoire | When one chain contributes disproportionately to binding; cross-reactivity engineering |
| Hotspot Combination | Beneficial single mutations identified in rounds 1–2 are combined into multi-mutation variants | Maximizing additive/synergistic affinity gains from individual hotspots |
Need an antibody to mature? Start discovery with Phage Display (naïve library hits are the most common input for maturation), Rabbit Single B Cell, or Rapid Hybridoma, then feed leads directly into our maturation pipeline. For kinetic measurement of your current antibody, see our SPR/Biacore Analysis service.
How much can affinity be improved?
Typical improvements are 10–100-fold in KD. A naïve library hit at 50 nM can often be matured to 0.5–5 nM. The achievable improvement depends on the starting affinity, the epitope, and the structural plasticity of the CDRs. We quantify improvement by SPR/BLI kinetic measurement before and after maturation, reporting both KD and the individual kon/koff changes.
What do I need to provide?
Your antibody sequence (VH and VL, or VHH for nanobodies) and the target antigen protein. If you have existing SPR/ELISA affinity data for the starting antibody, that helps us design the mutagenesis strategy. If you do not have the antigen, AtaGenix can express it using our protein expression platforms.
Which mutagenesis method should I choose?
If your antibody has a known crystal structure or docking model, CDR saturation mutagenesis at predicted contact residues is most efficient. If the binding interface is unknown, error-prone PCR across the full variable region explores all positions. Chain shuffling is best when you suspect one chain (VH or VL) limits affinity. In practice, we often combine methods — error-prone PCR in round 1 to identify hotspots, then CDR saturation in round 2 to optimize them, then hotspot combination in round 3.
Can affinity maturation affect specificity or developability?
Yes — this is why we validate matured variants not just for improved KD but also for retained specificity (cross-reactivity panel), expression yield, and thermal stability. Mutations that improve affinity but introduce aggregation, polyreactivity, or poor expression are flagged and excluded. Optional DSC thermostability and solubility profiling are available to assess developability of final candidates.
Affinity improvement is target- and sequence-dependent. Not all antibodies are equally amenable to maturation; CDR structure and germline constraints influence the achievable gain. SPR/BLI kinetic data for before-and-after comparison is included in all projects. Quote-based pricing.
Contact Us
+86-27-6552-3339
info@atagenix.com
Building C, R & D Building, No. 666, Shendun 4th Road, Donghu New Technology Development Zone, Wuhan
