Recombinant Antibody Production
antibody reformatting

Apart from canonical antibodies, which are around 150 kDa and composed of antigen-binding fragment (Fab) and Fc region, a wide range of antibody formats can be produced through recombinant expression technology. These reformatted antibodies can benefit certain applications by improving solubility, thermostability, immunogenicity, etc. For instance, scFv is used in medical imaging due to its increased tumor penetration. Bispecific antibodies, on the other hand, enable the binding of two target antigens at once, which has a great potential in cancer immunotherapy and drug delivery.

As an expert in producing recombinant antibodies, Bon Opus Biosciences offers a broad range of antibody reformatting services including scFv, Fab, bispecific. A variety of different antibody formats are available and can be tailored to customer requirements.

Canonical Antibody

Fab

scFv

Bispecific

(with or without Fc)

Non-canonical mAb Production Service Package

 

ScFv Expression and Purification 

-Single Chain Variable Fragment (ScFv) is produced by connecting VH and VL together via a linker peptide. 

  1. Gene synthesis and codon optimization of VH and VL regions

  2. Vector construction with selected purification tag

  3. Transient expression in Expi293, ExpiCHO or E. coli

  4. Purification with tag and additional columns as requested

  5. Endotoxin removal as low as 0.1EU/mg (optional) 

  6. QC analysis: SDS-PAGE, SEC-HPLC, endotoxin level test

Deliverable:

- Purified scFv antibodies with customized QC standard

Turnaround time:

4 weeks plus shipping

 

Fab Expression and Purification

-Fragment Antigen-Binding (Fab) is produced by co-expressing a VH-CH1 and a VL-CL

  1. Gene synthesis and codon optimization of VH-CH1 and VL-CL

  2. Vector construction with selected purification tag

  3. Transient expression in Expi293, ExpiCHO or E. coli

  4. Purification

  5. Endotoxin removal as low as 0.1EU/mg (optional) 

  6. QC analysis: SDS-PAGE, SEC-HPLC, endotoxin level test

Deliverable:

- Purified antibodies with customized QC standard

Turnaround time:

4 weeks plus shipping

 

Bispecific Antibody Expression and Purification

-Bispecific antibody is produced by assembling two Fab regions. It can be manufactured in several formats (with or 

 without Fc) using different approaches.

  1. Gene synthesis and codon optimization of target Fab regions

  2. Cloning into expression vectors

  3. Transient expression in Expi293, ExpiCHO

  4. Purification

  5. Endotoxin removal as low as 0.1EU/mg (optional) 

  6. QC analysis: SDS-PAGE, SEC-HPLC, endotoxin level test

Deliverable:

- Purified bispecific antibodies with customized QC standard

Turnaround time:

4 weeks plus shipping

 

Case Study

100mg ScFv Expression and Purification​

  • Expressed by Expi293 with VH-(GGGGS)3-VL-6x His and purified by Ni-NTA

SDS-PAGE and Anti-His WB image

M: Molecular weight marker

Me: Culture medium

FT: Flow-through

W: Washes

E: Elution

  • Final Sample

SDS-PAGE

SEC-HPLC

  • Deliverable: 100mg purified antibody, >SEC-HPLC 99% purity, endotoxin < 0.1EU/mg

 

20mL Fab Expression and Purification​

  • Co-expressed in Expi293 and purified by Ni-NTA

SDS-PAGE and Anti-His WB image

M: Molecular weight marker

Me: Culture medium

FT: Flow-through

W: Washes

E: Elution

  • Final Sample

SDS-PAGE

1: Reducing sample

2: Non-reducing sample

  • Deliverable: 120ug purified antibody, 95% SDS-PAGE purity

 

Bispecific Antibody Expression and Purification​

Antibody properties: Human IgG4, Human Kappa Light Chain

  • Gene synthesis of two VH and one VL regions

  • Cloning into premade Fc vectors (contains one chimeric Fc*)

  • Co-expression with three constructs in Expi293

  • Purification by MabSelect SuRe LX and pH gradient elution

Target purification UV280 profile

Pool1: Fc*Fc* and process-related impurities

Pool2: Bispecific peak (FcFc*)

Pool3: FcFc

  • Final Sample

SDS-PAGE

1: Reducing sample

2: Non-reducing sample

  • Deliverable: Purified bispecific antibody, SDS-PAGE >95% purity