Cell-Free Antibody Production Service: High-Yield Synthesis of scFvs, Fabs, and Full-Length IgGs

The Cell-Free Antibody Production Service utilizes advanced Cell-Free Protein Synthesis (CFPS) systems, typically based on E. coli or Insect Lysates, to rapidly and efficiently synthesize functional antibody molecules. This platform is particularly effective for fragments such as single-chain variable fragments (scFvs), Fab fragments, and VHH/Nanobodies , as well as complex multi-chain structures like full-length Immunoglobulin G (IgG).

CD Biosynsis offers a specialized service emphasizing the critical requirements for antibody expression: efficient disulfide bond formation and precise assembly of heavy and light chains. By precisely controlling the reaction environment, including the redox potential (GSH/GSSG ratio) and chaperone supplementation, we achieve superior yields and folding fidelity compared to traditional in vivo methods. This service is ideal for drug discovery, structural analysis, and diagnostic assay development where speed and quality are paramount.

Highlights

Key advantages of utilizing Cell-Free systems for antibody production:

• Enhanced Disulfide Bonding: Precise tuning of the redox environment is implemented to maximize the correct formation of stabilizing disulfide bonds in the antibody fragments.
• Rapid Screening and Prototyping: Synthesis and screening of hundreds of antibody variants (e.g., from phage display outputs) can be completed in hours, accelerating lead identification.
• High Yields of Fragments: Particularly high expression levels are achieved for scFvs, Fabs, and Nanobodies using optimized continuous-exchange E. coli CFPS systems.
• Flexibility for Toxins: Expression of antibody fragments targeting highly toxic antigens or containing toxic fusion tags, which is challenging in living cells.

Antibody Targets

Our Cell-Free Antibody Production Service supports the synthesis of various formats:

Single-Chain Fvs (scFvs)

Rapid, high-yield synthesis of single-chain fragments for binding assays, imaging, and structural studies.

Fab Fragments

Co-expression of heavy and light chain Fd fragments optimized for correct disulfide linkage and heterodimer assembly.

VHH / Nanobodies

High-efficiency production of these stable, small, single-domain antibodies, often requiring specific folding optimization.

Full-Length IgGs (IgG1, IgG4)

Co-expression of four chains (2 heavy, 2 light) using specialized CFPS systems for functional screening and initial characterization.

Key Features & Optimization

Technical features ensuring functional antibody production:

Redox Buffer Control

Precise balance of the glutathione couple (GSH/GSSG) to drive the formation of intra- and inter-chain disulfide bonds.

Multi-Chain Co-Expression

Optimized systems for the simultaneous co-expression of multiple DNA templates (heavy and light chains) for complex assembly.

Chaperone Supplementation

Addition of folding-assisting proteins (e.g., DsbC, protein disulfide isomerase) to enhance correct folding, especially in E. coli lysates.

Contained PTMs (Glycosylation)

While standard CFPS systems lack complex human glycosylation, Insect Lysates provide high-mannose glycosylation suitable for many applications.

Scale Flexibility

Seamless scaling from microliter screening volumes to milligram preparative synthesis using continuous-exchange CFPS methods.

Workflow

Our systematic approach for functional antibody synthesis:

• Template Preparation: Design and optimize separate DNA templates for the heavy and light chains, often including an affinity tag for purification.
• Co-Expression Setup: Templates are combined with the CFPS lysate (E. coli or Insect) and the customized redox buffer system for optimal disulfide bond formation.
• In Vitro Synthesis: Reaction is incubated in a continuous-exchange mode for high yield, promoting co-assembly of the chains.
• Purification: Antibody fragments or IgGs are purified via affinity chromatography (e.g., Protein A/G or specific tag).
• Quality Control (QC) & Functionality: Analysis includes SDS-PAGE/Western Blot for yield and assembly, plus a functional assay (e.g., ELISA, SPR) to confirm binding activity.

We provide essential assurance for high-quality antibody expression outcomes:

• Guaranteed Assembly: Confirmation of correct chain pairing and disulfide linkage for multi-chain fragments (Fab, IgG).
• Functional Binding: Assurance that the produced antibody fragment retains verified binding affinity to the target antigen.
• Yield Optimization: Implementation of continuous-exchange CFPS protocols to maximize final yield for preparative applications.
• Folding Expertise: Utilization of CFPS's open nature to optimize buffer conditions specifically for difficult-to-fold or unstable VHH/scFv targets.