Porcine Immunoglobulin G

What Are the Primary IgG Subclasses in Pigs and Their Functional Characteristics?

Porcine IgG subclasses include IgG1, IgG2a, IgG2b, IgG2c, IgG3, IgG4, IgG5b, and IgG5c. These differ significantly in Fc-mediated effector functions:

Key Methodological Considerations:

• Selection for Therapeutic Antibodies: Prioritize IgG1, IgG2a, IgG2b, IgG2c, or IgG4 for Fc-mediated effector functions. IgG3 and IgG5b are functionally limited .

• Assay Design: Use cytokine-activated porcine NK cells (e.g., IL-2, IL-12, IL-18) for ADCC/ADCP assays, and rabbit/pig complement for CDCC .

How Do Cytokines Influence IgG Isotype Production in Pigs?

Cytokine-driven polarization of immune responses governs IgG subclass expression:

Experimental Approaches:

• In Vitro Studies: Culture porcine B cells with recombinant cytokines (e.g., rpIL-10, rpIFN-γ) to analyze IgG1/IgG2 ratios .

• In Vivo Models: Use pathogens like Actinobacillus pleuropneumoniae to correlate IgG1:IgG2 ratios with protection .

What Methodologies Are Used to Characterize Porcine IgG-Binding Proteins?

To identify IgG-binding proteins (e.g., enolase), researchers employ:

A. Western Blotting

• Resolve proteins (e.g., enolase, casein) via 12% SDS-PAGE.

• Transfer to PVDF membranes and probe with porcine IgG (pIgG) or human IgG (hIgG) at 20 µg/ml.

• Detect binding via HRP-conjugated Staphylococcal Protein A (SPA) .

B. Dot Blot Analysis

• Spot recombinant proteins on PVDF membranes.

• Block with 5% skim milk, then incubate with pIgG/hIgG.

• Visualize binding using DAB .

Critical Controls:

• Negative Controls: Use casein to exclude non-specific binding .

• Validation: Confirm binding regions via N-terminal sequencing or mutagenesis.

How Do Genomic Organization and Evolution Shape Porcine IgG Subclass Functions?

Functionally similar subclasses cluster in the genome:

• IgG1, IgG2a, IgG2b, IgG2c, IgG4: Grouped in a region associated with strong effector functions.

• IgG3, IgG5b, IgG5c: Located in distinct regions with reduced functionality .

Bioinformatics Workflows:

• Align porcine IGHG genes (e.g., IGHG1: AB699686) to identify homologs.

• Use phylogenetic tools (e.g., RAxML) to infer subclass relationships.

• Correlate genomic clustering with functional data (e.g., ADCC/CDCC assays) .

What Are Key Discrepancies Between In Vitro and In Vivo Data for Porcine IgG?

Example: IgG3’s role in newborns vs. adults:

• In Vitro: No Fc-mediated functions (e.g., CD107a mobilization) .

• In Vivo: High IgG3 expression in neonatal gut-associated lymphoid tissues (GALT) suggests pre-adaptive immunity .

Resolving Contradictions:

• Epitope-Specificity: Validate in vivo efficacy with HA-sialic acid-binding antibodies .

• Model Systems: Use pig models for influenza to assess therapeutic IgG subclasses (e.g., pb27 IgG1) .

How Should Researchers Select IgG Subclasses for Therapeutic Antibodies?

Decision Framework:

Case Study: For anti-influenza antibodies, prioritize IgG1 for balanced neutralization and effector functions .

What Are Emerging Research Directions for Porcine IgG?

• Optimizing Fc Functions: Engineer IgG subclasses (e.g., IgG3) to enhance effector activity.

• Neonatal Immunity: Investigate IgG3’s role in pre-adaptive responses in piglets.

• Cross-Species Therapeutics: Develop porcine mAbs for swine models to avoid anti-human responses .

Methodological Innovations:

• Genome Editing: CRISPR/Cas9 to modify IGHG loci for subclass switching.

• Single-Cell RNA-seq: Profile B-cell responses to pathogens like African swine fever virus.