Preface

IgA mediates microbial homeostasis in the intestinal mucosa. Microbial-reactive IgA is produced in the gut through both T-cell-independent and T-cell-dependent pathways , playing a role in preventing and promoting bacterial colonization and influencing bacterial gene expression, thereby providing exquisite control over the microbiota. IgA-microbiota interactions are highly diverse across individuals and populations, yet the factors driving this diversity remain poorly understood.

(Data source: Kelsey E , et al. Nat Rev Immunol. 2021)

Research ideas

SIgA is the main antibody secreted by the intestine, but the relationship between its functional effects and antigen specificity is still unclear. Rollenske's team analyzed how the polyclonal IgA response induced by a single microorganism affects microbial physiology and metabolism in parallel.

(Data source: Tim R , et al. Nature. 2021)

Study Results

After germ-free wild-type (WT) mice were pretreated with the gastric microorganism E. coli-HA107, HA107-reactive monoclonal antibodies were obtained from plasma cells in the lamina propria. The IGHV and IGKV gene combinations were cloned and amplified.

Recombinantly expressed mouse dimeric IgA clones were used to identify monoclonal antibodies coated on the surface of living intact HA107 bacteria by flow cytometry to determine the antibody binding characteristics.

After the model was reconstructed, the secretion and elimination of monomeric or dimeric mIgA after intravenous injection after bile duct ligation (BDL) were detected by ELISA to reveal the functional consequences of recombinant mSIgA.

Rollenske and colleagues investigated the consequences of secretory immunoglobulin A (SIgA) binding to the gut microbiota. They propose that SIgA's universal and unique epitope-specific effects regulate the gut microbiota in parallel. SIgA modulates the composition and function of the gut microbiota and regulates its interactions with the host.

(Data source: Oliver P , et al. Nat Rev Gastroenterol Hepatol. 2022)

Supplementary IgA research methods

Ig A expression vector construction: IgAh and Ig κ or IgJ chain transcripts were amplified by PCR using primers with 5′and 3′ends homologous to the respective expression vectors (mouse IgA (Addgene plasmid ID #244892), Ig κ (Addgene plasmid #127157), IgJ (Addgene plasmid ID #244893);

Recombinant Monoclonal Dimeric Murine IgA Expression：HEK293F suspension cells were transfected with equal amounts of Ig Ah , the corresponding Ig κ, and Ig J using PEI-mediated co-transfection. After 24 hours, an equal amount of culture medium was added, and the cells were cultured for an additional 5 days under constant agitation, after which the supernatant was collected by centrifugation.

Antibody purification by affinity chromatography: Cell culture supernatant was incubated with CaptureSelect LC-kappa affinity medium (Thermo Fisher Scientific) overnight at 4°C on a rotary shaker. The supernatant was removed and the affinity medium was transferred to an equilibrated column and allowed to drain by gravity. The gravity column was equilibrated with 3 column volumes (CVs) of PBS. After equilibration, the column was placed in an empty collection tube, 250 µL of 0.1 M citrate buffer (pH 2.0) was added to the column, and the column was drained by gravity or gentle pressure. The column was then neutralized by adding a 1:2.5 volume of 1 M Tris-HCl (pH 9). The column was concentrated using a centrifugal concentrator with a 100 kDa cutoff membrane (Millipore) and dialyzed overnight at 4°C against PBS or 0.9% (w/v) NaCl.

Recombinant IgA Detection: Purified IgA was analyzed by SDS-PAGE using 4%–20% prepreg polyacrylamide gels under reducing or non-reducing conditions.

(Data source: Antonia G , et al. Eur J Immunol. 2025)