IL16 Antibody

What is IL-16 and what biological functions make it a relevant research target?

IL-16 is a chemoattractant cytokine and modulator of T-cell activation that has been proposed as a ligand for the co-receptor CD4. The secreted active form of IL-16 is commonly detected at sites of TH1-mediated inflammation, including those observed in autoimmune diseases, ischemic reperfusion injury (IRI), and tissue transplant rejection . IL-16 functions primarily in cytokine-mediated signaling pathways and immune response pathways. The protein has a canonical amino acid length of 1332 residues with a mass of approximately 141.8 kilodaltons and is localized in the nucleus and cytoplasm, with secreted forms being biologically active . IL-16 is expressed predominantly in the rectum, lymph node, colon, bone marrow, and appendix, making it relevant for research on diseases affecting these tissues .

What structural characteristics of IL-16 are important for antibody recognition?

Secreted IL-16 contains a characteristic PDZ domain that is crucial for its biological function and antibody recognition. Unlike typical PDZ domains that feature an exposed peptide-binding site located in a groove between the αB and βB structural elements, the solution structure of IL-16 reveals a tryptophan residue (Trp600) that obscures the recognition groove . This structural feature is significant because therapeutic antibodies like the 14.1 monoclonal antibody can induce conformational changes in this region, which involves rotation of the αB-helix and movement of the tryptophan residue to open up the binding site . Understanding these structural characteristics is essential for researchers designing experiments involving IL-16 targeting.

What types of IL-16 antibodies are available for research applications?

Researchers can select from over 449 commercially available IL-16 antibodies across 32 suppliers, with applications spanning Western blotting, ELISA, immunohistochemistry, and multiplex assays . The selection should be guided by the specific experimental requirements, including detection method, species reactivity, and whether a functional (neutralizing) or purely analytical approach is needed.

What is the mechanism of action for therapeutic IL-16 antibodies?

The mechanism of action of therapeutic anti-IL-16 antibodies, particularly the 14.1 monoclonal antibody, involves a surprising conformational change in the IL-16 PDZ domain upon binding. Structural analysis has revealed that antibody binding requires rotation of the αB-helix of IL-16, accompanied by movement of the peptide groove-obscuring tryptophan residue (Trp600), which opens up the binding site for interaction . This conformational change is critical for neutralizing IL-16 activity.

When the 14.1 antibody binds to IL-16, it prevents IL-16 from recruiting to its receptor (CD4), significantly attenuating inflammation and disease pathology in models of ischemic reperfusion injury and autoimmune diseases . The incubation of dendritic cells with this antibody results in reduced cell migration in cultures of epidermal cells and produces a measurable reduction in TH1-type inflammatory responses . This mechanism offers insights into how anti-IL-16 therapeutics might function in clinical settings.

How do the conformational changes in IL-16 affect antibody binding efficiency?

Experimental studies on the IL-16/antibody interaction reveal that the conformational change required for antibody binding has significant energetic implications. Researchers investigating this interaction created a W600A variant of IL-16 to probe the importance of the Trp600 side chain in regulating binding . Contrary to what might be expected, removing the blocking tryptophan side chain by substituting it with alanine decreased the affinity of the c14.1 antibody for IL-16 by approximately 10-fold (from an EC50 of 115 nM for native IL-16) .

This counterintuitive finding suggests that the energetic penalty arising from the conformational change induced in IL-16 upon antibody binding is more than offset by the network of Van der Waals interactions between the c14.1Fab CDR-H3 loop residues and the hydrophobic pocket formed by IL-16 residues Phe545, Leu547, Arg596, Ile603, and particularly the indole side chain from Trp600 . Researchers working with IL-16 antibodies should consider these structural dynamics when interpreting binding data or designing new therapeutic approaches.

How can IL-16 neutralizing antibodies be utilized in cancer therapy research?

Recent research has identified a novel application for IL-16 neutralizing antibodies in cancer therapy, particularly in combination with Aurora-A inhibitors. Studies have shown that tumor-intrinsic Aurora-A contributes to anti-tumor immunity depending on the status of lymphocyte infiltration . In colorectal cancer models, Aurora-A inhibition was found to upregulate IL-16, which may potentially impair the therapeutic effect of Aurora-A targeting alone .

The combination of IL-16 neutralization with Aurora-A inhibitors has been demonstrated to improve therapeutic responses in immune "hot" colorectal cancer tumors (those with high levels of lymphocyte infiltration) . This synergistic approach represents an emerging research direction, as bioinformatics analysis using TCGA datasets confirmed the upregulation of IL-16 in lower Aurora-A-expressed colorectal cancer, suggesting a regulatory role of Aurora-A in modulating the IL-16-mediated immune response .

Researchers investigating cancer immunotherapy should consider this interaction when designing experiments involving IL-16 neutralization or Aurora-A inhibition, as the combination may offer enhanced efficacy compared to single-agent approaches.

What experimental techniques are most effective for IL-16 detection using antibodies?

When selecting a detection method, researchers should consider that IL-16 exists in multiple forms - a full-length 141.8 kDa protein and processed secreted forms. For detecting secreted IL-16 in culture supernatants or serum, sandwich ELISA using capture and detection antibody pairs is highly effective . For cellular localization studies, immunohistochemistry with appropriate permeabilization is recommended to access both cytoplasmic and nuclear IL-16 .

How should IL-16 antibody specificity be validated for research applications?

Thorough validation of IL-16 antibody specificity is critical for obtaining reliable research results. A recommended validation approach includes:

• Knockout/knockdown controls: Compare antibody reactivity in wild-type samples versus those with IL-16 gene knockout or knockdown. This is particularly important given the multiple processing forms of IL-16.

• Peptide competition assays: Pre-incubate the antibody with recombinant IL-16 or specific peptides corresponding to the epitope to demonstrate signal reduction in subsequent detection assays.

• Cross-reactivity testing: For antibodies claimed to react with multiple species, confirm reactivity with recombinant IL-16 from each species. The conservation of IL-16 structure across species can vary, affecting antibody recognition.

• Binding site characterization: When possible, use structural information about the antibody-IL-16 interaction, such as that available for the 14.1 antibody, to predict and confirm epitope specificity .

• Application-specific validation: An antibody that works well in ELISA may not perform adequately in Western blot or IHC due to differences in how the epitope is presented in each technique.

What factors should be considered when designing experiments using IL-16 neutralizing antibodies?

When designing experiments with IL-16 neutralizing antibodies, researchers should consider several critical factors:

• Dose-response relationship: Establish a dose-response curve for the neutralizing antibody to determine optimal concentration. The EC50 for the c14.1 antibody binding to native hIL-16 has been reported as 115 nM, which can serve as a reference point .

• Timing of administration: For in vivo or cell culture studies, the timing of antibody administration relative to IL-16 production or stimulation is crucial. Based on studies in autoimmune disease models, pre-treatment or early intervention tends to be more effective .

• Duration of neutralization: Consider the half-life of the antibody in the experimental system and plan dosing accordingly. For in vivo studies, pharmacokinetic pilot studies may be necessary.

• Combination approaches: As demonstrated in cancer therapy research, IL-16 neutralization may have synergistic effects when combined with other treatments, such as Aurora-A inhibitors . Design experiments to test potential synergies relevant to your research question.

• Appropriate controls: Include isotype-matched control antibodies to account for non-specific effects of antibody administration. In the context of the 14.1 antibody studies, researchers included appropriate controls to distinguish specific IL-16 neutralization from general immunoglobulin effects .

• Confirmation of mechanism: When possible, include experiments that verify the proposed mechanism of neutralization, such as blocking IL-16 recruitment to CD4 or inducing conformational changes in the IL-16 PDZ domain .

How can researchers effectively troubleshoot IL-16 antibody experiments?

Common challenges with IL-16 antibody experiments and their solutions include: