Recombinant Mouse Interleukin-1 beta (Il1b) (Active)

What expression systems are used for producing recombinant mouse IL-1β, and how do they impact the protein's properties?

Multiple expression systems are employed for producing recombinant mouse IL-1β, each offering distinct advantages and limitations:

When selecting a source, researchers should consider that E. coli-derived proteins may lack certain post-translational modifications that could affect bioactivity, while yeast-produced proteins often exhibit folding patterns more similar to mammalian-expressed proteins . Regardless of source, quality control standards typically ensure >95% purity as determined by SDS-PAGE analysis and minimal endotoxin contamination (<0.001 ng/μg) .

What are the optimal storage and reconstitution protocols for maintaining recombinant mouse IL-1β activity?

Proper handling of recombinant mouse IL-1β is crucial for maintaining its biological activity:

For lyophilized protein:

• Store at -20°C to -80°C in the original sealed container

• Avoid repeated exposure to freeze-thaw cycles

• Protect from light and moisture

Reconstitution protocol:

• Centrifuge the vial before opening to collect all material at the bottom

• Reconstitute in sterile phosphate-buffered saline (PBS) containing at least 0.1% carrier protein (such as BSA)

• For specific applications, a neutral buffer such as 50mM Tris-HCl with 50mM NaCl at pH 8.0 may be optimal

• Gently mix by inversion rather than vortexing to prevent protein denaturation

• Prepare single-use aliquots to avoid repeated freeze-thaw cycles

After reconstitution, the solution should be stored at 4°C for short-term use (1-2 weeks) or aliquoted and stored at -20°C for longer periods. Working solutions should be prepared fresh before experiments to ensure maximum activity.

What are the key biological activities of IL-1β in immunological research, and how can they be measured?

Interleukin-1β serves as a master regulator of inflammatory responses through multiple mechanisms:

Primary biological activities:

• Activation of T helper cells, particularly driving Th1 and Th17 differentiation pathways

• Induction of acute phase proteins and pro-inflammatory cytokine cascades

• Regulation of cell proliferation, differentiation, and apoptosis

• Promotion of neutrophil recruitment and activation

• Induction of fever, hypotension, and systemic inflammatory responses

These activities can be measured through several experimental approaches:

When measuring IL-1β activity, researchers should include appropriate controls such as heat-inactivated protein, IL-1 receptor antagonist (IL-1Ra) blockade, and dose-response analyses to confirm specificity.

How does recombinant mouse IL-1β interact with cellular receptors, and what downstream signaling pathways are activated?

IL-1β mediates its biological effects through specific receptor interactions and signaling cascades:

Receptor interactions:

• Primary binding occurs with type I IL-1 receptor (IL-1R1)

• IL-1R accessory protein (IL-1RAcP) is recruited to form a heterodimeric signaling complex

• IL-1 receptor antagonist (IL-1Ra) serves as a natural inhibitor by competing for receptor binding

The formation of the IL-1β/IL-1R1/IL-1RAcP complex initiates multiple intracellular signaling pathways:

• MyD88-dependent pathway:

  • Recruitment of adapter proteins (MyD88, IRAK1/4, TRAF6)

  • Activation of TAK1 complex

  • Parallel activation of:

    • NF-κB pathway (leading to pro-inflammatory gene expression)

    • MAPK pathways (p38, JNK, ERK) regulating cellular responses

• NLRP3 inflammasome pathway:

  • IL-1β can trigger further inflammasome activation

  • This creates a potential amplification loop for IL-1β production

  • NLRP3 expression correlates with IL-1β levels in inflammatory conditions

These pathways ultimately lead to the expression of hundreds of genes involved in inflammation, including additional cytokines, chemokines, adhesion molecules, and acute phase proteins, creating a coordinated inflammatory response.

How should researchers design dose-response experiments with recombinant mouse IL-1β for different cell types?

Optimal experimental design for IL-1β stimulation requires careful consideration of multiple factors:

Dose selection strategy:

• Begin with a wide concentration range (typically 0.1-100 ng/mL)

• For E. coli-derived proteins, start with higher concentrations (1-100 ng/mL)

• For yeast-derived proteins with higher bioactivity, lower concentrations may be sufficient (0.1-10 ng/mL)

• Include at least 5-6 concentration points with 3-5 fold dilutions between points

• Always include unstimulated controls

Cell type considerations:

Time-course considerations:

• Signaling events: 5-60 minutes

• Gene expression changes: 1-24 hours

• Protein secretion: 6-48 hours

• Morphological changes: 24-72 hours

For rigorous experimental design, include both dose-response and time-course elements, and validate findings with appropriate antagonists or neutralizing antibodies to confirm specificity.

What are the best practices for using recombinant mouse IL-1β in complex in vitro and in vivo experimental systems?

When employing IL-1β in advanced experimental systems, researchers should consider these methodological approaches:

For 3D cell culture systems:

• Increase IL-1β concentrations by 2-5 fold compared to 2D cultures due to diffusion limitations

• Monitor penetration using immunostaining for phosphorylated signaling molecules

• Consider gradient formation in larger 3D structures

• Validate expected biological effects with appropriate readouts before complex experiments

For co-culture systems:

• Different cell types may respond differently and influence each other's responses

• Use cell-specific markers to track responses in mixed populations

• Consider using reporter systems (e.g., NF-κB reporters) for cell-specific readouts

• Account for potential IL-1β-induced cell-cell communication

For in vivo applications:

Essential controls for in vivo work include:

• Vehicle-treated groups (matching buffer composition)

• IL-1Ra-treated groups to confirm IL-1β specificity

• Dose-response studies to establish optimal dosing

• Time-course sampling to capture peak effects

Researchers should report detailed methodological information, including source and concentration of IL-1β, administration details, and validation approaches used to confirm specificity.

How does the choice of expression system affect recombinant mouse IL-1β structure-function relationships in experimental research?

The expression system used to produce recombinant IL-1β can significantly impact its structural characteristics and functional properties:

E. coli expression system (as used by several manufacturers) :

• Lacks eukaryotic post-translational modifications

• May have different folding patterns than native protein

• Often requires higher concentrations to achieve equivalent biological effects

• Potential for endotoxin contamination requiring rigorous purification

• Advantages include high yield and cost-effectiveness

Yeast (Pichia pastoris) expression system :

• Provides eukaryotic-like post-translational modifications

• More closely mimics natural protein folding patterns

• Generally exhibits higher specific activity

• Lower risk of endotoxin contamination

• May better recapitulate in vivo activities

These differences have important experimental implications:

• Receptor binding kinetics may vary between proteins from different sources

• Neutralizing antibodies may have different efficacy against differently-produced IL-1β

• Dose requirements must be established empirically for each protein source

• Experimental reproducibility requires consistent sourcing

What are the current methodological challenges in studying IL-1β's role in complex disease models, and how can they be addressed?

Researchers face several methodological challenges when investigating IL-1β's contributions to disease pathogenesis:

Challenge 1: Distinguishing direct IL-1β effects from secondary inflammatory cascades

• Solution: Use timed interventions with IL-1Ra or neutralizing antibodies

• Employ genetic approaches (conditional knockouts, cell-specific deletions)

• Develop reporter systems to track primary vs. secondary responses

Challenge 2: Temporal dynamics of IL-1β activity

• Solution: Implement time-course experiments with frequent sampling

• Use biosensors or reporter systems for real-time monitoring

• Compare acute vs. chronic IL-1β exposure models

Challenge 3: Tissue-specific effects and microenvironmental factors

• Solution: Utilize tissue-specific conditional genetic models

• Develop organoid or tissue-specific 3D culture systems

• Combine in vivo imaging with targeted sampling

Challenge 4: Redundancy within the IL-1 family cytokine network

• Solution: Use combined blockade approaches (IL-1α/β/receptor)

• Employ genetic models with multiple cytokine/receptor deletions

• Conduct comprehensive cytokine profiling alongside intervention studies

Challenge 5: Contradictory findings between acute and chronic models

• Solution: Carefully design models with varying exposure durations

• Consider adaptation and compensatory mechanisms

• Integrate data from multiple model systems and human samples

To address these challenges, researchers should implement comprehensive experimental designs that include:

• Appropriate temporal sampling

• Combined pharmacological and genetic approaches

• Multiple readout systems to capture direct and indirect effects

• Validation across different model systems

How do findings from mouse IL-1β research translate to human inflammatory conditions, and what are the key considerations?

While mouse IL-1β research has generated valuable insights into inflammatory mechanisms, several factors influence its translational relevance to human conditions:

Structural and functional homology:

• Mouse and human IL-1β share approximately 75% amino acid sequence identity

• Human IL-1β is active on mouse cells, indicating functional conservation

• Receptor binding characteristics are largely conserved between species

Recent translational insights:

• IL-1β blocking agents have proven effective for various inflammatory and autoimmune conditions identified in mouse models, including rheumatoid arthritis, ischemic stroke, diabetes, uveitis, multiple sclerosis, and myocarditis

• Biomarker studies have demonstrated elevated IL-1β in human cardiovascular conditions, correlating with mouse model findings

• IL-1β's role in modulating adaptive immunity through Th17 pathway modulation is conserved between species

To enhance translational relevance, researchers should:

• Validate findings across multiple mouse strains

• Compare results with human cell culture systems

• Correlate mouse findings with human biospecimen data when possible

• Consider humanized mouse models for advanced translational studies

What are the emerging applications of recombinant mouse IL-1β in advanced research areas such as immunotherapy and disease modeling?

Recombinant mouse IL-1β is increasingly being utilized in cutting-edge research areas:

Immunotherapy development:

• As a target for testing novel antagonist approaches

• In combinatorial immunotherapy models to understand cytokine network interactions

• For ex vivo conditioning of immune cells for adoptive transfer

• In developing IL-1β-based adjuvants for vaccine research

Advanced disease modeling:

• Patient-derived xenograft models with IL-1β manipulation

• Organoid systems with controlled IL-1β exposure

• Biomaterial-based models of IL-1β-mediated inflammation

• CRISPR-engineered reporter systems for IL-1β pathway visualization

Mechanistic research has revealed IL-1β's role beyond classical inflammation:

• As a mediator in the NLRP3 inflammasome pathway, correlating with mast cell activation in chronic urticaria

• In modulating immune checkpoint molecules including PDL1, CTLA4, TIM3, and TIGIT

• As a biomarker with diagnostic potential for inflammatory conditions, as demonstrated by ROC curve analysis in CSU studies

Innovative methodological approaches include:

• Ultrasensitive immunoassays capable of detecting sub-pg/mL levels of IL-1β in biological samples

• Combined pharmacological and genetic manipulation of the IL-1 system

• Multi-omics approaches to understand IL-1β-mediated inflammatory networks

• Systems biology modeling of IL-1β-dependent inflammatory cascades

Researchers exploring these frontier areas should implement rigorous experimental design, comprehensive controls, and integrated analytical approaches to maximize the translational impact of their findings.