Recombinant Mouse Limitin

Recombinant Mouse Limitin (also known as IFN-zeta) is used in research applications primarily for its unique antiviral activity and its ability to selectively inhibit B-lineage lymphoid-cell development in both in vivo and in vitro systems.

Key scientific reasons to use recombinant Mouse Limitin in your research include:

• Antiviral Activity: Limitin is a secreted interferon-like glycoprotein with demonstrated antiviral properties, making it valuable for studies on viral infection mechanisms, host-pathogen interactions, and the development of antiviral therapeutics.

• Selective Inhibition of B-Cell Development: Unlike other interferons, Limitin specifically inhibits the development of B-lineage lymphoid cells without broadly suppressing other hematopoietic lineages. This property is useful for dissecting the molecular pathways of B-cell differentiation and for modeling diseases or conditions where B-cell suppression is relevant.

• Immunomodulatory Research: Because of its lineage-specific effects, recombinant Mouse Limitin is a powerful tool for studying immune regulation, hematopoiesis, and the interplay between cytokines and immune cell development.

• Reproducibility and Consistency: Recombinant proteins offer batch-to-batch consistency, defined activity, and purity, which are critical for reproducible experimental results and for mechanistic studies where precise cytokine dosing is required.

• In Vivo and In Vitro Applications: Recombinant Mouse Limitin can be used in both cell culture and animal models, enabling translational research from basic immunology to preclinical studies.

In summary, recombinant Mouse Limitin is a specialized tool for immunology and virology research, particularly when investigating B-cell biology, antiviral responses, or the effects of interferon-like cytokines in mouse models. Its recombinant form ensures high purity, defined activity, and experimental reproducibility.

Recombinant Mouse Limitin (IFN-ζ) can be used as a standard for quantification or calibration in ELISA assays, provided it is compatible with your assay’s antibodies and detection system. However, several critical factors must be considered to ensure accurate and reliable quantification:

• Antibody Specificity: The recombinant Limitin must be recognized by the capture and detection antibodies used in your ELISA. If your assay is designed specifically for Limitin/IFN-ζ, and the antibodies are validated for this analyte, recombinant Limitin is appropriate as a standard.
• Protein Purity and Formulation: Use a highly purified recombinant protein. Some protocols recommend using a carrier protein (such as BSA) to stabilize the standard, especially for repeated freeze-thaw cycles or low-concentration dilutions.
• Standard Curve Preparation: Prepare the standard curve using serial dilutions of the recombinant Limitin in the same matrix as your samples (e.g., serum, plasma, or buffer), as matrix effects can influence assay performance.
• Validation: It is essential to validate the standard curve with your specific ELISA setup. This includes confirming linearity, recovery, and parallelism between the recombinant standard and endogenous Limitin in your sample matrix.
• Lot-to-Lot Variability: Recombinant proteins may show variability between lots. Assign the value of your standard based on its performance in the ELISA, not solely on the mass stated on the vial.

Best Practices:

• Always follow the manufacturer’s instructions for reconstitution and storage of the recombinant standard.
• Use curve-fitting software (e.g., 4-parameter logistic regression) for optimal standard curve generation.
• If using a custom or in-house ELISA, ensure the recombinant Limitin is immunologically equivalent to the endogenous protein in your samples.

Summary Table:

In conclusion: Recombinant Mouse Limitin is suitable as an ELISA standard if it is validated for your assay system and all the above considerations are addressed.

Recombinant Mouse Limitin (also known as IFN-zeta) has been validated in published research for several key applications, primarily in immunology and hematopoiesis studies:

• Antiviral activity assays: Limitin has been shown to possess antiviral properties, and its activity is commonly validated by measuring its ability to protect L-929 mouse fibroblast cells from viral lysis. This is a standard bioassay for type I interferons, including Limitin.

• Inhibition of B-lineage lymphoid-cell development: Limitin selectively inhibits the development of B-lineage lymphoid cells both in vivo and in vitro, making it a valuable tool for studying lymphopoiesis and immune regulation.

• Modulation of immune cell proliferation and function: Limitin has been reported to modify the proliferation and function of peripheral T lymphocytes, natural killer (NK) cells, and bone marrow stromal cells, supporting its use in studies of immune cell biology and hematopoietic microenvironments.

• Receptor binding and neutralization assays: Recombinant mouse IFN-alpha/beta receptor proteins have been used to validate Limitin’s activity by inhibiting its protective effects in cell-based assays, confirming its specificity and mechanism of action.

Additional validated uses (based on broader interferon research and vendor protocols, where Limitin is included among type I IFNs):

• Cell signaling studies: Limitin is used to investigate interferon receptor signaling pathways, including STAT phosphorylation and downstream gene expression, in various cell types.

• ELISA standard: Carrier-free recombinant Limitin is used as a standard in ELISA assays to quantify endogenous or recombinant protein levels in biological samples.

Summary Table: Validated Applications of Recombinant Mouse Limitin

Key details:

• Most published research uses recombinant mouse Limitin in in vitro cell-based assays, in vivo mouse models, and as a standard for immunoassays.
• Its primary scientific value lies in dissecting type I interferon biology, immune regulation, and antiviral defense mechanisms.

If you require protocols or more specific experimental details for any of these applications, please specify the context or assay type.

To reconstitute and prepare Recombinant Mouse Limitin protein for cell culture experiments, follow these general steps based on best practices for recombinant cytokines and interferons:

1. Centrifuge the vial briefly (20–30 seconds in a microcentrifuge) before opening to ensure all lyophilized protein is at the bottom of the vial.

2. Reconstitution buffer:

   • Most recombinant proteins, including Limitin/IFN-zeta, are reconstituted in sterile phosphate-buffered saline (PBS) or sterile distilled water.
   • For enhanced stability and to prevent adsorption to plastic, add a carrier protein such as 0.1–1% bovine serum albumin (BSA) if recommended.
   • Typical reconstitution concentrations are 10–100 μg/mL in sterile PBS with carrier protein.

3. Reconstitution procedure:

   • Add the appropriate volume of buffer to achieve the desired concentration (e.g., 10 μg/mL or 100 μg/mL, depending on your experimental needs).
   • Gently mix by pipetting or slow inversion.
   • Allow the protein to dissolve for 15–30 minutes at room temperature with gentle agitation. Avoid vigorous shaking or vortexing to prevent foaming and protein denaturation.

4. Aliquot and storage:

   • Once fully dissolved, aliquot the solution to avoid repeated freeze-thaw cycles.
   • Store aliquots at –20°C to –70°C for long-term storage, or at 2–8°C for short-term use (up to one month).
   • Avoid repeated freeze-thaw cycles, which can reduce protein activity.

5. Preparation for cell culture:

   • Dilute the reconstituted stock into your cell culture medium immediately before use.
   • Ensure the final concentration matches your experimental design (e.g., dose-response studies).
   • If using serum-free medium, confirm that the carrier protein does not interfere with your assay.

6. Quality check:

   • Confirm protein presence and integrity by SDS-PAGE if needed.
   • Use endotoxin-free reagents and sterile technique throughout to prevent contamination.

Example protocol for 10 μg/mL stock:

- Centrifuge vial briefly.- Add sterile PBS (with 0.1% BSA) to the vial to achieve 10 μg/mL.- Mix gently and let dissolve at room temperature for 15–30 min.- Aliquot and store at –20°C or below.- Dilute into cell culture medium for experiments.

Notes:

• Always consult the specific product datasheet for recommended buffer and concentration, as some preparations may require different conditions.
• Recombinant Limitin is a secreted glycoprotein with IFN-like activity, so handle as you would other recombinant interferons.

If your preparation is not lyophilized or has a different formulation, adjust accordingly. For optimal results, use endotoxin-free buffers and maintain sterile conditions throughout.