Recombinant Mouse Angiopoietin-3

Recombinant mouse ANGPTL3 is a valuable tool for lipid metabolism research, offering several compelling advantages for your experimental work.

Structural and Functional Characteristics

Recombinant mouse ANGPTL3 is a secreted glycoprotein with well-defined structural domains that make it ideal for mechanistic studies. The mature protein contains an N-terminal coiled-coil domain and a C-terminal fibrinogen-like domain. This structural organization is functionally important because proteolytic removal of the fibrinogen-like domain activates ANGPTL3 and enhances its ability to inhibit lipoprotein lipase (LPL) both in vitro and in vivo.

Primary Research Applications

Lipid Metabolism Studies

The primary application of recombinant mouse ANGPTL3 is investigating lipoprotein metabolism. ANGPTL3 directly inhibits both lipoprotein lipase and endothelial lipase, enzymes responsible for hydrolyzing circulating triglycerides. This inhibitory activity requires a putative heparin-binding motif located N-terminal to the coiled-coil domain. By using recombinant ANGPTL3, you can study how this protein regulates triglyceride levels without altering VLDL or HDL secretion or uptake.

Disease Model Development

Recombinant mouse ANGPTL3 is particularly useful for studying dysregulated lipid metabolism associated with obesity and diabetes. Dysregulated ANGPTL3 expression and elevated plasma triglyceride levels are characteristic of certain obese and diabetic mouse strains. Using recombinant protein allows you to experimentally manipulate ANGPTL3 levels and observe downstream metabolic effects in controlled conditions.

Cell Biology and Vascular Research

Beyond lipid metabolism, the fibrinogen-like domain of ANGPTL3 interacts with integrin alpha V beta 3 to induce endothelial cell adhesion, migration, and neovascularization. Additionally, ANGPTL3 secreted by fetal liver cells promotes the expansion of hematopoietic stem cells, making recombinant protein useful for stem cell biology applications.

Experimental Advantages

Using recombinant mouse ANGPTL3 provides several experimental benefits: you can achieve precise protein concentration control, conduct reproducible in vitro assays, and perform dose-response studies that would be difficult with endogenous protein. The mouse ortholog shares 77% amino acid sequence identity with human ANGPTL3, making findings translatable to human biology while maintaining the advantages of mouse model compatibility.

Yes, you can use recombinant Mouse Angiopoietin-3 (ANGPTL3) as a standard for quantification or calibration in your ELISA assays, provided it is of high purity and its concentration is accurately determined. This is a common and accepted practice in quantitative ELISA development and validation.

Key considerations and supporting details:

• Recombinant proteins are widely used as ELISA standards when purified native protein is unavailable or impractical to obtain. The standard curve in ELISA is typically generated using a purified or recombinant protein whose concentration is known.
• Commercial mouse ANGPTL3 ELISA kits routinely use recombinant ANGPTL3 as the standard for calibration, as indicated in multiple kit manuals and technical datasheets. These kits demonstrate that recombinant ANGPTL3 is suitable for generating standard curves and quantifying ANGPTL3 in biological samples.
• Critical requirements for use as a standard:
  • The recombinant ANGPTL3 must be of high purity and free from contaminants that could interfere with antibody binding or detection.
  • The protein concentration must be accurately determined, ideally by a method such as absorbance at 280 nm (using a known extinction coefficient), BCA assay, or amino acid analysis.
  • The recombinant protein should be reconstituted and diluted according to best practices to ensure stability and reproducibility.
• Matrix effects: When quantifying ANGPTL3 in complex biological samples (e.g., serum, plasma), ensure that the recombinant standard behaves similarly to endogenous ANGPTL3 in your assay system. Some kits report recovery and linearity data for recombinant ANGPTL3 spiked into various matrices, supporting its use as a standard.
• Validation: If you are developing a custom ELISA, validate that your recombinant ANGPTL3 standard produces a standard curve that is parallel to the response of endogenous ANGPTL3 in your samples. This ensures accurate quantification.

Summary of best practices:

• Use a highly purified recombinant Mouse ANGPTL3 with a well-characterized concentration.
• Prepare the standard curve in the same matrix as your samples if possible, or validate recovery and parallelism.
• Follow standard ELISA guidelines for standard preparation, dilution, and curve fitting (e.g., 4-PL or 5-PL regression).

In conclusion, recombinant Mouse Angiopoietin-3 is appropriate for use as a standard in ELISA quantification, provided you adhere to best practices for protein standards and assay validation.

Recombinant Mouse Angiopoietin-like 3 (ANGPTL3) has been validated for several key applications in published research, primarily related to lipid metabolism, hematopoietic stem cell regulation, and endothelial cell function. The main applications supported by published studies include:

1. Lipoprotein Lipase (LPL) Inhibition Assays

   • Recombinant Mouse ANGPTL3 has been used to study its direct inhibition of LPL activity, a key enzyme in triglyceride metabolism. Dose-dependent inhibition of LPL activity has been demonstrated in bioassays, confirming its role in regulating circulating triglyceride levels.
   • Reference: Bio-Techne (Catalog # 9899-AN), Nature (2025), R&D Systems (136-AN-050).

2. Hematopoietic Stem Cell (HSC) Expansion and Regulation

   • ANGPTL3 has been shown to promote the expansion of hematopoietic stem cells, particularly in ex vivo culture systems. Studies using recombinant ANGPTL3 have demonstrated its ability to support HSC expansion and maintenance.
   • Reference: R&D Systems, BMC Nephrol (2015), PMC3031476.

3. Endothelial Cell Adhesion, Migration, and Neovascularization

   • The fibrinogen-like domain of ANGPTL3 interacts with integrin αVβ3, promoting endothelial cell adhesion, migration, and neovascularization. Recombinant ANGPTL3 has been used in assays to study these processes.
   • Reference: R&D Systems, PMC3511699.

4. ELISA and Protein Quantification

   • Recombinant Mouse ANGPTL3 is used as a standard in ELISA kits for quantifying ANGPTL3 levels in biological samples such as serum, plasma, and cell culture supernatants.
   • Reference: BioVendor, R&D Systems.

5. In Vivo and In Vitro Functional Studies

   • Recombinant ANGPTL3 has been used in both in vitro and in vivo models to study its effects on lipid metabolism, including triglyceride and cholesterol regulation, as well as its interaction with other ANGPTL family members (e.g., ANGPTL8).
   • Reference: PMC3511699, Nature (2025), ACC.org.

6. Protein-Protein Interaction Studies

   • Recombinant ANGPTL3 has been used in co-immunoprecipitation and interaction assays to study its physical and functional interactions with other proteins, such as ANGPTL8.
   • Reference: PMC3511699.

In summary, Recombinant Mouse Angiopoietin-like 3 has been validated for use in bioassays, ELISA, HSC expansion, endothelial cell function studies, lipid metabolism research, and protein interaction studies.

To reconstitute and prepare Recombinant Mouse Angiopoietin-3 (ANGPTL3) protein for cell culture experiments, follow these steps based on best practices and product-specific protocols:

• Reconstitution:

  • Use sterile phosphate-buffered saline (PBS) to reconstitute the lyophilized protein.
  • The recommended concentration for reconstitution is 100 μg/mL in sterile PBS.
  • Before opening the vial, briefly centrifuge it to collect all lyophilized material at the bottom.
  • Add the calculated volume of PBS directly to the vial, gently swirl or invert to dissolve. Avoid vigorous vortexing or foaming, as this can denature the protein.
  • Allow the solution to sit at room temperature for at least 15 minutes with gentle agitation to ensure complete dissolution.

• Aliquoting and Storage:

  • Once reconstituted, aliquot the protein into small volumes to avoid repeated freeze-thaw cycles, which can degrade protein activity.
  • Store aliquots at -20 °C to -70 °C for long-term storage (up to 3 months), or at 2–8 °C for short-term use (up to 1 month) under sterile conditions.
  • Avoid repeated freeze-thaw cycles.

• Preparation for Cell Culture:

  • Before adding to cell culture, dilute the reconstituted stock to the desired working concentration using sterile cell culture medium or PBS.
  • Typical working concentrations for functional assays (e.g., stimulation of hematopoietic stem cells) range from 20–120 ng/mL, but optimal concentrations should be determined empirically for your specific assay.
  • If the protein contains additives (e.g., CHAPS, MOPS), ensure that the final concentration of these components is compatible with your cell culture system.

• General Best Practices:

  • Always use sterile technique to prevent contamination.
  • If the protein is sensitive to oxidation or aggregation, consider adding carrier proteins (e.g., 0.1% BSA) to the dilution buffer, unless this interferes with your assay.
  • Consult the product-specific datasheet or certificate of analysis for any unique handling instructions.

Summary Table: Reconstitution and Handling

These guidelines ensure the protein remains stable and functional for cell culture applications. Always verify with the specific product datasheet for any additional requirements.