SERPINF2 Mouse

What is the molecular structure and primary function of mouse SERPINF2?

Mouse SERPINF2 (alpha-2-antiplasmin) is synthesized as a 491 amino acid precursor with a 27 amino acid signal peptide and a short propeptide (residues 28-39) . The mature protein functions as the primary physiological inhibitor of plasmin, which is responsible for dissolving fibrin clots . Beyond plasmin inhibition, SERPINF2 also efficiently inhibits trypsin and chymotrypsin .

The protein has a molecular mass of approximately 53.2 kDa, though in SDS-PAGE analysis it typically appears at 70-85 kDa due to post-translational modifications . The amino acid sequence for mouse SERPINF2 (Val28-Lys491) corresponds to UniProt/NCBI accession number Q61247 .

Methodologically, researchers can study SERPINF2's inhibitory function through:

• Plasmin inhibition assays using chromogenic substrates

• Clot lysis assays comparing wild-type and SERPINF2-deficient plasma

• Binding studies examining SERPINF2-protease complex formation

What is the tissue expression pattern of SERPINF2 in mice?

Mouse SERPINF2 shows a tissue-specific expression pattern with primary production sites in:

• Liver: High expression (major production site)

• Kidney: High expression

Moderate expression levels are found in:

• Muscle tissue

• Intestine (confirmed by immunohistochemistry)

• Central nervous system

• Placenta

This distribution suggests SERPINF2 serves as a key regulator of plasmin-mediated proteolysis across multiple tissues . The widespread but differential expression indicates importance beyond blood coagulation, potentially in tissue remodeling and other proteolytic processes.

For experimental detection, immunohistochemistry protocols using specific antibodies (such as Goat Anti-Mouse Serpin F2/α2-Antiplasmin at 5 μg/mL) have successfully visualized SERPINF2 in mouse intestinal tissue using HRP-DAB detection systems .

What phenotypes are observed in SERPINF2-deficient mice?

Mice homozygous for disruptions in the Serpinf2 gene exhibit a surprisingly mild phenotype considering the protein's important role in fibrinolysis:

The enhanced clot lysis aligns with SERPINF2's known function as an inhibitor of plasmin. Without SERPINF2's inhibitory action, plasmin activity increases, leading to faster dissolution of fibrin clots . This relatively mild phenotype suggests potential compensatory mechanisms within the fibrinolytic system.

For researchers, these observations highlight the importance of challenge models that may reveal more pronounced phenotypes than baseline conditions alone. Standardized clot lysis assays and specialized thrombosis models are recommended for comprehensive phenotypic characterization.

What mouse models are available for studying SERPINF2?

Several mouse models are available for SERPINF2 research:

• SERPINF2 knockout mice:

  • Complete deletion of functional Serpinf2 gene

  • Show enhanced clot lysis but otherwise normal phenotype

  • Valuable for studying basic physiological roles

• B-hSERPINF2 humanized mice:

  • Catalog number: 112340

  • Strain name: C57BL/6N-Serpinf2/Bcgen tm1(SERPINF2)Bcgen

  • Background: C57BL/6N

  • Gene targeting strategy: Mouse Serpinf2 exons 2-10 replaced with human SERPINF2 exons 2-10

  • Human SERPINF2 mRNA detectable only in homozygous mice

  • Application: Pharmacodynamics and safety evaluation of anticoagulant and antitumor drugs

These models provide complementary platforms for investigating loss-of-function effects and human-relevant therapeutic targets. The humanized model is particularly valuable for translational research, allowing evaluation of human-targeted therapies in an in vivo system.

What experimental techniques provide optimal results when studying SERPINF2 in mouse models?

Protein Detection and Localization:

• Western Blotting: Using specific antibodies such as Goat Anti-Mouse Serpin F2/α2-Antiplasmin Antigen Affinity-purified Polyclonal Antibody . Note that approximately 15% cross-reactivity with recombinant human Serpin F2 may occur .

• Immunohistochemistry: Most effective on frozen sections using 5 μg/mL antibody concentration with HRP-DAB detection systems . Perfusion fixation improves tissue preservation and antigen accessibility.

• ELISA: Direct ELISA techniques can quantify SERPINF2 in plasma and tissue extracts with high sensitivity.

Functional Assays:

• Plasmin Inhibition Assays: Measuring SERPINF2's inhibitory effect using chromogenic or fluorogenic substrates specific for plasmin.

• Clot Lysis Assays: Ex vivo assessment of fibrinolysis rates comparing plasma from wild-type versus SERPINF2-deficient mice.

• Thromboelastography: Provides comprehensive assessment of clot formation, stability, and lysis relevant to SERPINF2 function.

Gene Expression Analysis:

• RNA Sequencing: For tissue or cell-type specific transcriptomic profiling following proper cell isolation techniques .

• RT-qPCR: For targeted expression analysis using validated primer sets spanning exon junctions.

Cell-Type Specific Analysis:
For cell-specific studies, isolation protocols have been validated:

• Microglia isolation using magnetic cell separation

• Astrocyte purification using immunopanning with HepaCAM antibodies

These techniques enable comprehensive investigation of SERPINF2's expression, localization, and function in diverse experimental contexts.

How do mouse and human SERPINF2 compare, and what are the implications for translational research?

Mouse and human SERPINF2 share significant structural and functional similarities with important differences:

Structural Comparison:

• Similar size and domain organization

• Sufficient sequence homology that antibodies against mouse SERPINF2 show approximately 15% cross-reactivity with human SERPINF2

• Both contain characteristic serpin domain structure with reactive center loop

Functional Comparison:

• Both primarily inhibit plasmin and secondarily inhibit trypsin and chymotrypsin

• Both are expressed predominantly in liver and kidney with moderate expression in other tissues

Research Implications:
The B-hSERPINF2 humanized mouse model provides a valuable translational platform:

• Expression validation confirms human SERPINF2 mRNA is detectable only in homozygous B-hSERPINF2 mice

• This model enables preclinical evaluation of therapeutics targeting human SERPINF2

For translational researchers, the humanized model offers advantages over standard mouse models, particularly for:

• Testing human-specific therapeutics

• Evaluating anticoagulant drugs' efficacy and safety

• Investigating SERPINF2's role in tumor biology

When designing translational studies, researchers should consider both the similarities and differences between species, using the humanized model when human-specific interactions are paramount.

What are critical experimental design considerations when using SERPINF2 mouse models?

When designing experiments with SERPINF2 knockout or humanized mouse models, several critical factors should be considered:

Genetic Background and Controls:

• SERPINF2-related phenotypes may vary with genetic background

• The B-hSERPINF2 model is on C57BL/6N background

• Use littermate controls whenever possible

• For humanized models, appropriate controls include both wild-type mice and species-specific knockouts

Age and Sex Variables:

• Include both male and female mice with appropriate sample sizes

• Age-dependent effects may be significant; published microglia studies used 1-month-old mice while astrocyte studies used 2-month-old mice

• Analyze data by sex to identify potential sex-specific effects in coagulation parameters

Model Validation:

• Confirm genotype using PCR-based methods

• Validate protein expression/absence using Western blotting

• For humanized models, confirm functional activity of human protein in mouse environment

Challenge Models:

• Baseline phenotypes may be subtle; challenge models can reveal cryptic phenotypes

• Consider thrombotic challenges, inflammatory stimuli (e.g., LPS), or tissue-specific injury models

• The relatively normal phenotype of knockout mice suggests compensatory mechanisms that may be overcome under stress conditions

Special Handling Requirements:
For recombinant SERPINF2 protein used in experiments:

• Use manual defrost freezer and avoid repeated freeze-thaw cycles

• Store at -20 to -70°C for long-term stability; after reconstitution, store at 2-8°C for up to 1 month or -20 to -70°C for up to 6 months under sterile conditions

Careful attention to these considerations will enhance experimental reproducibility and translational relevance.

How does SERPINF2 interact with other components of the fibrinolytic and coagulation systems?

Understanding SERPINF2's interactions within the broader hemostatic system provides crucial mechanistic insights:

Plasminogen Activation System:

• SERPINF2 is the primary physiological inhibitor of plasmin

• Plasmin is generated from plasminogen by tissue-type (tPA) or urokinase-type (uPA) plasminogen activators

• In SERPINF2-deficient mice, enhanced spontaneous clot lysis occurs due to uninhibited plasmin activity

Interaction with Other Serpins:

• The fibrinolytic system contains multiple regulatory serpins

• Plasminogen activator inhibitor-1 (PAI-1/SERPINE1) inhibits tPA and uPA

• Functional redundancy with other serpins may explain the relatively mild phenotype in knockout mice

Methodological Approaches to Study These Interactions:

• Co-immunoprecipitation to detect physical interactions between SERPINF2 and other proteins

• Plasma reconstitution experiments comparing clotting/lysis with components added to SERPINF2-deficient plasma

• Multi-parameter coagulation testing including:

  • Thromboelastography (TEG)

  • Specialized fibrinolysis assays

  • Global coagulation assays

• Comparative studies between single and double knockout models

The enhanced spontaneous clot lysis in SERPINF2-deficient mice without significant bleeding time alterations suggests complex compensatory mechanisms within the hemostatic system . Understanding these interactions could reveal new therapeutic approaches for thrombotic or hemorrhagic disorders.