Profilin Antibody
Description
Profilin Antibody Overview
Profilin antibodies are immunoglobulins raised against epitopes of Profilin proteins. They are categorized by their specificity to isoforms (e.g., Profilin-I, Profilin-II) and applications such as Western blot (WB), immunoprecipitation (IP), or immunofluorescence (IF). Recent studies highlight the importance of rigorous validation to ensure antibody specificity, particularly in research on neurodegenerative diseases like amyotrophic lateral sclerosis (ALS), where PFN1 mutations have been identified .
Profilin-1 Antibody Characterization
A 2023 study systematically evaluated 16 commercial Profilin-1 antibodies across multiple experimental platforms . Key findings include:
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Western Blot:
Antibodies were tested for specificity using knockout (KO) and wild-type (WT) cell lysates. High-performing antibodies (e.g., Abcam ab51263, Santa Cruz sc-365703) exhibited strong signal in WT lysates and no cross-reactivity in KO lysates. -
Immunoprecipitation:
Antibodies like CST #9167 demonstrated efficient pull-down of Profilin-1 from cell extracts, confirming their utility in biochemical assays. -
Immunofluorescence:
A mosaic staining strategy (WT and KO cells co-cultured) revealed that antibodies such as Sigma-Aldrich HPA026793 achieved robust, specific staining of Profilin-1 in WT cells.
Table 1: Performance of Top-Performing Profilin-1 Antibodies
| Antibody Vendor/Catalog Number | Western Blot | Immunoprecipitation | Immunofluorescence |
|---|---|---|---|
| Abcam ab51263 | ✅ | ✅ | ✅ |
| Santa Cruz sc-365703 | ✅ | ✅ | ✅ |
| Sigma-Aldrich HPA026793 | ✅ | ✅ | ✅ |
| CST #9167 | ✅ | ✅ | ✅ |
Applications in Research
Profilin antibodies are integral to studying actin dynamics and disease mechanisms:
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Neurodegeneration: PFN1 mutations linked to familial ALS necessitate antibodies for mechanistic studies. For example, mutant PFN1 (G118V) in mice shows motor defects, and antibodies enable detection of protein aggregates .
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Cancer: Profilin’s role in cell migration makes antibodies valuable for investigating metastasis pathways .
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Pollen Allergies: Profilin is a major allergen in grass and birch pollen, and antibodies assist in diagnostics and therapeutic development .
Best Practices for Antibody Selection
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Use Knockout Controls: Validate specificity by comparing signals in WT and KO cells.
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Optimize Conditions: Adjust antibody dilutions and blocking buffers to minimize background noise.
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Cross-Platform Testing: Ensure antibodies are compatible with multiple techniques (e.g., WB, IF) for comprehensive analysis.
Future Directions
References Wikipedia contributors. (2023). Profilin. Wikipedia. Retrieved from https://en.wikipedia.org/wiki/Profilin. Ayoubi et al. (2023). The identification of high-performing antibodies for Profilin-1. PubMed. Retrieved from https://pubmed.ncbi.nlm.nih.gov/37576538/. Ayoubi et al. (2023). The identification of high-performing antibodies for Profilin-1. PMC. Retrieved from https://pmc.ncbi.nlm.nih.gov/articles/PMC10415725/.
Product Specs
**Constituents:** 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
UniGene: Ppe.19069
Q&A
What are the primary applications of Profilin-1 antibodies in cellular biology studies?
Profilin-1 antibodies are essential for studying actin dynamics, cytoskeletal remodeling, and cellular processes like motility and division. Common applications include:
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Western blotting: Quantifying Profilin-1 expression levels in cell lysates or tissue samples .
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Immunoprecipitation: Isolating Profilin-1 complexes to study protein interactions (e.g., with actin-binding partners like dynamin I or synapsin) .
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Immunofluorescence: Visualizing Profilin-1 localization in axonal/dendritic processes or subcellular compartments .
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ELISA: Detecting secreted Profilin-1 in extracellular matrices or biofluids (e.g., urine in bladder cancer studies) .
Methodological Note: Antibody performance varies by technique. For example, conjugated antibodies (e.g., HRP, Alexa Fluor®) may optimize signal-to-noise ratios in Western blotting or imaging .
How do I select a Profilin-1 antibody for cross-species studies?
Profilin-1 antibodies must be validated for cross-reactivity with target species. Key considerations:
Critical Tip: Profilin-1 and Profilin-2 are encoded by distinct genes. Use isoform-specific antibodies to avoid cross-reactivity .
What controls should I include in Profilin-1 antibody validation?
Validating antibodies requires rigorous controls to ensure specificity:
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Knockout (KO) cells: Compare signal intensity between wild-type (WT) and PFN1 KO cells to confirm antibody specificity .
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Isogenic parental controls: Use HAP1 WT vs. PFN1 KO cells for Western blot or immunofluorescence .
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Negative controls: Omit primary antibody or use non-specific IgG to assess background noise .
Data Table: Antibody Performance in Validation Studies
| Antibody Source | Western Blot | Immunoprecipitation | Immunofluorescence |
|---|---|---|---|
| SCBT (B-10) | High | Moderate | Moderate |
| Horizon (Rabbit) | Moderate | High | High |
| Alexis (Polyclonal) | Low | Low | Moderate |
How do I resolve conflicting data between Profilin-1 antibodies in neurodegenerative disease models?
Conflicting results often arise from antibody cross-reactivity or epitope masking. Strategies to address this:
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Epitope mapping: Use antibodies targeting distinct regions of Profilin-1 (e.g., N-terminal vs. C-terminal domains) .
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KO cell validation: Confirm antibody specificity by testing in PFN1 KO cells .
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Quantitative analysis: Compare signal intensity ratios (WT/KO) across antibodies to identify outliers .
Case Study: In ALS models, PFN1 G118V mutant mice show motor defects. Antibodies with high specificity in WT vs. KO cells are critical for studying pathogenic mechanisms .
What experimental designs optimize Profilin-1 antibody use in actin polymerization assays?
To study actin dynamics, pair Profilin-1 antibodies with functional assays:
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TIRFM (Total Internal Reflection Fluorescence Microscopy): Use antibodies to visualize actin filament elongation at physiological profilin/actin ratios .
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Nucleotide exchange assays: Measure ATP charging of actin monomers in the presence of Profilin-1 .
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Cell motility assays: Knock down Profilin-1 and assess rescue with antibody-neutralized Profilin-1 to study motility defects .
Key Parameter: Maintain near-physiological ionic strength (0.133 M) to preserve actin/profilin binding kinetics .
How can Profilin-1 antibodies be leveraged in biomarker discovery for bladder cancer?
Profilin-1 is a potential biomarker for bladder cancer aggressiveness. Experimental workflows:
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Urinary Profilin-1 detection: Use ELISA kits with validated antibodies to quantify secreted Profilin-1 .
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Immunohistochemistry (IHC): Assess stromal vs. epithelial expression patterns using rabbit polyclonal antibodies .
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Functional validation: Block Profilin-1 with neutralizing antibodies to test effects on cell motility and actin polymerization .
Clinical Relevance: High stromal Profilin-1 expression correlates with poor prognosis. Antibody-based detection in urine may enable non-invasive monitoring .
What challenges arise when using Profilin-1 antibodies in plant allergen studies?
Plant-derived Profilin is a pan-allergen. Challenges include:
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Cross-reactivity: Antibodies may bind conserved epitopes in human and plant Profilin-1 .
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Sample preparation: Denatured Profilin (e.g., in cooked foods) may alter epitope accessibility .
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ELISA optimization: Use affinity-purified antibodies to reduce non-specific binding to food matrices .
Solution: Validate antibodies against recombinant plant Profilin-1 (e.g., celery, peanut) to confirm specificity .
Why does my Profilin-1 Western blot show non-specific bands?
Common causes and fixes:
| Issue | Solution |
|---|---|
| Cross-reactivity | Test in PFN1 KO cells |
| Insufficient blocking | Increase BSA/goat serum concentration |
| Poor antigen retrieval | Optimize PFA fixation time (15 min) |
Advanced Tip: Use Alexa Fluor®-conjugated secondary antibodies for immunofluorescence to minimize background .
How can I improve signal-to-noise in Profilin-1 immunofluorescence?
Optimize staining protocols:
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Fixation: Use 4% PFA for 15 min to preserve cytoskeletal structures .
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Permeabilization: Triton X-100 (0.1%) for membrane disruption without excessive actin depolymerization .
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Blocking: PBS + 5% BSA + 5% goat serum to reduce non-specific binding .
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Primary antibody incubation: Overnight at 4°C to enhance epitope accessibility .
Quantification: Use DAPI for nuclear counterstaining and image analysis software to normalize fluorescence intensity .
Can Profilin-1 antibodies advance studies of synaptic plasticity?
Yes. Profilin-1 co-localizes with dynamin I and synapsin in axons/dendrites. Applications:
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Live-cell imaging: Track Profilin-1 dynamics during synaptic vesicle recycling .
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CRISPR editing: Use antibodies to validate PFN1 knockout in neuronal models .
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Protein interaction mapping: Immunoprecipitation to identify Profilin-1 partners in synaptic terminals .
Key Insight: Profilin-1 overexpression enhances cell adhesion to fibronectin, suggesting roles in synaptic adhesion .
How do I integrate Profilin-1 antibody data with proteomic/network analysis?
Combine antibody-based workflows with omics data:
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Proteomics: Use mass spectrometry to identify Profilin-1 binding partners .
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Bioinformatics: Map Profilin-1 to actin-regulatory pathways (e.g., WASP, VASP) .
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Single-cell RNA-seq: Correlate PFN1 transcript levels with cytoskeletal gene modules .
Example: In bladder cancer, link urinary Profilin-1 levels to stromal-epithelial crosstalk networks .
