FER3 Antibody
Description
Introduction to FER3 Antibody
The term "FER3 Antibody" refers to antibodies targeting Fer3-like protein (FERD3L), a basic helix-loop-helix (bHLH) transcription factor involved in regulating neurogenesis, cell differentiation, and cancer biology . FERD3L binds to E-box DNA motifs and inhibits transcriptional activation by sequestering E proteins, playing roles in developmental processes and disease states such as cancer . Antibodies against FERD3L are critical tools for detecting its expression in research and clinical settings, particularly in studies of cancer biomarkers and transcriptional regulation .
Role in Cancer Biology
FERD3L is overexpressed in cancers such as colorectal and kidney carcinomas, where it promotes tumor progression by modulating transcriptional networks . Studies using FERD3L antibodies (e.g., PACO39622) have revealed:
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Biomarker Potential: Elevated FERD3L levels correlate with metastasis and poor prognosis .
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Therapeutic Target: Inhibiting FERD3L disrupts cancer cell proliferation in vitro, suggesting its utility in targeted therapies .
Neurological Functions
FERD3L is expressed in neural tissues, where it regulates neurogenesis and floor plate development . Knockdown experiments in mice demonstrate its necessity for early neuronal differentiation .
Experimental Use Cases
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Western Blot: Antibodies like ab126381 detect FERD3L at ~19 kDa in human cell lysates .
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Immunohistochemistry: PACO39622 highlights cytoplasmic and membranous FERD3L in colon cancer tissues .
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Functional Studies: Neutralizing FERD3L activity using antibodies reduces transcription of oncogenic targets like ASCL1 .
Technical Considerations
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Dilution Ranges: Optimal dilutions vary (e.g., 1:100 for IHC, 1:2000 for WB) .
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Storage: Antibodies are stable in glycerol-based buffers at -20°C .
Future Directions
Current research focuses on:
Product Specs
**Constituents:** 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
Q&A
Here’s a structured collection of FAQs tailored for researchers working with FER3 (FERDL3) antibodies, synthesized from peer-reviewed methodologies and technical data:
What experimental applications are validated for FER3 antibodies in academic research?
FER3 antibodies (e.g., ab126381) are primarily validated for:
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Western Blot (WB): Detects endogenous FERDL3 at ~19 kDa in human tissues (e.g., colon) .
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Immunohistochemistry-Paraffin (IHC-P): Localizes FERDL3 in formalin-fixed, paraffin-embedded tissues .
Methodological Considerations:
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For WB, use lysates from HEK293T cells transfected with FERDL3-myc-DDK for positive controls .
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For IHC-P, optimize antigen retrieval with citrate buffer (pH 6.0) and block with 5% BSA to reduce non-specific binding .
| Application | Validated Species | Recommended Dilution | Key Controls |
|---|---|---|---|
| WB | Human | 1:50–1:100 | Vector-only HEK293T lysate |
| IHC-P | Human | 1:50 | Isotype-matched IgG |
How to troubleshoot non-specific bands in FER3 Western blots?
Common Causes & Solutions:
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Protein Degradation: Use fresh protease inhibitors (e.g., PMSF) and confirm lysate integrity via β-actin blot.
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Cross-reactivity: Pre-adsorb antibody with FERDL3 knockout lysate.
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Buffer Optimization: Include 0.1% Tween-20 in TBST and increase blocking time to 2 hrs .
Data Validation:
Compare observed bands to the predicted 19 kDa size. Deviations may indicate splice variants or post-translational modifications.
How to design experiments to map FER3 antibody epitopes?
Stepwise Approach:
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Peptide Array: Synthesize overlapping 15-mer peptides spanning FERDL3 (aa 1–100 immunogen region) .
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ELISA: Test antibody binding to immobilized peptides.
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Alanine Scanning: Identify critical residues by substituting sequential amino acids with alanine.
Outcome: Epitope mapping clarifies specificity and guides mutagenesis studies for functional assays.
What strategies resolve contradictions in FER3 expression data across studies?
Root-Cause Analysis:
Example: Discrepancies in neuronal vs. epithelial FERDL3 levels may reflect tissue-specific isoform expression.
How to assess FER3 antibody cross-reactivity in non-human models?
Methodology:
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Phylogenetic Alignment: Compare FERDL3 sequences (e.g., mouse: 82% homology to human) .
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Functional Testing: Use siRNA knockdown in murine cell lines (e.g., 3T3-L1) and monitor off-target effects via RNA-seq.
| Species | Homology to Human FERDL3 | Observed Cross-Reactivity |
|---|---|---|
| Mouse | 82% | Predicted (untested) |
| Rat | 78% | Not recommended |
What computational tools predict FER3 antibody utility in novel assays?
Workflow:
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Structural Modeling: Use AlphaFold2 to predict FERDL3-E protein dimerization interfaces.
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Epitope Accessibility: Analyze solvent accessibility with PDBePISA.
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Machine Learning: Train classifiers on antibody performance data (e.g., Antibodypedia) to predict success in ChIP-seq.
Methodological Best Practices
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Positive Controls: Always include HEK293T lysates with overexpressed FERDL3-myc-DDK .
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Negative Controls: Use CRISPR-generated FERDL3 knockout cell lines to confirm signal specificity.
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Quantification: Normalize WB bands to housekeeping proteins (e.g., GAPDH) and report as fold-change relative to controls.
