DEFA1 Antibody
Description
Research Applications
The DEFA1 antibody is validated for use in:
Protocols:
-
Antigen Retrieval: For IHC, citrate buffer (pH 6.0) or TE buffer (pH 9.0) is recommended.
-
Blocking: Non-specific binding is minimized using 1–5% BSA or normal serum.
Role in Cancer Progression
-
Tumor Invasion: DEFA1 overexpression at the invasive tumor front promotes cancer cell migration and growth via the JNK MAPK/NF-κB pathway. Monocyte-derived DEFA1 enhances this effect, suggesting a microenvironment-driven mechanism .
-
Biomarker Potential: Elevated DEFA1 levels correlate with colorectal and bladder cancer progression, with studies indicating its utility as a diagnostic marker .
Immune Response Modulation
-
COVID-19 Severity: DEFA1 levels are elevated in severe COVID-19 cases, serving as a biomarker with 68% sensitivity and 91% specificity for disease severity prediction .
-
Periodontitis: DEFA1 inhibits osteoclast differentiation and is proposed as a diagnostic target for periodontitis, with significantly higher levels in gingival crevicular fluid (GCF) of affected patients .
Mechanistic Insights
-
Antimicrobial Activity: DEFA1 disrupts microbial membranes and inhibits bacterial cell wall synthesis, contributing to innate immunity .
-
Immune Cell Activation: It induces pro-inflammatory cytokines (e.g., IL-1β, TNF-α) and modulates dendritic cell maturation .
Clinical and Diagnostic Implications
Product Specs
Target Background
- Lower DEFA1/DEFA3 copy number (CNV < 7) is associated with increased susceptibility to hospital-acquired infections (HAIs) in critically ill patients. This suggests that host genetic factors contribute to the development of HAIs. PMID: 29950924
- Human Neutrophil Peptide 1 directly binds and neutralizes Exotoxin A produced by Pseudomonas aeruginosa. PMID: 29738776
- Pretreating component b with human alpha-defensin-1 prior to adding component a of Clostridium perfringens iota toxin prevents intoxication. This implies that alpha-defensin-1 interacts with the toxin b component to inhibit the formation of biologically active iota toxin on cells. PMID: 29635426
- DEFA1, DEFA3, and PPBP expression levels are significantly elevated in hyperlipidemia and coronary heart disease patients compared to controls. PMID: 28420383
- HNP1 upregulation of cytokine expression in pDCs is inhibited by blocking NF-kappaB activation or by knocking down IRF1, highlighting the importance of these two signaling pathways in HNP1-induced pDC activation. PMID: 27031443
- These findings demonstrate that HNPs1-3 may be potent inhibitors of ADAMTS13 activity, likely by binding to the central A2 domain of VWF and physically preventing ADAMTS13 binding. PMID: 27207796
- Elevated alpha-defensin expression is associated with an increased risk of coronary heart disease in hyperlipidemia patients. PMID: 27430968
- Serum and cerebrospinal fluid levels are elevated in Alzheimer's disease patients compared to controls. PMID: 27082275
- The reduced HNP-1 production in polymorphonuclear phagocytes from elderly individuals might contribute to their increased susceptibility to infectious diseases. PMID: 26323500
- Increased levels of neutrophil defensin 1, apolipoprotein E, clusterin, and zinc-alpha-2-glycoprotein are found in carotid atherosclerotic plaque secretomes. PMID: 26795031
- HNP1 acts as a "molecular brake" on macrophage-driven inflammation, ensuring both pathogen clearance and the resolution of inflammation with minimal collateral tissue damage. PMID: 27044108
- HP1 binding to the chromosomal passenger complex becomes particularly crucial when Aurora B phosphorylates kinetochore targets to eliminate erroneous microtubule attachments. PMID: 26954544
- Data indicate that HNP-1 induces IL-8 production not only through P2Y6 but also through additional P2 receptors via an ERK1/2-dependent mechanism in intestinal epithelial cells. PMID: 25816245
- Results suggest that NO and NaNO2 can be considered as factors regulating the chemoattractant properties of defensin HNP1 in neutrophils. PMID: 25772994
- Overexpression of DEFA1 retained independent prognostic significance for B-ALL outcome. PMID: 25802479
- The expression of alpha-defensins 1, 2, and 3 is upregulated in hypercholesterolemia. PMID: 25300997
- Defensins promote the differentiation of dendritic cells into activated CD91(bright) dendritic cells. PMID: 25351513
- Elevated DEFA1 levels in diabetes are independent of DEFA1 copy numbers. PMID: 25083086
- The AD-1 assay provides another highly sensitive and specific test for diagnosing prosthetic joint infections. PMID: 25256621
- The concentrations of LL37, alpha-1, beta-1, and beta-2 defensins were determined by ELISA. Serum AMPs did not change during attacks and did not correlate with acute phase reactants. PMID: 24747281
- DEFA1 encodes an antibacterial peptide that is bactericidal against S. aureus, E. coli, and P. aeruginosa. PMID: 2997278
- Side chain hydrophobicity is the key factor determining HNP1 protein binding potential. PMID: 24236072
- A reduction in salivary HNP 1-3 levels might be a contributing factor to the predisposition to oral ulcers in patients with Behcet's disease and oral infections in healthy individuals. PMID: 22861387
- HNP-1 exacerbates hepatic fibrosis in fatty liver by stimulating hepatic stellate cell proliferation. PMID: 23682724
- Alpha-defensin 1 enhances EFS-induced contractions of rat detrusor muscles. PMID: 23542712
- Sub-inhibitory doses of HNP-1 significantly enhance the activity of several anti-gp41 antibodies and peptide inhibitors. PMID: 23785290
- Human defensin alpha-1 is an innate immune molecule secreted by HCT116 cells in response to Trypanosoma cruzi infection. It inhibits Trypanosoma cruzi motility and plays a significant role in reducing cellular infection. PMID: 23980110
- Results show that DEFA 1/3 and 4 gene expression is significantly increased in all tumors, except for a significant decrease in DEFA 4 gene expression in pleomorphic adenomas. PMID: 23050799
- This study suggests that HNPs 1-3 promote tumor invasion and are potential indicators of disease progression in patients with bladder cancer. PMID: 23011762
- The invariant glycine residue is essential for alpha-defensin folding and dimerization. PMID: 22496447
- Alpha-Defensin (DEFA1) was the most differentially overexpressed gene in this sample and may be related to the onset of Bell's palsy and Ramsay Hunt Syndrome. PMID: 22737966
- Expression levels of HNPs 1, 2, and 3 were significantly higher and their distributions overlapped in cancerous tissues of gastric cancer patients. PMID: 22297599
- Dimerization contributes to some, but not all, of the activities of HNP1. PMID: 22270360
- A high DEFA1 gene copy number was significantly associated with intestinal involvement in BD patients. PMID: 22219625
- Directly isolated dendritic cells secrete alpha-defensins 1-3. E2 inhibits this secretion. The same trend is observed in myeloid dendritic cells isolated from pregnant women in their first trimester (low plasma E2) and third trimester (high plasma E2). PMID: 21861873
- Elevated levels of human neutrophil peptides 1, 2, and 3 in peritoneal fluid of patients with endometriosis: association with neutrophils, T cells, and IL-8. PMID: 21831449
- Human granulocyte precursors transfected with siRNA against serglycin showed reduced capability to retain fully processed HNP-1. PMID: 21849484
- HNP1-3 concentrations in patients with multidrug-resistant tuberculosis were significantly lower than in drug-susceptible pulmonary TB and healthy controls. PMID: 21333105
- Changes in the expression pattern of DEFA 1/3 appear to be involved in the development of gingival irritation fibromas, while chronic inflammation might be less important. PMID: 21187770
- Alpha-defensins 1-3 in T cells from patients with SJS/TEN may play a role in the pathogenesis of these life-threatening medication-induced diseases. PMID: 20880148
- Defensin-alpha(1) is a biologically active peptide exhibiting a dose-dependent trypanocidal effect in vitro against trypomastigotes and amastigotes of Trypanosoma cruzi line Tulahuen. PMID: 21165432
- HNP-1 (alpha), HBD-2 (beta), and RTD-1 (theta) exhibit anti-HIV-1 activity, but through different mechanisms. PMID: 20305815
- HNP-1 mainly enhanced the expression of IL-8 in epithelial cells, while it enhanced transforming growth factor-beta and vascular endothelial growth factor expressions in lung fibroblasts. PMID: 20715983
- Our report of DEFA1-3 expression by human omental adipocytes adds to the understanding of adipocytes' role in primary defense against bacterial infection. PMID: 20424487
- Data suggest that high production of alpha-defensins1-3 by immature DCs acts as a host protective factor against the progression of HIV-1 infection, implying potential diagnostic, therapeutic, and preventive implications. PMID: 20195543
- The total amount of gingival crevicular fluid human neutrophil peptides 1 and 2 did not differ among periodontitis, gingivitis, and health control groups. There was no correlation with clinical periodontal parameters. PMID: 20151808
- HNP1 binds to the cell wall precursor lipid II, and reducing lipid II levels in the bacterial membrane significantly reduces bacterial killing. PMID: 20214904
- Combining distance constraints from the 3D experiment and the chemical-shift-derived torsion angles, we obtained a de novo high-resolution NMR structure of HNP-1. PMID: 19963419
- HNP1 mediates host immune responses to tumors in situ through the recruitment and subsequent activation of immature dendritic cells. PMID: 19861439
- The solid-state NMR structure of HNP-1 exhibits close similarity to the crystal structures of the HNP family, with the exception of the loop region between the first and second beta-strands. PMID: 20097206
Q&A
How do researchers validate the specificity of DEFA1 antibodies in Western blotting?
Specificity validation requires a multi-step approach:
-
Knockout controls: Use lysates from DEFA1-knockout cell lines or tissues to confirm absence of non-specific bands .
-
Peptide competition assays: Pre-incubate the antibody with excess immunizing peptide (e.g., synthetic human DEFA1 peptide) to block target binding .
-
Orthogonal validation: Compare results with RNA interference (siRNA-mediated DEFA1 suppression) or mass spectrometry .
Table 1: Validation Metrics for Common DEFA1 Antibodies
| Antibody Clone | Host | Applications Validated | Cross-Reactivity Tested |
|---|---|---|---|
| Polyclonal (ABIN7237958) | Rabbit | WB, ELISA | Human, Mouse |
| ab134706 | Rabbit | IHC-P, WB | Human |
| Monoclonal 1B20 | Mouse | ELISA | Human |
What sample preparation protocols optimize DEFA1 detection in immunohistochemistry?
-
Fixation: Limit formalin fixation to <24 hours to prevent epitope masking .
-
Antigen retrieval: Use citrate buffer (pH 6.0) with high-temperature (95°C) unmasking for 20–30 minutes .
-
Blocking: 5% BSA + 0.1% Tween-20 reduces non-specific binding in neutrophil-rich tissues .
Critical consideration: DEFA1’s high cationic charge necessitates low-ionic-strength buffers (e.g., 10 mM Tris-HCl) to prevent electrostatic interactions with non-target proteins .
How do ELISA kits quantify DEFA1 in complex biological fluids?
Sandwich ELISA kits (e.g., ABIN6955300) employ:
-
Capture antibody: Coated with monoclonal anti-DEFA1 (AA 20–94) to ensure epitope accessibility .
-
Detection range: 0.31–20 ng/mL, requiring sample dilution for plasma/serum (typically 1:10–1:50) .
-
Interference mitigation: Pre-treat samples with heterophilic blocking reagents to neutralize rheumatoid factors .
How should researchers resolve contradictions in DEFA1 levels between ELISA and Western blot data?
Discrepancies often arise from:
-
Post-translational modifications: DEFA1’s disulfide bonds may alter ELISA epitopes but not Western blot linearized epitopes .
-
Proteolytic degradation: Add protease inhibitors (e.g., 1 mM PMSF) during neutrophil isolation .
-
Matrix effects: Compare spike-and-recovery rates across matrices (e.g., 92% recovery in serum vs. 68% in cell lysates) .
Table 2: DEFA1 Recovery Rates in Common Matrices (ABIN6955300 Kit)
| Matrix | Spiked Concentration (ng/mL) | Recovery (%) |
|---|---|---|
| Human Serum | 5.0 | 92 ± 4.1 |
| Cell Lysate | 5.0 | 68 ± 6.3 |
| Tissue Homogenate | 5.0 | 74 ± 5.8 |
What strategies enable multiplexed detection of DEFA1 with co-expressed antimicrobial peptides?
-
Antibody pairing: Combine monoclonal DEFA1 (clone 1B20, mouse host) with rabbit polyclonals for HNP-2/3 to avoid cross-reactivity .
-
Spatial resolution: Sequential immunohistochemistry with heat-mediated antibody stripping (>95°C for 30 min between rounds) .
-
Signal amplification: Use tyramide-based systems (e.g., Opal 520/620/690) for spectral separation .
Case study: In COVID-19 cohorts, multiplex assays revealed DEFA1 levels correlated with S100A8/A9 (r=0.72, p<0.001) but not LL-37, suggesting distinct regulatory pathways .
How do longitudinal studies account for DEFA1’s dynamic expression in inflammatory models?
-
Sampling frequency: Collect samples at ≤24-hour intervals due to DEFA1’s 3.8-hour plasma half-life .
-
Normalization: Use neutrophil count-adjusted values (DEFA1 per 10^6 neutrophils) to distinguish synthesis from neutrophilia .
-
Confounder control: Monitor renal function (eGFR <60 mL/min elevates serum DEFA1 via reduced clearance) .
Key finding: In severe COVID-19, DEFA1 >314 ng/mL predicted mortality with 91% specificity (AUC=0.78) .
What controls are essential when quantifying DEFA1 in autoimmune disease models?
-
Endogenous controls: Include α-defensin-free matrices (e.g., fibroblast lysates) to identify background signals .
-
Pre-analytical variables: Standardize anticoagulants (heparin > EDTA for plasma stability) .
-
Batch effects: Calibrate inter-assay variability using lyophilized reference standards (CV <12%) .
How can researchers differentiate DEFA1 from homologous defensins in functional assays?
-
CRISPR-Cas9 knockout: Generate DEFA1-specific null lines while retaining DEFA3 expression .
-
Charge-based separation: Use cation-exchange chromatography (pH 4.5) to isolate DEFA1 (pI 8.5) from DEFA4 (pI 9.2) .
-
Functional blockade: Apply inhibitory antibodies targeting DEFA1’s lipid II binding domain (AA 28–62) .
