DMD Antibody, Biotin conjugated
Description
Diagnostic and Therapeutic Utility
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Muscular Dystrophy Diagnosis: Biotin-conjugated DMD antibodies are used in IHC to detect dystrophin expression in muscle biopsies. For example, BosterBio’s anti-dystrophin antibody (PB9276) demonstrated robust staining in human, mouse, and rat tissues, with specificity confirmed in Duchenne patient samples .
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Therapeutic Development: In mdx mice (a DMD model), anti-myostatin antibodies like mRK35 improved muscle mass and strength, highlighting the potential of antibody-based therapies for DMD . While not directly using biotin-DMD conjugates, this underscores the role of targeted antibody delivery in neuromuscular diseases.
Mechanistic Insights
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Biotin-Streptavidin Systems: Biotinylated antibodies enable modular detection; streptavidin-enzyme/fluorophore conjugates amplify signals in low-abundance targets. For example, Bio-Techne’s biotin-DMD antibodies (NBP2-79917B, NBP3-20751B) are validated for WB, IHC, and protein arrays .
Key Considerations
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SMVT Transport: Free biotin’s carboxylic acid group is essential for uptake via the sodium-dependent multivitamin transporter (SMVT). Biotin-drug conjugates may bypass SMVT, relying instead on receptor-mediated endocytosis .
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Stability: Biotin-avidin binding ( K d ≈ 10⁻¹⁵ M) ensures durable complexes, critical for high-sensitivity assays .
Future Directions in DMD Antibody Research
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Multiplexed Assays: Combining biotin-DMD antibodies with other tagged probes (e.g., fluorophores) could enable simultaneous detection of dystrophin isoforms.
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Gene Therapy Integration: Adeno-associated virus (AAV) vectors delivering DMD transgenes paired with biotinylated antibodies may enhance dystrophin tracking in vivo .
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- Case Report: A dystrophin mutation (p.1667 del Ala) was identified in a patient exhibiting Becker muscular dystrophy-associated cardiomyopathy. The observed pathology included substantial fibrofatty replacement in the sub-epicardial layer of the ventricle. PMID: 30103083
- DMD gene mutations affecting the hinge 3 region, actin-binding domain, and exons 45-49, alongside the LTBP4 IAAM haplotype, were not found to be correlated with the age of left ventricular dysfunction onset in Duchenne muscular dystrophy. PMID: 29766838
- A novel 9358-9359insA mutation within the dystrophin gene was discovered in a Chinese boy with muscular dystrophy and his mother. PMID: 29336709
- An analysis of the mutation spectrum within the DMD gene in 68 Kuwaiti families diagnosed with Duchenne/Becker muscular dystrophy was reported. PMID: 29847600
- Lowered dystrophin expression has been associated with both Becker and Duchenne muscular dystrophy. PMID: 29641567
- This research characterized the developmental profile of dystrophin expression across human brain regions to define the temporal profile of astrocytic endfoot development. PMID: 28509351
- A novel small mutation at the first exon-intron boundary splicing site of the DMD gene was identified in a patient exhibiting elevated serum CK levels within their family. This mutation was linked to X-linked dilated cardiomyopathy. PMID: 29901616
- Newly discovered mutations in Duchenne muscular dystrophy/Becker muscular dystrophy patients carrying deletions were found to be significantly more frequent than those with duplications and small mutations. PMID: 28680110
- Ambulant and steroid-naive Japanese Duchenne muscular dystrophy patients were observed to be significantly shorter than individuals with normal stature. Additionally, Becker muscular dystrophy patients were slightly shorter than normal. This suggested a correlation between dystrophin expression levels and short stature. The higher incidence of short stature in the Dp71 subgroup points to a potential role of Dp71 in growth. PMID: 28734761
- Data indicate that Becker muscular dystrophy (BMD) patients carry deletions within the rod domain of the exons in the dystrophin gene. PMID: 29419852
- It has been concluded that patients with dystrophin "del x-51" or "del 48" mutations experience mild or asymptomatic Becker muscular dystrophy, whereas "del 45-x" mutations cause comparatively severe weakness and functional deterioration within a year. PMID: 27582364
- Further investigation and validation of DMD as a potential player in tumor development and its role as a prognostic factor for tumor progression and survival are warranted. PMID: 27391342
- The entire DMD locus undergoes dynamic transcription by RNA polymerase II, highlighting mechanisms involved in the regulation of dystrophin gene expression. PMID: 28867298
- This study suggests that the Dp71-lamin B1 protein complex plays a significant role in the newly identified tumor suppressive function of Dp71. PMID: 27449096
- By increasing FAK in the cytoplasmic FAK-Dp71 complex, lamin B1 in the nuclear laminB1-Dp71 complex, and HBE-Dp71d and HBE-Dp71f cells, alterations in proliferation, migration, invasion, cell cycle, and apoptosis rate induced by H2O2 were observed. PMID: 29059680
- MyoD-transformed cells may serve as a suitable myogenic model for studying dystrophin gene expression. Native urine stem cells could be utilized to investigate the dystrophin transcript, potentially contributing to both diagnostic procedures and splicing modulation therapies in both patients and control subjects, without the need for invasive and costly collection methods. PMID: 27530229
- The de novo deletion of exons 17 to 29 of the DMD gene detected in a fetus may result in either BMD or DMD. PMID: 28777860
- Recent findings demonstrate that multi-step events are required for the removal of long DMD introns. The role of temporary intron retention in the occurrence of alternative splicing events is also discussed. PMID: 28597072
- This study provides evidence that dystrophin contains multiple independent membrane-binding domains. These structurally and functionally distinct membrane-binding domains form a molecular framework enabling dystrophin to act as a shock absorber and signaling hub. PMID: 27378693
- Multiplex ligation-dependent probe amplification combined with next-generation sequencing proved effective in detecting mutations in dystrophin gene exons in patients with Duchenne and Becker muscular dystrophies. PMID: 27750387
- A rarely reported deletion of a single exon 48 of the dystrophin gene caused a subclinical or very mild form of Becker muscular dystrophy in nine non-consanguineous families. PMID: 28247318
- This study describes a series of patients of Greek origin carrying a c.5068_5070delCAC mutation in the DMD gene. PMID: 27178005
- These data support the hypothesis that skewed XCI is involved in the onset of phenotype in DMD carriers, where the X chromosome carrying the normal DMD gene is preferentially inactivated, leading to moderate-severe muscle involvement. PMID: 27098336
- DMD reversion leads to somatic mosaicism in DMD patients. PMID: 26956251
- This study focused on the phenotype of BMD patients with in-frame deletions starting at exon 45 of the DMD gene to investigate the appropriate restoration of the reading frame by exon skipping therapy. PMID: 27974813
- It was concluded that Dp71, comprising Dp71b and Dp71ab, was exclusively expressed in HEK293 cells and that Dp71ab was specifically localized to the nucleus. These findings suggest that Dp71ab in the nucleus contributes to the diverse functions of HEK293 cells. PMID: 27109495
- This research aimed to provide in vitro quantitative evidence of the ability of human mesoangioblasts to restore dystrophin, in terms of protein accumulation and distribution, within myotubes derived from patients with Duchenne muscular dystrophy. PMID: 27502519
- In TMD patients, a novel locus at genome-wide level of significance (rs73460075, OR = 0.56, P = 3.8 x 10(-8)) in the intron of the dystrophin gene DMD (X chromosome), and a suggestive locus on chromosome 7 (rs73271865, P = 2.9 x 10(-7)) upstream of the Sp4 Transcription Factor ( SP4) gene were identified in the discovery cohort, but neither of these was replicated. PMID: 28081371
- As exon 44 skipping-amenable DMD exhibits a later loss of ambulation, mutation-specific randomization and selection of placebo groups are crucial for the success of clinical trials. PMID: 27343068
- This study focused on four prevalent mutated proteins deleted in RDelta45-47, RDelta45-48, RDelta45-49, and RDelta45-51, analyzing protein/membrane interactions. Mutants RDelta45-48 and RDelta45-51 led to mild pathologies and displayed a similar triple coiled-coil structure as the full-length DYS R16-21, whereas the two others, RDelta45-47 and RDelta45-49, induced more severe pathologies and showed "fractional" structures unre... PMID: 27367833
- The dystrophin expression plasmids described here will be valuable in cell and gene therapy studies aimed at mitigating Duchenne muscular dystrophy. PMID: 28139886
- In Korean boys, 117 different deletions, 48 duplications, and 90 pathogenic sequence variations, including 30 novel variations, were identified. Deletions and duplications accounted for 65.4% and 13.3% of Korean dystrophinopathy, respectively, suggesting that the incidence of large rearrangements in dystrophin is similar among different ethnic groups. PMID: 27593222
- The expression of endogenous exons 44-56 connected mRNA transcript of the DMD was detected using total RNAs derived from human normal skeletal muscle by reverse transcription polymerase chain reaction. A total of eight types of multiple exon skipping products around the mutation hotspot were identified. PMID: 27754374
- Multiplex Ligation Probe Amplification identified 56 mutations (45 deletions, 9 duplications and 2 point mutations), confirming the clinical diagnosis in 63% (51/81) of patients and symptomatic females. It also established the carrier status of 54% (20/37) of females at-risk and 3 male villus samples. An association between the most frequent deletion intron breakpoints and the abundance of dinucleotide microsatellites was established. PMID: 27206868
- Using NGS, we have identified a pathogenic DMD mutation from degraded DNA and low-level somatic mosaicism, which would have been overlooked using Sanger sequencing. PMID: 26740235
- Four nonsense, one frameshift, and two splice site mutations, as well as two missense variants, were found in the dystrophin gene in Iranian Duchenne and Becker muscular dystrophy patients. PMID: 27350676
- Four missense mutations (p.Arg2937Gln, p.Asp882Gly, p.Lys2366Gln, and p.Arg1745His) that are known multiple-polymorphic sites were found in the coding region of the DMD gene. A hemizygous splicing mutation IVS44ds +1G-A (c.6438 +1G>A) was located in intron 44. PMID: 27421007
- Results provide evidence for recursive splicing in the dystrophin transcript and that the order at which introns are removed is not consecutive. PMID: 26670121
- DMD mutations affecting different DMD isoforms are associated with characteristically abnormal scotopic ERGs and severe neurodevelopmental problems in Duchenne muscular dystrophy patients. PMID: 26081639
- DMD gene mutations may be suspected in girls with persistently elevated levels of creatine kinase and scoliosis, calf hypertrophy, or myopathic pattern at electromyography. PMID: 26718981
- The deletion patterns and distribution characteristics of the dystrophin gene in a Chinese population of patients with Duchenne muscular dystrophy (DMD) or Becker muscular dystrophy are reported. PMID: 26786758
- The range of phenotypes associated with the region Xp21 has been expanding. The mild end of the spectrum includes phenotypes such as muscle cramps with myoglobinuria and isolated quadriceps myopathy, while the severe end encompasses progressive muscle diseases. PMID: 25416089
- Attention should be paid to the possibility of severe arrhythmias in patients with the severe phenotype of Becker muscular dystrophy. PMID: 26631896
- Males with mutations at the 3' end of the DMD gene affecting all protein isoforms have higher rates of intellectual disability and clusters of neurodevelopmental, emotional, and behavioral symptoms in Duchenne muscular dystrophy. PMID: 26365034
- This study determined the frequency of dystrophin gene alterations in Iranian Duchenne and Becker muscular dystrophies patients using two newly proposed sets of primer pairs of M-PCR to facilitate further mutation detection in patients and their families. PMID: 26081009
- Correlation of Utrophin Levels with the Dystrophin Protein Complex and Muscle Fibre Regeneration in Duchenne and Becker Muscular Dystrophy Muscle Biopsies. PMID: 26974331
- Mutations of Dystrophin and Duchenne and Becker muscular dystrophies (review). PMID: 26295289
- Data show that viral vector-mediated transient designer nuclease expression leads to permanent and regulated dystrophin synthesis from corrected native Duchenne muscular dystrophy (DMD) alleles. PMID: 26762977
- MicroRNAs contribute to variable dystrophin levels in muscular dystrophy. PMID: 26321630
- Knocking down Dp71 expression can significantly inhibit the A549 xenograft tumor growth in nude mice. PMID: 26691328
Q&A
Key Methodological Considerations for Experimental Design
Biotin-conjugated DMD antibodies enable precise detection of dystrophin in complex biological systems through streptavidin-mediated signal amplification. This FAQ synthesizes technical insights from peer-reviewed studies , commercial validation data , and detection system optimizations to address critical challenges in muscular dystrophy research.
What experimental workflows utilize biotin-conjugated DMD antibodies most effectively?
Biotin-conjugated DMD antibodies are foundational in:
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Immunohistochemistry (IHC): Paraffin-embedded human skeletal muscle sections require antigen retrieval with cell conditioning solution (CC1, pH 8.5) at 100°C for 32 min, followed by antibody incubation at 1:100–1:800 dilution . Co-staining with spectrin (1:20 dilution) enables membrane integrity assessment in dystrophinopathies .
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Proximity labeling: The Biotinylation by Antibody Recognition (BAR) method uses primary antibodies against DMD and HRP-conjugated secondaries to biotinylate adjacent proteins in fixed tissues, enabling interactome studies via mass spectrometry .
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ELISA quantification: Recombinant human dystrophin (3400–3558AA) serves as the immunogen for polyclonal antibodies, with biotin conjugation enhancing sensitivity in capture assays .
Table 1: Antibody Performance Across Assays
How do researchers validate antibody specificity in dystrophin-deficient models?
Three orthogonal strategies are critical:
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Genetic controls: Compare staining in mdx (dystrophin-null) iPSCs vs. gene-corrected isogenic lines using anti-spectrin as a normalization marker .
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Blocking peptides: Pre-incubate antibodies with recombinant dystrophin (3400–3558AA) to confirm signal loss in wild-type muscle .
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Cross-species reactivity: Validate in murine models by testing antibody binding to both human (UniProt P11532) and mouse dystrophin isoforms .
How can proximity labeling with biotinylated DMD antibodies resolve dynamic interactomes in muscular dystrophy?
The BAR method addresses this through:
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Fixation-permeabilization: 4% PFA fixation preserves native protein interactions while allowing antibody penetration.
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Controlled radical generation: HRP-conjugated secondaries + H₂O₂/phenol biotin deposit biotin within a 20 nm radius of the antigen.
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Differential proteomics: SILAC labeling distinguishes true interactors (heavy isotope-labeled) from background proteins in lamin A/C interactome studies .
Table 2: BAR vs. Traditional Immunoprecipitation
What explains contradictory dystrophin localization data across studies using biotinylated antibodies?
Discrepancies often arise from:
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Epitope accessibility: Antibodies targeting residues 3400–3558AA may miss truncated isoforms in Becker muscular dystrophy vs. full-length deletions in Duchenne .
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Fixation artifacts: Over-fixation in FFPE masks epitopes detectable in frozen sections .
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Signal amplification bias: The ABC method generates larger complexes (≤500 kDa) than LSAB, limiting penetration into dense muscle matrices .
Resolution protocol:
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Parallel detection: Compare ABC (avidin-biotin complex) and LSAB (labeled streptavidin biotin) in the same sample .
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Multiplex validation: Use RNAscope® to correlate dystrophin mRNA distribution with protein localization patterns .
How to optimize biotin-streptavidin systems for low-abundance dystrophin detection?
Tandem amplification achieves 8–12x sensitivity gains :
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Primary amplification: Biotinylated anti-DMD (1:200) + streptavidin-APC (1:1000).
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Secondary amplification: Anti-APC-biotin (1:500) + streptavidin-HRP.
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Threshold calibration: Use mdx muscle spiked with 0.01–1% wild-type myofibers to establish detection limits .
Can biotinylated DMD antibodies enable single-myofiber analysis in heterogeneous tissues?
Emerging workflows combine:
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Laser capture microdissection: Isolate dystrophin+ vs. dystrophin– fibers from IHC-stained sections.
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SWATH-MS: Quantify 40+ DGC proteins per fiber using the NBP3-20751B antibody .
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Data normalization: Express dystrophin levels as a ratio to β-spectrin to account for section thickness variability .
What computational tools reconcile interactome data from biotinylation-based studies?
The 2024 BAR-DMD pipeline integrates:
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Proximity score: Weight proteins by biotinylation frequency across 5 replicates.
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Contaminant filtering: Subtract proteins appearing in ≥3/6 negative controls (e.g., IgG-only BAR).
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Pathway mapping: Enrichment for ECM-receptor interaction (p=1.2e-12) and hypertrophic cardiomyopathy (p=3.4e-09) .
How to distinguish non-specific biotinylation from true signals?
Implement a three-tiered approach:
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SILAC ratios: True interactors show >3:1 heavy/light isotope ratios in BAR experiments .
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Spatial profiling: Confocal microscopy confirms perinuclear ER localization (dystrophin-sarcoglycan complexes) vs. cytoplasmic artifacts .
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Kinetic controls: Omit H₂O₂ in parallel samples to disable radical-based biotinylation .
What quality metrics ensure reproducibility in multi-center studies?
Adopt the MUSCLE-BiQ (Multisite Standardization of Conjugated Antibody Quality) framework:
