TGF b Antibody
Description
Definition and Structure
TGF-β antibodies are typically IgG-class monoclonal antibodies designed to bind TGF-β isoforms with high specificity. They are produced via phage display or hybridoma technologies and can target single or multiple TGF-β isoforms. For example:
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Monospecific antibodies: Bind to a single isoform (e.g., TGF-β1).
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Pan-isoform antibodies: Neutralize all three isoforms (e.g., XPA.42.681) .
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Bispecific antibodies: Simultaneously target TGF-β and other molecules (e.g., PD-L1) .
These antibodies are often engineered for enhanced affinity, stability, and tumor penetration.
Mechanisms of Action
TGF-β antibodies exert their effects by:
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Blocking receptor binding: Preventing TGF-β from activating Smad-dependent signaling pathways .
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Inhibiting immune suppression: Reversing TGF-β-mediated downregulation of cytotoxic T cells and natural killer (NK) cells .
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Reducing fibrosis: Inhibiting TGF-β-driven extracellular matrix deposition .
Key Signaling Pathway Inhibition
TGF-β antibodies disrupt the canonical Smad2/3 phosphorylation cascade, which regulates genes involved in cell proliferation and apoptosis. For example, anti-TGF-β antibodies suppress Smad2 phosphorylation in Detroit 562 tumor cells .
Table 1: TGF-β Antibody Applications in Disease Models
Cancer Immunotherapy
Bispecific antibodies targeting TGF-β and PD-L1 (e.g., BiTP, YM101) synergistically enhance tumor immunity:
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BiTP: Blocks TGF-β and PD-L1, promoting T-cell infiltration and reducing immunosuppression in triple-negative breast cancer (TNBC) models .
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M7824: Combines PD-L1 inhibition with TGF-β trapping, showing efficacy in phase I trials but mixed results in phase III .
Fibrosis and Tissue Repair
Anti-TGF-β antibodies (e.g., XPA.42.681) reduce fibrotic scarring in skin, lung, and liver models by inhibiting TGF-β1/2/3 signaling .
Affinity and Neutralization Potency
Table 2: Affinity of Pan-isoform TGF-β Antibodies
| Antibody | K<sub>D</sub> (pM) for TGF-β1 | K<sub>D</sub> (pM) for TGF-β2 | K<sub>D</sub> (pM) for TGF-β3 |
|---|---|---|---|
| XPA.42.681 | ≤10 | ≤10 | ≤10 |
| XPA.42.089 | ≤10 | 25 | 1,400 |
| XPA.42.068 | 59 | 51 | 455 |
Data from affinity-matured clones in Detroit 562 xenograft models .
In Vitro Bioactivity
Anti-TGF-β antibodies inhibit:
In Vivo Efficacy
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4T1 metastatic breast cancer: 1D11 antibody reduced metastasis by enhancing CD8+ T-cell responses .
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Glioblastoma: TGF-β2 antisense DNA therapy improved immune recognition of tumor cells .
Challenges and Limitations
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Clinical Translation: Despite preclinical success, TGF-β inhibitors (e.g., M7824) have shown limited efficacy in phase III trials, highlighting the need for biomarkers to identify responsive patients .
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Toxicity: While preclinical studies report low toxicity , human trials must carefully monitor off-target effects.
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Tumor Microenvironment Heterogeneity: TGF-β-driven immune exclusion varies across tumor types, necessitating combination therapies (e.g., STING agonists) .
Future Directions
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Bispecific Antibody Development: Next-generation BsAbs (e.g., BiTP) aim to improve tumor specificity and reduce systemic toxicity .
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Pan-isoform Inhibitors: Antibodies like XPA.42.681, which neutralize all TGF-β isoforms, may address redundancy in signaling pathways .
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Combination Therapies: Co-targeting TGF-β with checkpoint inhibitors, cytokines, or STING agonists to overcome resistance .
Product Specs
Q&A
How does antibody selection impact detection of TGF-β isoforms in Western blot versus immunohistochemistry?
The critical methodological distinction lies in epitope accessibility and post-translational processing. For Western blot, antibodies targeting linear epitopes in denatured TGF-β1 (e.g., Proteintech 26155-1-AP ) require validation against recombinant proteins across species – murine studies show 97% cross-reactivity is insufficient for quantitative comparisons between human xenografts and host stroma . In contrast, IHC applications demand antibodies recognizing conformational epitopes in paraffin-embedded tissues, with successful protocols requiring antigen retrieval using citrate buffer (pH 6.0) and 3 µg/mL primary antibody concentration . A comparative analysis reveals:
What validation framework ensures antibody specificity in flow cytometry applications?
Three-tiered validation is essential:
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Biological controls: Compare staining intensity in wild-type versus TGF-β1 knockout lymphocytes, noting that complete knockout induces 4.2-fold increase in IL-9 expression during Th1/Th2 polarization .
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Competition assays: Pre-incubate antibody with 10-fold molar excess of recombinant TGF-β1 for 1 hour at 37°C – effective neutralization should reduce signal by ≥90% in intracellular staining protocols .
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Cross-platform correlation: Validate flow cytometry results against MLEC luciferase bioassays, which detect functional TGF-β activity with 25-100 pg/mL sensitivity .
How can researchers distinguish antibody-mediated detection of latent vs. active TGF-β in ELISA?
The acid activation step creates critical methodological divergence:
Active TGF-β Protocol
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Directly plate serum/tissue lysate in MEM/BSA
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Use untreated MLEC reporter cells
Total TGF-β Protocol
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Acidify samples with 1N HCl (pH 2.0) for 15 min
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Neutralize with 1.2N NaOH/0.5M HEPES
Discrepancies >30% between active/total measurements indicate improper latent complex dissociation – a common artifact in tumor lysates with high α2-macroglobulin content.
What experimental design parameters optimize in vivo therapeutic studies with anti-TGF-β antibodies?
The 4T1 metastatic breast cancer model demonstrates three critical considerations:
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Metastasis induction method
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Dosing regimen
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Endpoint selection
Primary Endpoint Detection Method Pitfall Macroscopic metastases India ink staining Misses micrometastases <0.5 mm Immune infiltration Multiplex IHC (CD8+/Granzyme B+) Regional heterogeneity requires 5-section sampling
How should researchers resolve contradictory findings in TGF-β antibody-mediated immune effects?
A 2025 analysis of 47 studies identified three major conflict points and resolution strategies:
Contradiction 1: Pro-metastatic vs anti-metastatic effects in PDAC models
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Resolution: Preclinical models using antibodies targeting TGF-β1 versus pan-TGF-β show opposing outcomes. Selective β1 inhibition (e.g., Bio-Rad AHP1734 ) increases CD8+ infiltration by 40%, while pan-inhibition dysregulates NK cell homing .
Contradiction 2: Dual pro-/anti-angiogenic reports
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Resolution: Dose-dependent VEGF modulation – 1D11 at 5 mg/kg reduces microvessel density by 35%, while 20 mg/kg induces compensatory FGF2 upregulation .
Methodological Solution Matrix
| Conflict Type | Diagnostic Assay | Recommended Approach |
|---|---|---|
| Immune cell paradox | Phospho-Smad2/3 IHC | Combine with CXCR4 inhibition |
| Metabolic adaptation | Seahorse XF analysis | Pair with 2-DG pretreatment |
What integrative strategies combine phospho-specific antibody arrays with functional genomics?
The TGF-β Phospho Antibody Array generates 176 phosphorylation profiles per sample, requiring computational integration with RNA-seq data:
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Data alignment
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Map phospho-sites to KEGG pathways (e.g., Smad2 Ser465/467 phosphorylation correlates with ID1 expression; r=0.78, p<0.001)
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Apply time-series clustering to separate primary (0-2 hr) vs secondary (4-8 hr) signaling events
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Experimental validation triage
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Prioritize targets showing ≥2-fold phosphorylation change AND differential gene expression
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Exclude auto-phosphorylation artifacts via kinase inhibitor controls (e.g., 10 µM SB431542)
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Cross-species modeling
This integrated approach recently identified PKCθ Thr538 phosphorylation as a novel amplifier of TGF-β-mediated EMT – a finding validated through CRISPRi knockdown showing 60% reduction in Smad4 nuclear translocation .
How does TGF-β antibody selection influence single-cell RNA-seq experimental outcomes?
Three critical interference mechanisms have been documented:
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Cell surface receptor internalization
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Anti-TGFBR2 antibodies induce 83% receptor internalization within 15 min, distorting downstream signaling readouts
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scRNA-seq library preparation artifacts
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Clone-dependent RNA integrity impacts: 7G3 antibody increases RIN scores by 0.8 versus 1D11-treated cells
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Data interpretation frameworks
Analysis Layer Antibody-Specific Confounder Correction Method Differential expression Antibody-mediated NFκB activation Include IgG isotype controls Trajectory inference Altered cell cycle progression Synchronize cultures pre-treatment
Best practices mandate parallel bulk RNA-seq validation for any scRNA-seq findings using TGF-β antibodies, with particular attention to IL-9/GATA3/Tbet expression ratios as internal consistency controls .
What emerging techniques address TGF-β antibody limitations in live-cell imaging?
The 2024 Live-Smad Translocation Assay (LSTA) overcomes three key challenges:
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Temporal resolution
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Quantitative analysis
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Multiplex capacity
This methodology recently uncovered bistable TGF-β signaling states that eluded traditional endpoint assays – a critical consideration for time-course experiment design.
Product Science Overview
Mouse-anti human TGF-β antibodies are monoclonal antibodies developed in mice that specifically target human TGF-β. These antibodies are used in research and therapeutic applications to neutralize the activity of TGF-β and study its role in various biological processes and diseases . For example, the mouse monoclonal pan-TGF-β neutralizing antibody 1D11 has been shown to inhibit the TGF-β pathway in human basal cell-like breast carcinoma cells .
Applications in Research and Therapy
Mouse-anti human TGF-β antibodies are valuable tools in studying the TGF-β signaling pathway and its involvement in disease progression. They are used to investigate the effects of TGF-β inhibition on cell growth, differentiation, and immune responses . Additionally, these antibodies have therapeutic potential in treating diseases associated with dysregulated TGF-β signaling, such as cancer and fibrosis .
