TNFSF13B Antibody

What is TNFSF13B and what are its alternative designations in scientific literature?

TNFSF13B (Tumor Necrosis Factor Superfamily Member 13B) is a type II transmembrane glycoprotein belonging to the TNF superfamily. In the scientific literature, it is also known as BAFF (B-cell Activating Factor), BLyS (B Lymphocyte Stimulator), TALL-1, THANK, TNFSF20, ZTNF4, and CD257 . Human TNFSF13B is a 285 amino acid protein consisting of a 218 aa extracellular domain, a 21 aa transmembrane region, and a 46 aa cytoplasmic tail . The protein can exist in both membrane-bound and soluble forms, with the soluble form resulting from proteolytic cleavage between R133 and Ala134 .

What are the primary receptors for TNFSF13B and their cellular distribution?

TNFSF13B interacts with three distinct TNF receptor family members:

All three receptors are type III transmembrane proteins primarily expressed on B cells, though expression on other immune cell types has been documented . TNFSF13B shares with APRIL (another TNF family member) the ability to bind to BCMA and TACI, while BAFF-R is specific for TNFSF13B .

What is the physiological role of TNFSF13B in the immune system?

TNFSF13B serves as a potent B cell activator and plays critical roles in:

• Providing survival signals to T2 B cells (B cells with intermediate maturation status)

• Supporting B cell development, maturation, and differentiation

• Promoting B cell proliferation and antibody class switching

• Contributing to the development of lymphoid tissue architecture

• Enhancing survival of activated memory B cells

Its expression is primarily found in peripheral blood mononuclear cells, monocytes, macrophages, dendritic cells, and is upregulated by inflammatory stimuli including IFN-alpha, IFN-beta, LPS, and IL-10 .

What key criteria should be considered when selecting a TNFSF13B antibody for specific experimental applications?

When selecting a TNFSF13B antibody, researchers should consider:

• Application compatibility: Ensure the antibody is validated for your specific application (WB, IHC, flow cytometry, ELISA, neutralization) .

• Species reactivity: Verify cross-reactivity with your target species. Note that human and mouse TNFSF13B share 86% amino acid sequence identity .

• Epitope recognition: Consider whether you need to detect:

  • Membrane-bound vs. soluble forms

  • Full-length vs. cleaved forms

  • Denatured vs. native conformations

• Clone type: Choose between:

  • Monoclonal antibodies (like clones #137317 and #137314) for specific epitope recognition

  • Polyclonal antibodies for broader epitope detection

• Conjugation needs: Depending on your application, select from:

  • Unconjugated antibodies for flexibility

  • Fluorophore-conjugated antibodies (e.g., Alexa Fluor 488) for direct detection

What are the optimal protocols for detecting TNFSF13B using immunohistochemistry?

For optimal TNFSF13B detection in tissue sections:

• Tissue preparation:

  • Use heat-induced epitope retrieval with Antigen Retrieval Reagent-Basic (pH 9.0) or citrate buffer (pH 6.0)

  • For paraffin-embedded sections, immersion fixation is recommended

• Antibody concentration and incubation:

  • For human lymph node tissue, use Mouse Anti-Human TNFSF13B at 5 μg/mL

  • Incubate for 1 hour at room temperature

  • Follow with appropriate secondary antibody (e.g., Anti-Mouse IgG HRP Polymer)

• Detection system:

  • DAB (3,3'-diaminobenzidine) substrate yields brown staining

  • Counterstain with hematoxylin for nuclear visualization

• Expected results:

  • Specific staining should be localized to cell surface and cytoplasm

  • Positive staining has been reported in human lymph node and intrahepatic cholangiocarcinoma tissues

How can flow cytometry be optimized for TNFSF13B detection?

For successful TNFSF13B detection by flow cytometry:

• Cell preparation:

  • THP-1 cells (human monocytic cell line) serve as a positive control

  • Ensure single-cell suspension and appropriate fixation

• Staining protocol:

  • Use Mouse Anti-Human TNFSF13B monoclonal antibody (e.g., MAB1241)

  • Compare with isotype control antibody to assess background staining

  • Follow with fluorochrome-conjugated secondary antibody (e.g., PE-conjugated Anti-Mouse IgG)

• Gating strategy:

  • Use forward/side scatter to identify viable cells

  • Compare fluorescence intensity between experimental and isotype control samples

• Analysis considerations:

  • TNFSF13B expression may vary with cell activation state

  • Surface and intracellular staining may be needed to detect both membrane-bound and intracellular forms

How can researchers accurately distinguish between membrane-bound and soluble TNFSF13B in experimental systems?

Differentiating between membrane-bound and soluble TNFSF13B requires specific methodological approaches:

• Cell surface vs. secreted protein:

  • Flow cytometry with non-permeabilized cells detects only membrane-bound TNFSF13B

  • ELISA of culture supernatants or biological fluids identifies soluble TNFSF13B

• Western blot analysis:

  • Membrane-bound form: ~32 kDa band in cell lysates

  • Soluble form: ~17 kDa band in supernatants/serum

  • Note that glycosylated forms may appear at 45-52 kDa

• Proteolytic processing detection:

  • Antibodies recognizing epitopes near the R133-Ala134 cleavage site can differentiate intact vs. cleaved forms

  • Inhibitors of TNFSF13B-cleaving proteases can help quantify processing rates

• Functional assessment:

  • Neutralizing antibodies against soluble TNFSF13B can help determine the relative contribution of membrane vs. soluble forms in biological responses

What is the relationship between TNFSF13B genetic variants and autoimmune disease susceptibility?

Research has revealed significant associations between TNFSF13B genetic variants and autoimmune diseases:

• BAFF-var insertion-deletion variant:

  • A rare variant (GCTGT > A) in TNFSF13B (called BAFF-var) has been significantly associated with:

    • Systemic Lupus Erythematosus (SLE) in Spanish populations (p = 0.001, OR = 1.41)

    • SLE in German populations (p = 0.030, OR = 1.86)

    • Rheumatoid Arthritis (RA)

  • Meta-analysis confirmed this association (p = 0.0002, OR = 1.46)

• rs9514828 (-871 C > T) polymorphism:

  • Associated with altered TNFSF13B mRNA expression in healthy subjects and SLE patients

  • Linked to susceptibility in multiple autoimmune disorders

• rs1041569 (-2701 A > T) polymorphism:

  • Associated with increased risk of inflammatory bowel disease in Greek populations

• Mechanism of action:

  • BAFF-var results in a shorter transcript that escapes microRNA inhibition

  • This leads to increased production of soluble BAFF protein

  • Associated with augmented levels of total IgG and IgM and reduced monocyte counts

How can TNFSF13B expression be used as a prognostic biomarker in disease settings?

TNFSF13B has emerging value as a prognostic biomarker in several diseases:

• Cancer prognosis:

  • High TNFSF13B expression correlates with poor prognosis in adrenocortical carcinoma (ACC)

  • TNFSF13B has been identified as a prognostic biomarker for renal clear cell carcinoma

  • Gene Set Enrichment Analysis (GSEA) shows TNFSF13B expression relates to immune signaling pathways and lymphocyte infiltration

• Inflammatory disease severity:

  • Plasma TNFSF13B levels serve as inflammatory indicators in severe HAdV pneumonia in pediatric patients

  • ROC analysis shows plasma TNFSF13B has 74.38% accuracy in predicting severe HAdV pneumonia cases

  • At a cut-off value of 11,324 pg/ml, TNFSF13B provides 76.19% sensitivity and 80.95% specificity in predicting severe cases

• Autoimmune disease activity:

  • Increased TNFSF13B expression correlates with autoantibody production in SLE and RA

  • Mouse models of SLE and RA showed elevated serum TNFSF13B levels, and BAFF blockade reduced disease manifestations

What are the common technical challenges when detecting TNFSF13B by Western blot and how can they be addressed?

Researchers frequently encounter these challenges when detecting TNFSF13B by Western blot:

• Multiple bands:

  • Expected molecular weights: 32 kDa (primary band), 45-52 kDa (glycosylated forms)

  • Challenge: Additional bands may represent:

    • Post-translational modifications

    • Proteolytic fragments

    • Multimeric forms (TNFSF13B forms homotrimers)

  • Solution: Include positive controls (e.g., recombinant TNFSF13B) and verify with multiple antibodies recognizing different epitopes

• Low signal intensity:

  • Challenge: Endogenous TNFSF13B may be expressed at low levels in some cell types

  • Solution: Enrich for TNFSF13B-expressing cells or stimulate with IFN-alpha, IFN-beta, LPS, or IL-10 to upregulate expression

• Sample preparation:

  • Challenge: Protein degradation during preparation

  • Solution: Use fresh samples with protease inhibitors; avoid repeated freeze-thaw cycles

• Antibody dilution optimization:

  • Challenge: Finding optimal antibody concentration

  • Solution: Test a range of dilutions (1:500-1:2000 recommended for WB with polyclonal anti-TNFSF13B)

How can researchers validate the functional neutralization activity of anti-TNFSF13B antibodies?

To validate neutralizing activity of anti-TNFSF13B antibodies:

• B cell proliferation assay:

  • Recombinant TNFSF13B stimulates B cell proliferation in the presence of anti-IgM antibodies

  • Measure neutralization by adding increasing concentrations of anti-TNFSF13B antibody

  • Calculate the neutralization dose (ND50) that inhibits 50% of TNFSF13B-induced proliferation

  • For mouse TNFSF13B, the ND50 is typically 0.01-0.04 μg/mL in the presence of 3 ng/mL recombinant protein

• Receptor binding inhibition:

  • Use ELISA-based competition assays to determine if the antibody blocks TNFSF13B-receptor interactions

  • Test against all three receptors (BAFF-R, TACI, BCMA) as neutralization may be receptor-specific

• Cell signaling inhibition:

  • Assess whether the antibody blocks TNFSF13B-induced NF-κB activation

  • Monitor phosphorylation of downstream signaling molecules

• In vivo validation:

  • In mouse models of SLE or RA, neutralizing antibodies should reduce disease manifestations

  • Measure serum immunoglobulin levels and autoantibody production before and after treatment

How is TNFSF13B being investigated as a therapeutic target in autoimmune diseases?

TNFSF13B has become an important therapeutic target in autoimmune diseases:

• Current therapeutic approaches:

  • BAFF inhibitors are in clinical trials for systemic lupus erythematosus (SLE)

  • Strategies include:

    • Monoclonal antibodies against soluble TNFSF13B

    • Receptor fusion proteins that act as decoy receptors

    • Small molecule inhibitors of TNFSF13B-receptor interactions

• Therapeutic rationale:

  • TNFSF13B is overexpressed in multiple autoimmune diseases including SLE and RA

  • BAFF-var genetic variant leads to increased TNFSF13B production and is associated with autoimmune disease risk

  • In mouse models, TNFSF13B blockade reduces disease manifestations

• Considerations for therapeutic development:

  • Targeting specificity: Distinguish TNFSF13B from APRIL which shares some receptors

  • Balance immunosuppression against infection risk

  • Monitor effects on normal B cell development and antibody responses

What role does TNFSF13B play in inflammatory respiratory diseases and potential therapeutic applications?

TNFSF13B has emerging significance in respiratory inflammation:

• Expression in pulmonary conditions:

  • Elevated TNFSF13B levels detected in:

    • Bronchoalveolar lavage fluid of patients with idiopathic pulmonary fibrosis

    • Airways during respiratory syncytial virus (RSV) infections

    • Excessive expression in chronic obstructive pulmonary disease (COPD) leads to aggravated lung inflammation and alveolar wall destruction

• Role in viral pneumonia:

  • TNFSF13B levels are dramatically increased in both plasma and BAL fluid of severe HAdV pneumonia patients

  • Plasma TNFSF13B positively correlates with C-reactive protein (CRP) levels

  • Functions as a key molecule in local and systemic inflammatory networks

• Diagnostic potential:

  • ROC analysis shows plasma TNFSF13B can predict severe HAdV pneumonia cases with 74.38% accuracy

  • At a cut-off value of 11,324 pg/ml, provides 76.19% sensitivity and 80.95% specificity

• Future therapeutic directions:

  • TNFSF13B neutralization might reduce pulmonary inflammation

  • Potential applications in severe viral pneumonia, including HAdV infection

  • May benefit conditions with excessive B cell activation in the lung