TNFSF8 Antibody

Introduction to TNFSF8 and its Antibodies

TNFSF8, also known as CD30 Ligand (CD30L) or CD153, is a type II membrane protein belonging to the tumor necrosis factor (TNF) superfamily. The human TNFSF8 protein has a calculated molecular weight of approximately 26 kDa, although glycosylation often results in an observed molecular weight of 35-40 kDa in experimental settings . The protein sequence corresponds to Gln63-Asp234 of the full human TNFSF8 protein, as referenced in multiple antibody specifications .

TNFSF8 antibodies are immunoglobulins specifically designed to recognize and bind to the CD30 ligand protein. These antibodies come in various formats including monoclonal, polyclonal, and recombinant variants, each with specific applications in research and potential diagnostic settings. The development of these antibodies has significantly advanced our understanding of CD30/CD30L signaling pathways and their role in both normal immune function and disease states .

Molecular Structure

TNFSF8 is characterized as a type II transmembrane protein, meaning its N-terminus is located in the cytoplasm while the C-terminus extends extracellularly. The protein belongs to the tumor necrosis factor (TNF) superfamily, sharing structural homology with other members of this important group of cytokines . The human TNFSF8 gene is located on chromosome 9q33, and the protein contains 234 amino acids with a predicted molecular mass of approximately 26 kDa .

Cellular Expression

TNFSF8 exhibits a distinct expression pattern primarily within the immune system. The protein is expressed on the cell surface of several immune cell types, including:

• Activated T cells, B cells, and monocytes

• Granulocytes (constitutively expressed)

• Medullary thymic epithelial cells (constitutively expressed)

• Macrophages, eosinophils, neutrophils, and mast cells

Immunohistochemical studies using TNFSF8 antibodies have demonstrated specific staining in human thymus (particularly in Hassall's corpuscles), placenta (endothelial cells in villi), and spleen tissues . Flow cytometry analysis has confirmed expression on activated peripheral blood mononuclear cells (PBMCs), particularly after stimulation with agents such as PMA and calcium ionomycin .

Receptor Interaction

The specific receptor for TNFSF8 is CD30 (TNFRSF8), a type I transmembrane glycoprotein belonging to the TNF receptor superfamily. CD30 was originally identified as a cell surface antigen of Hodgkin's and Reed-Sternberg cells using the monoclonal antibody Ki-1 .

CD30 expression is observed on:

• Hodgkin's and Reed-Sternberg cells

• Certain non-Hodgkin's lymphomas

• Virus-infected T and B cells

• Normal T and B cells following activation

• CD4+/CD8+ thymocytes co-expressing CD45RO and IL-4 receptor

• A subset of T cells producing Th2-type cytokines

Signaling Effects

The binding of TNFSF8 to CD30 initiates a cascade of intracellular signaling events that lead to diverse cellular responses. These effects include:

• Cell proliferation and activation

• Cellular differentiation

• Apoptotic cell death in certain contexts

• NF-κB pathway activation

• Production of cytokines

The CD30/CD30L signaling axis plays a critical role in immune regulation, particularly in T cell development and function. Research has demonstrated its importance in thymic negative selection, a process essential for eliminating self-reactive T cells and preventing autoimmunity .

Types of Available Antibodies

Multiple commercial vendors offer TNFSF8 antibodies with varying specifications. The following table summarizes key commercially available antibodies:

FC: Flow Cytometry; IHC: Immunohistochemistry; WB: Western Blot; IF: Immunofluorescence; ICC: Immunocytochemistry; ELISA: Enzyme-Linked Immunosorbent Assay

Research Applications

TNFSF8 antibodies serve numerous research applications that help elucidate the biology of CD30/CD30L signaling:

Flow Cytometry

Flow cytometry applications allow for the detection and quantification of TNFSF8 expression on cell surfaces. Studies have demonstrated successful staining of activated human PBMCs, particularly after stimulation with agents such as PMA and calcium ionomycin . This technique enables researchers to identify specific cell populations expressing TNFSF8 and monitor expression changes under various conditions.

Immunohistochemistry

TNFSF8 antibodies have been validated for immunohistochemical detection in various human tissues. Notable findings include:

• Localization to Hassall's corpuscles in human thymus

• Expression in endothelial cells of placental villi

• Detection on cell surfaces in splenic nodules

These applications provide valuable insights into the tissue distribution and cellular localization of TNFSF8 in physiological and pathological states.

Protein Interaction Studies

Antibodies against TNFSF8 have been utilized in protein interaction studies to investigate binding with CD30 receptor. For example, immobilized Human CD30 Protein can bind Human CD30 Ligand Protein with a linear range of 0.039-0.313 μg/mL, as determined through in vitro binding assays .

Expression Analysis

Flow cytometry analysis has revealed that TNFSF8 expression is significantly upregulated following cellular activation. In human peripheral blood mononuclear cells (PBMCs) treated with activating agents such as PMA (50 ng/mL) and calcium ionomycin (200 ng/mL), TNFSF8 expression becomes readily detectable after 16 hours of stimulation . This inducible expression pattern underscores the protein's role in activated immune responses rather than homeostatic conditions.

In mouse models, anti-CD3/CD28 treatment of splenocytes induces detectable TNFSF8 expression, confirming the conservation of this activation-dependent expression pattern across species .

Binding and Functional Studies

Binding studies utilizing bio-layer interferometry (BLI) have determined that human TNFSF8 protein binds to CD30 with an affinity constant of approximately 132 nM . This quantitative measurement provides important insights into the strength of the receptor-ligand interaction.

Functional assays have demonstrated that TNFSF8 can induce IL-6 secretion when bound to CD30, highlighting its role in cytokine production and inflammatory responses . Furthermore, this interaction can be blocked by neutralizing anti-CD30 antibodies, with an IC50 of approximately 0.2401 μg/mL in certain experimental systems .

Tissue Localization Studies

Immunohistochemical analyses have provided detailed information about TNFSF8 localization in various tissues:

1. Human Thymus: TNFSF8 shows specific localization to Hassall's corpuscles, specialized structures within the medulla of the thymus that play roles in T cell development and selection .

2. Human Placenta: TNFSF8 expression is detected in endothelial cells within the villi, suggesting potential roles in placental immunity or vascular function .

3. Human Spleen: TNFSF8 staining is observed on cell surfaces within splenic nodules, consistent with its expression on activated lymphocytes .

These tissue distribution patterns provide important clues about the physiological roles of TNFSF8 in different organ systems and microenvironments.

Role in Lymphoma and Cancer

The CD30/CD30L signaling axis has significant clinical relevance, particularly in lymphoid malignancies. CD30 (the receptor for TNFSF8) serves as an important clinical marker for Hodgkin lymphoma and anaplastic large cell lymphomas, where it shows notably high expression . This expression pattern has made CD30 a valuable diagnostic marker and therapeutic target in these conditions.

While TNFSF8 itself has been less extensively studied in the clinical context than its receptor, understanding its expression and function could provide additional insights into the pathophysiology of CD30-positive malignancies and potentially reveal new therapeutic approaches .

Immunoregulatory Functions

Beyond cancer, the CD30/CD30L system plays important roles in immunoregulation. Research suggests involvement in:

• Thymic negative selection (eliminating self-reactive T cells)

• Regulation of Th1/Th2 balance

• Modulation of inflammatory responses

Dysregulation of this signaling pathway may contribute to various immunological disorders, though further research is needed to fully elucidate these connections.

Therapeutic Potential

While current TNFSF8 antibodies are primarily research tools, their continued development could potentially lead to therapeutic applications. Given the established role of CD30 as a therapeutic target in certain lymphomas (e.g., with brentuximab vedotin), modulation of the ligand side of this interaction represents an alternative approach worthy of investigation .

Diagnostic Applications

The specificity of TNFSF8 antibodies for their target makes them valuable tools for diagnostic applications. Further development of standardized immunohistochemical or flow cytometry protocols using these antibodies could enhance the precision of lymphoma classification or identification of specific immune cell subsets in various disease states .

Technical Advancements

Ongoing improvements in antibody engineering technologies are likely to yield TNFSF8 antibodies with enhanced specificity, sensitivity, and functionality. The development of recombinant antibody formats, already seen in some commercial offerings, represents an important advancement that may improve reproducibility and reduce batch-to-batch variation .