Human CD30/TNFRSF8 Antibody

R&D Systems, part of Bio-Techne | Catalog # AF229

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AF229
AF229-SP
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Key Product Details

Species Reactivity

Human

Applications

Western Blot, ELISA, Immunocytochemistry, Simple Western, Agonist Activity

Label

Unconjugated

Antibody Source

Polyclonal Goat IgG

View all CD30/TNFRSF8 Antibodies »

Product Specifications

Immunogen

S. frugiperda insect ovarian cell line Sf 21-derived recombinant human CD30
Phe19-Lys379
Accession # P28908

Specificity

Detects human CD30 in direct ELISAs and Western blots.

Clonality

Polyclonal

Host

Goat

Isotype

IgG

Endotoxin Level

<0.10 EU per 1 μg of the antibody by the LAL method.

Scientific Data Images for Human CD30/TNFRSF8 Antibody

CD30/TNFRSF8 antibody in Jurkat Human Cell Line by Immunocytochemistry (ICC).

CD30/TNFRSF8 in Jurkat Human Cell Line.

CD30/TNFRSF8 was detected in immersion fixed Jurkat human acute T cell leukemia cell line using Goat Anti-Human CD30/TNFRSF8 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF229) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; NL001) and counterstained with DAPI (blue). Specific staining was localized to cell membrane and cytoplasm. View our protocol for Fluorescent ICC Staining of Non-adherent Cells.

Detection of Human CD30/TNFRSF8 by Simple WesternTM.

Left: Simple Western lane view shows lysates of HDLM-2 human Hodgkin’s lymphoma cell line, loaded at 0.5 mg/ml. A specific band was detected for CD30/TNFRSF8 at approximately 178 kDa (as indicated) using both 10 µg/ml and 50 µg/ml of Goat Anti-Human CD30/TNFRSF8 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF229) followed by HRP-conjugated Donkey Anti-Goat Secondary Antibody (Catalog # 043-522-2). This experiment was conducted under reducing conditions and using the 12-230kDa separation system. Right: Simple Western electropherogram showing the same Goat Anti-Human CD30/TNFRSF8 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF229) tested at 10 µg/ml (blue line) and 50 µg/ml (green line) in the HDLM-2 human Hodgkin’s lymphoma cell line.

Human CD30/TNFRSF8 ELISA Standard Curve.

Recombinant Human CD30/TNFRSF8 protein was serially diluted 2-fold and captured by Mouse Anti-Human CD30/TNFRSF8 Monoclonal Antibody (MAB2291) coated on a Clear Polystyrene Microplate (DY990). Goat Anti-Human CD30/TNFRSF8 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF229) was biotinylated and incubated with the protein captured on the plate. Detection of the standard curve was achieved by incubating Streptavidin-HRP (DY998) followed by Substrate Solution (DY999) and stopping the enzymatic reaction with Stop Solution (DY994).

Applications for Human CD30/TNFRSF8 Antibody

Application
Recommended Usage

Agonist Activity

Measured by its ability to stimulate human IL-6 secretion by HDLM-2 human Hodgkin’s lymphoma cells. Gruss H.J. et al. (1996) Blood 87(6):2443.
The ED50 for this effect is typically 0.2 - 0.6 μg/mL.

ELISA

This antibody functions as an ELISA detection antibody when paired with Mouse Anti-Human CD30/TNFRSF8 Monoclonal Antibody (Catalog # MAB2291).

This product is intended for assay development on various assay platforms requiring antibody pairs. We recommend the Human CD30/TNFRSF8 DuoSet ELISA Kit (Catalog # DY6126-05) for convenient development of a sandwich ELISA.

Immunocytochemistry

5-15 µg/mL
Sample: Immersion fixed Jurkat human acute T cell leukemia cell line

Simple Western

10-50 µg/mL
Sample: HDLM-2 human Hodgkin's lymphoma cell line

Western Blot

0.1 µg/mL
Sample: Recombinant Human CD30/TNFRSF8 Fc Chimera (Catalog # 813-CD)

Reviewed Applications

Read 1 review rated 5 using AF229 in the following applications:

Formulation, Preparation, and Storage

Purification

Antigen Affinity-purified

Reconstitution

Reconstitute at 0.2 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.

Reconstitution Buffer Available:
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Qty

Formulation

Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. See Certificate of Analysis for details.
*Small pack size (-SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

Stability & Storage

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.

Background: CD30/TNFRSF8

CD30, also known as Ki-1 antigen and TNFRSF8, is a 120 kDa type I transmembrane glycoprotein belonging to the TNF receptor superfamily (1, 2). Mature human CD30 consists of a 361 amino acid (aa) extracellular domain (ECD) with six cysteine-rich repeats, a 28 aa transmembrane segment, and a 188 aa cytoplasmic domain (3). In contrast, mouse and rat CD30 lack 90 aa of the ECD and contain only three cysteine-rich repeats. Within common regions of the ECD, human CD30 shares 53% and 49% aa sequence identity with mouse and rat CD30, respectively. Alternate splicing of human CD30 generates an isoform that includes only the C‑terminal 132 aa of the cytoplasmic domain. CD30 is normally expressed on antigen-stimulated Th cells and B cells (4 - 6). However, it is upregulated in Hodgkin’s disease (on Reed-Sternberg cells), other lymphomas, chronic inflammation, and autoimmunity (7). CD30 binds to CD30 Ligand/TNFSF8 which is expressed on activated Th cells, monocytes, granulocytes and medullary thymic epithelial cells (1, 5). CD30 signaling costimulates antigen-induced Th0 and Th2 proliferation and cytokine secretion but favors a Th2-biased immune response (8). In the absence of antigenic stimulation, it can still induce T cell expression of IL-13 (9). CD30 contributes to thymic negative selection by inducing the apoptotic cell death of CD4+CD8+ T cells (10, 11). In B cells, CD30 ligation promotes cellular proliferation and antibody production in addition to the expression of CXCR4, CCL3, and CCL5 (5, 12). An 85-90 kDa soluble form of CD30 is shed from the cell surface by TACE-mediated cleavage (13, 14). Soluble CD30 retains the ability to bind CD30 Ligand and functions as an inhibitor of normal CD30 signaling (15).

References

  1. Kennedy, M.K. et al. (2006) Immunology 118:143.
  2. Tarkowski, M. (2003) Curr. Opin. Hematol. 10:267.
  3. Durkop, H. et al. (1992) Cell 68:421.
  4. Hamann, D. et al. (1996) J. Immunol. 156:1387.
  5. Shanebeck, S.D. et al. (1995) Eur. J. Immunol. 25:2147.
  6. Gruss, H.-J. et al. (1994) Blood 83:2045.
  7. Oflzoglu E. et al. (2009) Adv. Exp. Med. Biol. 647:174.
  8. Del Prete, G. et al. (1995) J. Exp. Med. 182:1655.
  9. Harlin, H. et al. (2002) J. Immunol. 169:2451.
  10. Amakawa, R. et al. (1996) Cell 84:551.
  11. Chiarle, R. et al. (1999) J. Immunol. 163:194.
  12. Vinante, F. et al. (2002) Blood 99:52.
  13. Hansen, H.P. et al. (1995) Int. J. Cancer 63:750.
  14. Hansen, H.P. et al. (2000) J. Immunol. 165:6703.
  15. Hargreaves, P.G. and A. Al-Shamkhani (2002) Eur. J. Immunol. 32:163.

Alternate Names

CD30, TNFRSF8

Entrez Gene IDs

943 (Human); 21941 (Mouse); 25069 (Rat)

Gene Symbol

TNFRSF8

UniProt

P28908

Additional CD30/TNFRSF8 Products

Product Documents for Human CD30/TNFRSF8 Antibody

Product Datasheets

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Human CD30/TNFRSF8 Antibody

For research use only

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