Glypican 3 Antibody [Unconjugated]

Western blot shows lysates of HepG2 human hepatocellular carcinoma cell line. PVDF membrane was probed with 2 µg/mL of Sheep Anti-Human Glypican 3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2119) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for Glypican 3 at approximately 75 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Glypican 3 was detected in immersion fixed HepG2 human hepatocellular carcinoma cell line using Sheep Anti-Human Glypican 3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2119) at 1.7 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Sheep IgG Secondary Antibody (red; Catalog # NL010) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm and cell membranes. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

Glypican 3 was detected in immersion fixed paraffin-embedded sections of human breast using 5 µg/mL Sheep Anti-Human Glypican 3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2119) overnight at 4 °C. Tissue was stained with the Anti-Sheep HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS019) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Glypican 3 was detected in immersion fixed paraffin-embedded sections of human liver cancer tissue using Sheep Anti-Human Glypican 3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2119) at 10 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Sheep HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS019) and counterstained with hematoxylin (blue). Lower panel shows a lack of labeling if primary antibodies are omitted and tissue is stained only with secondary antibody followed by incubation with detection reagents. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Recombinant Human Glypican 3 protein was serially diluted 2-fold and captured by Mouse Anti-Human Glypican 3 Monoclonal Antibody (MAB21191) coated on a Clear Polystyrene Microplate (DY990). Sheep Anti-Human Glypican 3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2119) 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).

• Reactivity: Hu
• Applications: WB, ELISA, Flow, IHC, CyTOF-ready, ICC/IF
• Clonality: Polyclonal
• Host: Sheep
• Conjugate: Unconjugated
• Concentration: LYOPH

Glypican 3 Antibody [Unconjugated] Summary

• Immunogen: Mouse myeloma cell line NS0-derived recombinant human Glypican 3
  Gln25-Val558
  Accession # P51654
• Specificity: Detects human Glypican 3 in direct ELISAs and Western blots. In direct ELISAs, less than 10% cross-reactivity with recombinant mouse Glypican 3 is observed and less than 5% cross-reactivity with recombinant human (rh) Glypican 2, rhGlypican 5, and rhGlypican 6 is observed.
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Sheep
• Gene: GPC3
• Purity Statement: Antigen Affinity-purified

Applications/Dilutions

• Dilutions:
  • CyTOF-ready
  • ELISA
  • Flow Cytometry 2.5 ug/10^6 cells
  • Immunocytochemistry 1-15 ug/mL
  • Immunohistochemistry 5-15 ug/mL
  • Western Blot 2 ug/mL
• Application Notes: Use in IHC-Fr at 1:50 dilution with IF detection for Rat frozen tissues is reported by Grozdanov et al. in PMID 17117158.

Packaging, Storage & Formulations

• 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.
• Buffer: Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
• Preservative: No Preservative
• Concentration: LYOPH
• Reconstitution Instructions: Reconstitute at 0.2 mg/mL in sterile PBS.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Glypican 3 Antibody [Unconjugated]

• DGSX
• Glypican 3
• glypican proteoglycan 3
• glypican-3
• GPC3
• GTR2-2
• heparan sulphate proteoglycan
• Intestinal protein OCI-5
• MXR7
• OCI5
• OCI-5
• secreted glypican-3
• SGB
• SGBS
• SGBS1SDYS

Background

Glypicans (GPC) are a family of heparan sulfate proteoglycans that are attached to the cell surface by a glycosylphosphatidylinositol (GPI) anchor. Six members of this family have been identified in mammals (GPC1-GPC6). All glypican core proteins contain an N-terminal signal peptide, a large globular cysteine-rich domain (CRD) with 14 invariant cysteine residues, a stalk-like region containing the heparan sulfate attachment sites, and a C-terminal GPI attachment site. While glypican proteins do not share strong amino acid sequence identity (they range from 17-63%), the conserved cysteine residues in their CRDs suggests similarity in their three‑dimensional structure (1, 2).

Mutations in GPC3 cause a rare disorder in humans, Simpson-Golabi-Behmel Syndrome, which is characterized by pre and postnatal overgrowth of multiple tissues and organs and an increased risk for developing embryonic tumors (3). These features are also present in the mouse knock-out of GPC3 indicating that GPC3 regulates cell survival and inhibits cell proliferation during development (4). Glypican 3 has been implicated in regulating many different signaling pathways including: IGF, FGF, BMP, and Wnt. An endoproteolytic processing of GPC3 by proprotein convertases is required for the modulation of Wnt signaling (5). Direct interaction with FGF-basic has been observed and is mediated by the heparan sulfate chains (6).

1. Filmus, J. and S.B. Selleck (2001) J. Clinical Invest. 108:497. 
2. De Cat, B and G. David (2001) Seminars in Cell & Dev. Biol. 12:117. 
3. Pilia, G. et al. (1996) Nat. Genet. 12: 241. 
4. Cano-Gauci, D.F. et al. (1999) J. Cell Biol. 146: 255.
5. De Cat, B. et al. (2003) J. Cell Biol. 163:625.
6. Song, H.H. et al. (1997) J. Biol. Chem. 272:7574.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for Glypican 3 Antibody (AF2119)(6)

Publications using AF2119

• N Sun, YT Lee, RY Zhang, R Kao, PC Teng, Y Yang, P Yang, JJ Wang, M Smalley, PJ Chen, M Kim, SJ Chou, L Bao, J Wang, X Zhang, D Qi, J Palomique, N Nissen, SB Han, S Sadeghi, RS Finn, S Saab, RW Busuttil, D Markovic, D Elashoff, HH Yu, H Li, AP Heaney, E Posadas, S You, JD Yang, R Pei, VG Agopian, HR Tseng, Y Zhu Purification of HCC-specific extracellular vesicles on nanosubstrates for early HCC detection by digital scoring Nat Commun, 2020-09-07;11(1):4489. 2020-09-07 [PMID: 32895384] (ELISA Capture, Human)
• YH Khalifa, GM Mourad, WM Stephanos, SA Omar, RA Mehanna Bone Marrow-Derived Mesenchymal Stem Cell Potential Regression of Dysplasia Associating Experimental Liver Fibrosis in Albino Rats Biomed Res Int, 2019-11-06;2019(0):5376165. 2019-11-06 [PMID: 31781620] (IHC-P, Rat)
• M Rahbari, M Pecqueux, D Aust, H Stephan, O Tiebel, A Chatzigeor, T Tonn, F Baenke, V Rao, N Ziegler, H Greif, K Lin, J Weitz, NN Rahbari, C Kahlert Expression of Glypican 3 is an Independent Prognostic Biomarker in Primary Gastro-Esophageal Adenocarcinoma and Corresponding Serum Exosomes J Clin Med, 2019-05-16;8(5):. 2019-05-16 [PMID: 31100935] (Flow Cytometry, Western Blot, Human)
• Phase I trial of a glypican-3-derived peptide vaccine for advanced hepatocellular carcinoma: immunologic evidence and potential for improving overall survival. Clin. Cancer Res., 2012-05-10;18(13):3686-96. 2012-05-10 [PMID: 22577059] (ELISA Development, Human)
• Grozdanov PN, Yovchev MI, Dabeva MD The oncofetal protein glypican-3 is a novel marker of hepatic progenitor/oval cells. Lab. Invest., 2006-10-02;86(12):1272-84. 2006-10-02 [PMID: 17117158] (IHC-Fr, Rat)
• Manton KJ, Sadasivam M, Cool SM, Nurcombe V Bone-specific heparan sulfates induce osteoblast growth arrest and downregulation of retinoblastoma protein. J. Cell. Physiol., 2006-10-01;209(1):219-29. 2006-10-01 [PMID: 16826571] (Western Blot, Human)

Bioinformatics

• Gene Symbol: GPC3
• Uniprot:
  • P51654()