gp96/HSP90B1/GRP94 Antibody (816803) [Unconjugated]

Western blot shows lysates of 293T human embryonic kidney cell line, HeLa human cervical epithelial carcinoma cell line, A20 mouse B cell lymphoma cell line, CH-1 mouse B cell lymphoma cell line, and L6 rat myoblast cell line. PVDF membrane was probed with 0.2 µg/mL of Mouse Anti-Human gp96/HSP90B1 Monoclonal Antibody (Catalog # MAB7606) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF018). A specific band was detected for gp96/HSP90B1 at approximately 100 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

gp96/HSP90B1 was detected in immersion fixed HeLa human cervical epithelial carcinoma cell line using Mouse Anti-Human gp96/HSP90B1 Monoclonal Antibody (Catalog # MAB7606) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Mouse IgG Secondary Antibody (red; Catalog # NL007) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

gp96/HSP90B1 was detected in immersion fixed paraffin-embedded sections of human mesothelioma using Mouse Anti-Human gp96/HSP90B1 Monoclonal Antibody (Catalog # MAB7606) at 15 µg/mL overnight at 4 °C. Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using the Anti-Mouse HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS002) and counterstained with hematoxylin (blue). Specific staining was localized to plasma membranes and cytoplasm. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Simple Western lane view shows lysates of HeLa human cervical epithelial carcinoma cell line, loaded at 0.5 mg/mL. A specific band was detected for gp96/HSP90B1 at approximately 108 kDa (as indicated) using 2 µg/mL of Mouse Anti-Human gp96/HSP90B1 Monoclonal Antibody (Catalog # MAB7606). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Western blot shows lysates of HEK293T human embryonic kidney parental cell line and gp96/HSP90B1 knockout HEK293T cell line (KO). PVDF membrane was probed with 0.2 µg/mL of Mouse Anti-Human gp96/HSP90B1 Monoclonal Antibody (Catalog # MAB7606) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF018). A specific band was detected for gp96/HSP90B1 at approximately 100 kDa (as indicated) in the parental HEK293T cell line, but is not detectable in knockout HEK293T cell line. GAPDH (Catalog # MAB5718) is shown as a loading control. This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

• Reactivity: Hu
• Applications: WB, Simple Western, IHC, ICC/IF, KO
• Clone: 816803
• Clonality: Monoclonal
• Host: Mouse
• Conjugate: Unconjugated
• Concentration: LYOPH

gp96/HSP90B1/GRP94 Antibody (816803) [Unconjugated] Summary

• Immunogen: E. coli-derived recombinant human gp96/HSP90B1
  Arg503-Arg660
  Accession # P14625
• Specificity: Detects human gp96/HSP90B1 in direct ELISAs and Western blots. Detects mouse and rat gp96/HSP90B1 in Western blots.
• Source: N/A
• Isotype: IgG2a
• Clonality: Monoclonal
• Host: Mouse
• Gene: HSP90B1
• Purity Statement: Protein A or G purified from hybridoma culture supernatant

Applications/Dilutions

• Dilutions:
  • Immunocytochemistry 8-25 ug/mL
  • Immunohistochemistry 8-25 ug/mL
  • Knockout Validated
  • Simple Western 2 ug/mL
  • Western Blot 0.2 ug/mL
• Application Notes: In Simple Western only 10-15 uL of the recommended dilution is used per data point.

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: Sterile PBS to a final concentration of 0.5 mg/mL.

Notes

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

Alternate Names for gp96/HSP90B1/GRP94 Antibody (816803) [Unconjugated]

• 94 kDa glucose-regulated protein
• ECGP
• Endoplasmin
• endothelial cell (HBMEC) glycoprotein
• glucose regulated protein, 94 kDa
• gp96
• GP96ECGP
• Grp94
• GRP-94
• GRP94gp96 homolog
• Heat shock protein 90 kDa beta member 1
• heat shock protein 90kDa beta (Grp94), member 1
• HSP90B1
• TRA1endoplasmin
• tumor rejection antigen (gp96) 1
• Tumor rejection antigen 1
• Tumor rejection antigen-1 (gp96)

Background

Glycoprotein 96 (gp96; also endoplasmin, GRP-94, TRA1 and HSP90B1) is a 94-100 kDa member of the HSP 90 family of proteins. gp96 is a ubiquitously-expressed ER resident protein that is found in a preformed complex with BiP, CaBP1 and UDP-glucosyltransferase. This is a chaperone complex that binds unfolded protein substrates. When folded properly, the substrate is forwarded to calnexin-containing chaperone complexes that promote its maturation. gp96 clients are restricted and include disulfide-bonded integrins, TLRs, LDLR and CD180. Within the complex, gp96 exists as a disulfide-linked homodimer that may form higher-order oligomers. gp96 also appears on the cell surface and may serve as a receptor for bacteria. Mature human gp96 is a 782 amino acid (aa) membrane-associated protein
(aa 22-803). It is not a transmembrane protein but utilizes an ER retention signal (aa 800-803) to interact with the ER membrane. The molecule possesses a
HATPase-C like region (aa 98-219) plus multiple ATP binding and two utilized phosphorylation sites. Over aa 503-660, human gp96 shares 98% aa sequence identity with mouse gp96.

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 gp96/HSP90B1/GRP94 Antibody (MAB7606)(4)

Publications using MAB7606

• DJ Busch, Y Zhang, A Kumar, SC Huhn, Z Du, R Liu Identification of RNA Content of CHO-derived Extracellular Vesicles from a Production Process Journal of biotechnology, 2022-03-12;0(0):. 2022-03-12 [PMID: 35292346] (Western Blot, Hamster, Virus)
• N Jongkamonw, MA Ramirez, S Edassery, ACY Wong, J Yu, T Abbott, K Pak, AF Ryan, JN Savas Noise Exposures Causing Hearing Loss Generate Proteotoxic Stress and Activate the Proteostasis Network Cell Rep, 2020-11-24;33(8):108431. 2020-11-24 [PMID: 33238128] (IHC-Fr, Mouse)
• D Matthies, NYJ Lee, I Gatera, HA Pasolli, X Zhao, H Liu, D Walpita, Z Liu, Z Yu, MS Ioannou Microdomains form on the luminal face of neuronal extracellular vesicle membranes Sci Rep, 2020-07-20;10(1):11953. 2020-07-20 [PMID: 32686698] (Western Blot, Rat)
• Natalia A. Cherepanova, Sergey V. Venev, John D. Leszyk, Scott A. Shaffer, Reid Gilmore Quantitative glycoproteomics reveals new classes of STT3A- and STT3B-dependent N-glycosylation sites Journal of Cell Biology 8/5/2019 [PMID: 31296534]

Bioinformatics

• Gene Symbol: HSP90B1
• Uniprot:
  • P14625(Human)