Cathepsin B Antibody [Unconjugated]

Western blot shows lysates of HepG2 human hepatocellular carcinoma cell line. PVDF membrane was probed with 0.25 µg/mL of Goat Anti-Human Cathepsin B Antigen Affinity-purified Polyclonal Antibody (Catalog # AF953) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF019). A specific band was detected for Cathepsin B at approximately 25-30 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Cathepsin B was detected in immersion fixed paraffin-embedded sections of human brain (cortex) using Goat Anti-Human Cathepsin B Antigen Affinity-purified Polyclonal Antibody (Catalog # AF953) at 10 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS008) 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.

Simple Western lane view shows lysates of HepG2 human hepatocellular carcinoma cell line, loaded at 0.2 mg/mL. A specific band was detected for Cathepsin B at approximately 34 kDa (as indicated) using 2.5 µg/mL of Goat Anti-Human Cathepsin B Antigen Affinity-purified Polyclonal Antibody (Catalog # AF953) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Western blot shows lysates of HeLa human cervical epithelial carcinoma parental cell line and Cathepsin B knockout HeLa cell line (KO). PVDF membrane was probed with 0.25 µg/mL of Goat Anti-Human Cathepsin B Antigen Affinity-purified Polyclonal Antibody (Catalog # AF953) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). Specific bands were detected for Cathepsin B at approximately 25-45 kDa (as indicated) in the parental HeLa cell line, but is not detectable in knockout HeLa cell line. GAPDH (Catalog # AF5718) is shown as a loading control. This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.New adjunct appears with knockout cell line.

Cathepsin B was detected in immersion fixed U‑87 MG human glioblastoma/astrocytoma cell line (Positive) & Daudi human Burkitt's lymphoma cell line (Negative) using Goat Anti-Human Cathepsin B Antigen Affinity-purified Polyclonal Antibody (Catalog # AF953) at 5 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; Catalog # NL001) and counterstained with DAPI (blue). Specific staining was localized to cell surface. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

• Reactivity: Hu
• Applications: WB, Simple Western, IHC, IP, ICC/IF, KO
• Clonality: Polyclonal
• Host: Goat
• Conjugate: Unconjugated
• Concentration: LYOPH

Cathepsin B Antibody [Unconjugated] Summary

• Immunogen: Mouse myeloma cell line NS0-derived recombinant human Cathepsin B
  Arg18-Ile339
  Accession # P07858
• Specificity: Detects human Cathepsin B in direct ELISAs and Western blots. In direct ELISAs, approximately 35% cross-reactivity with recombinant mouse (rm) Cathepsin B is observed and less than 5% cross-reactivity with recombinant human (rh) Cathepsin C, rmCathepsin H, and rhCathepsin L is observed.
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Goat
• Gene: CTSB
• Purity Statement: Antigen Affinity-purified

Applications/Dilutions

• Dilutions:
  • Immunocytochemistry 5-25 ug/mL
  • Immunohistochemistry 5-15 ug/mL
  • Immunoprecipitation 25 ug/mL
  • Knockout Validated
  • Simple Western 2.5 ug/mL
  • Western Blot 0.25 ug/mL

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 Cathepsin B Antibody [Unconjugated]

• APP secretase
• APPS
• Cathepsin B
• Cathepsin B1
• CPSBamyloid precursor protein secretase
• CTSB
• cysteine protease
• EC 3.4.22
• EC 3.4.22.1

Background

Cathepsin B is the first described member of the family of lysosomal cysteine proteases (1). Cathepsin B possesses both endopeptidase and exopeptidase activities, in the latter case acting as a peptidyl-dipeptidase. It is known to process a number of proteins, including pro and active caspases, prorenin, and secretory leucoprotease inhibitor (SLPI) (2-4). Therefore, Cathepsin B may play a role in activation and inactivation of caspases, activation of renin and inactivation of SLPI, the key steps in apoptosis, angiotensin production, and progression of emphysema, respectively. Because of its increased levels and redistribution of the enzyme in human and animal tumors, Cathepsin B may also have role in invasion and metastasis (5).

In addition to lysosome, Cathepsin B can be secreted or associated with plasma membrane, cytoplasm, and nucleus. It is synthesized as a preproenzyme. Following removal of the signal peptide, the inactive proenzyme undergoes further modifications including removal of the pro region to result in the active enzyme (1).

1. Mort, J.S. (2004) in Handbook of Proteolytic Enzymes. Barrett, A.J. et al. (eds): Academic Press, San Diego, p. 1079.
2. Vancompernolle, K. et al. (1998) FEBS Lett. 438:150.
3. Jutras, I. and T.L. Reudelhuber (1999) FEBS Lett. 443:48.
4. Taggart, C.C. et al. (2001) J. Biol. Chem. 276:33345.
5. Bergquin, I.M. and B.F. Sloane (1996) Adv. Exp. Med. Biol. 389:281.

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 Cathepsin B Antibody (AF953)(18)

Publications using AF953

• Y Kovalyova, DW Bak, EM Gordon, C Fung, JHB Shuman, TL Cover, MR Amieva, E Weerapana, SK Hatzios An infection-induced oxidation site regulates legumain processing and tumor growth Nature Chemical Biology, 2022;0(0):. 2022 [PMID: 35332331] (Western Blot, Gerbil)
• U Kern, K Fröhlich, J Bedacht, N Schmidt, ML Biniossek, N Gensch, K Baerenfall, O Schilling Impact of DJ-1 and Helix 8 on the Proteome and Degradome of Neuron-Like Cells Cells, 2021;10(2):. 2021 [PMID: 33669258] (Western Blot, Human)
• D Oelschlaeg, T Weiss Sada, S Salpeter, S Krug, G Blum, W Schmitz, A Schulze, P Michl Cathepsin Inhibition Modulates Metabolism and Polarization of Tumor-Associated Macrophages Cancers (Basel), 2020;12(9):. 2020 [PMID: 32927704] (IHC, Human)
• Y Xu, J Ren, X He, H Chen, T Wei, W Feng YWHA/14-3-3 proteins recognize phosphorylated TFEB by a noncanonical mode for controlling TFEB cytoplasmic localization Autophagy, 2019;0(0):1-14. 2019 [PMID: 30653408] (Western Blot, Human)
• AP Tran, S Sundar, M Yu, BT Lang, J Silver Modulation of Receptor Protein Tyrosine Phosphatase Sigma Increases Chondroitin Sulfate Proteoglycan Degradation through Cathepsin B Secretion to Enhance Axon Outgrowth J. Neurosci., 2018;38(23):5399-5414. 2018 [PMID: 29760175] (Western Blot, Rat)
• CW Lee, JN Stankowski, J Chew, CN Cook, YW Lam, S Almeida, Y Carlomagno, KF Lau, M Prudencio, FB Gao, M Bogyo, DW Dickson, L Petrucelli The lysosomal protein cathepsin L is a progranulin protease Mol Neurodegener, 2017;12(1):55. 2017 [PMID: 28743268] (Western Blot, Human)
• The Unusual Resistance of Avian Defensin AvBD7 to Proteolytic Enzymes Preserves Its Antibacterial Activity PLoS ONE, 2016;11(8):e0161573. 2016 [PMID: 27561012] (Western Blot, Chicken)
• Cathepsin S attenuates endosomal EGFR signalling: A mechanical rationale for the combination of cathepsin S and EGFR tyrosine kinase inhibitors Sci Rep, 2016;6(0):29256. 2016 [PMID: 27387133] (Western Blot, Human)
• Smith R, Solberg R, Jacobsen L, Voreland A, Rustan A, Thoresen G, Johansen H Simvastatin inhibits glucose metabolism and legumain activity in human myotubes. PLoS ONE, 2014;9(1):e85721. 2014 [PMID: 24416446] (Western Blot, Human)
• Ho I, Toh H, Ng W, Teo Y, Guo C, Hui K, Lam P Human bone marrow-derived mesenchymal stem cells suppress human glioma growth through inhibition of angiogenesis. Stem Cells, 2013;31(1):146-55. 2013 [PMID: 23034897] (IHC-Fr, Human)
• Differential secretome analysis reveals CST6 as a suppressor of breast cancer bone metastasis. Cell Res., 2012;22(9):1356-73. 2012 [PMID: 22688893] (Western Blot, Human)
• Noda S, Asano Y, Akamata K, Aozasa N, Taniguchi T, Takahashi T, Ichimura Y, Toyama T, Sumida H, Yanaba K, Tada Y, Sugaya M, Kadono T, Sato S A possible contribution of altered cathepsin B expression to the development of skin sclerosis and vasculopathy in systemic sclerosis. PLoS ONE, 2012;7(2):e32272. 2012 [PMID: 22384200] (IHC-P, Human)
• Shree T, Olson OC, Elie BT Macrophages and cathepsin proteases blunt chemotherapeutic response in breast cancer. Genes Dev., 2011;25(23):2465-79. 2011 [PMID: 22156207] (IHC-P, Human)
• Jak M, van Bochove GG, Reits EA CD40 stimulation sensitizes CLL cells to lysosomal cell death induction by type II anti-CD20 mAb GA101. Blood, 2011;118(19):5178-88. 2011 [PMID: 21948297] (Western Blot, Human)
• Sevenich L, Werner F, Gajda M, Schurigt U, Sieber C, Muller S, Follo M, Peters C, Reinheckel T Transgenic expression of human cathepsin B promotes progression and metastasis of polyoma-middle-T-induced breast cancer in mice. Oncogene, 2011;30(1):54-64. 2011 [PMID: 20818432] (Western Blot, Human)
• Halle A, Hornung V, Petzold GC, Stewart CR, Monks BG, Reinheckel T, Fitzgerald KA, Latz E, Moore KJ, Golenbock DT The NALP3 inflammasome is involved in the innate immune response to amyloid-beta. Nat. Immunol., 2008;9(8):857-65. 2008 [PMID: 18604209] (ICC, Mouse)
• Gocheva V, Zeng W, Ke D, Klimstra D, Reinheckel T, Peters C, Hanahan D, Joyce JA Distinct roles for cysteine cathepsin genes in multistage tumorigenesis. Genes Dev., 2006;20(5):543-56. 2006 [PMID: 16481467] (IHC-P, Human)
• Gronborg M, Kristiansen TZ, Iwahori A, Chang R, Reddy R, Sato N, Molina H, Jensen ON, Hruban RH, Goggins MG, Maitra A, Pandey A Biomarker discovery from pancreatic cancer secretome using a differential proteomic approach. Mol. Cell Proteomics, 2006;5(1):157-71. 2006 [PMID: 16215274] (Western Blot, Human)

Bioinformatics

• Gene Symbol: CTSB
• Uniprot:
  • P07858()