Human Legumain/Asparaginyl Endopeptidase Antibody

Catalog # Availability Size / Price Qty
AF2199
AF2199-SP
Detection of Human Legumain/ Asparaginyl Endopeptidase by Western Blot.
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Product Details
Citations (17)
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Human Legumain/Asparaginyl Endopeptidase Antibody Summary

Species Reactivity
Human
Specificity
Detects human Legumain/Asparaginyl Endopeptidase in direct ELISAs and Western blots.
Source
Polyclonal Goat IgG
Purification
Antigen Affinity-purified
Immunogen
Mouse myeloma cell line NS0-derived recombinant human Legumain/Asparaginyl Endopeptidase
Ile18-Tyr433
Accession # Q99538
Formulation
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.
Label
Unconjugated

Applications

Recommended Concentration
Sample
Western Blot
1 µg/mL
See below
Immunoprecipitation
25 µg/mL
Conditioned cell culture medium spiked with Recombinant Human Legumain (Catalog # 2199‑CY), see our available Western blot detection antibodies

Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.

Scientific Data

Western Blot Detection of Human Legumain/ Asparaginyl Endopeptidase antibody by Western Blot. View Larger

Detection of Human Legumain/ Asparaginyl Endopeptidase by Western Blot. Western blot shows lysates of human kidney tissue, human heart tissue, and human placenta tissue. PVDF membrane was probed with 1 µg/mL of Goat Anti-Human Legumain/Asparaginyl Endopeptidase Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2199) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). Specific bands were detected for Pro-Legumain/ Asparaginyl Endopeptidase at approximately 56 kDa and mature Legumain/Asparaginyl Endopeptidase 37 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Western Blot Detection of Human Legumain/Asparaginyl Endopeptidase by Western Blot View Larger

Detection of Human Legumain/Asparaginyl Endopeptidase by Western Blot The expression of AEP was higher in diffuse type gastric cancer than that in intestinal type gastric cancerA. AEP expression in intestinal type gastric cancer and diffuse type gastric cancer were detected by western blot (Representation: I, intestinal type gastric cancer; D, diffuse type gastric cancer, P=0.032). B. Patients' basic characteristics. Image collected and cropped by CiteAb from the following publication (https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.8879), licensed under a CC-BY license. Not internally tested by R&D Systems.

Immunocytochemistry/ Immunofluorescence Detection of Mouse Legumain/Asparaginyl Endopeptidase by Immunocytochemistry/Immunofluorescence View Larger

Detection of Mouse Legumain/Asparaginyl Endopeptidase by Immunocytochemistry/Immunofluorescence Representative sections showing overall higher expression of legumain (diffuse yellowish fluorescence staining) in the untreated (A) versus the Andosan™-treated (B) intestine.Notably, legumain expression was higher in tumor tissue seen in untreated animals. Scale bars represent 200 μm. Image collected and cropped by CiteAb from the following publication (https://dx.plos.org/10.1371/journal.pone.0167754), licensed under a CC-BY license. Not internally tested by R&D Systems.

Immunohistochemistry Detection of Human Legumain/Asparaginyl Endopeptidase by Immunohistochemistry View Larger

Detection of Human Legumain/Asparaginyl Endopeptidase by Immunohistochemistry AEP and E-cadherin expression were detected both in primary gastric cancer and peritoneal metastatic lociA. The expression of AEP and E-cadherin in primary gastric cancer and metastatic peritoneal loci were firstly investigated by immunohistochemistry. AEP expression were higher and E-cadherin expression was lower in peritoneal metastatic lesions than in primary gastric cancer (magnified 50× in column 1,3; magnified 200× in column 2,4). B. The expression of AEP and E-cadherin in primary gastric cancer and metastatic peritoneal loci were investigated by immunofluorescence assay (magnified 200×). AEP, the secondary antibody of which were labeled with green fluorescence, was more strongly expressed in peritoneal metastatic lesions than in primary gastric cancer. E-cadherin, the secondary antibody of which was labeled with red fluorescence, was expressed in primary gastric cancer and could not be identified in peritoneal metastatic loci. Image collected and cropped by CiteAb from the following publication (https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.8879), licensed under a CC-BY license. Not internally tested by R&D Systems.

Western Blot Detection of Mouse Legumain/Asparaginyl Endopeptidase by Western Blot View Larger

Detection of Mouse Legumain/Asparaginyl Endopeptidase by Western Blot AEP knockdown and overexpressive vectors were constructed and stably transfected gastric cancer cell linesA. Constructing of the AEP knocked-down lentiviral vector, it was verified by western blot that AEP was inhibited corresponding to AEP knockdown. B. Overexpressing AEP lentiviral vector was constructed and AEP indeed increased corresponding to AEP overexpression. C. The proliferative curve in response to AEP overexpression or knockdown as evidenced by the CCK8 method and quantification of proliferative rate. *P<0.05, **P<0.01. (GFP-NC: control of empty vector; AEP-OE: AEP-overexpression; NC: negatively control; AEP-KD1: AEP-knocking down 1; AEP-KD2: AEP-knocking down 2) Image collected and cropped by CiteAb from the following publication (https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.8879), licensed under a CC-BY license. Not internally tested by R&D Systems.

Immunocytochemistry/ Immunofluorescence Detection of Human Legumain/Asparaginyl Endopeptidase by Immunocytochemistry/Immunofluorescence View Larger

Detection of Human Legumain/Asparaginyl Endopeptidase by Immunocytochemistry/Immunofluorescence AEP and E-cadherin expression were detected both in primary gastric cancer and peritoneal metastatic lociA. The expression of AEP and E-cadherin in primary gastric cancer and metastatic peritoneal loci were firstly investigated by immunohistochemistry. AEP expression were higher and E-cadherin expression was lower in peritoneal metastatic lesions than in primary gastric cancer (magnified 50× in column 1,3; magnified 200× in column 2,4). B. The expression of AEP and E-cadherin in primary gastric cancer and metastatic peritoneal loci were investigated by immunofluorescence assay (magnified 200×). AEP, the secondary antibody of which were labeled with green fluorescence, was more strongly expressed in peritoneal metastatic lesions than in primary gastric cancer. E-cadherin, the secondary antibody of which was labeled with red fluorescence, was expressed in primary gastric cancer and could not be identified in peritoneal metastatic loci. Image collected and cropped by CiteAb from the following publication (https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.8879), licensed under a CC-BY license. Not internally tested by R&D Systems.

Immunocytochemistry/ Immunofluorescence Detection of Mouse Legumain/Asparaginyl Endopeptidase by Immunocytochemistry/Immunofluorescence View Larger

Detection of Mouse Legumain/Asparaginyl Endopeptidase by Immunocytochemistry/Immunofluorescence Representative sections showing overall higher expression of legumain (diffuse yellowish fluorescence staining) in the untreated (A) versus the Andosan™-treated (B) intestine.Notably, legumain expression was higher in tumor tissue seen in untreated animals. Scale bars represent 200 μm. Image collected and cropped by CiteAb from the following publication (https://dx.plos.org/10.1371/journal.pone.0167754), licensed under a CC-BY license. Not internally tested by R&D Systems.

Western Blot Detection of Mouse Legumain/Asparaginyl Endopeptidase by Western Blot View Larger

Detection of Mouse Legumain/Asparaginyl Endopeptidase by Western Blot AEP knockdown and overexpressive vectors were constructed and stably transfected gastric cancer cell linesA. Constructing of the AEP knocked-down lentiviral vector, it was verified by western blot that AEP was inhibited corresponding to AEP knockdown. B. Overexpressing AEP lentiviral vector was constructed and AEP indeed increased corresponding to AEP overexpression. C. The proliferative curve in response to AEP overexpression or knockdown as evidenced by the CCK8 method and quantification of proliferative rate. *P<0.05, **P<0.01. (GFP-NC: control of empty vector; AEP-OE: AEP-overexpression; NC: negatively control; AEP-KD1: AEP-knocking down 1; AEP-KD2: AEP-knocking down 2) Image collected and cropped by CiteAb from the following publication (https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.8879), licensed under a CC-BY license. Not internally tested by R&D Systems.

Western Blot Detection of Human Legumain/Asparaginyl Endopeptidase by Western Blot View Larger

Detection of Human Legumain/Asparaginyl Endopeptidase by Western Blot Legumain, cathepsin B and L expressions in cytosolic and membrane fractions of myotubes.Myotubes were treated with (+) or without (−) 30 µM simvastatin for 48 h before subcellular fractionation was performed. One representative immunoblot of legumain, cathepsin B and L in the cytosolic and membrane fractions is shown. All lanes were loaded with equal amount of total proteins and probed with antibodies as indicated. LAMP-2 and alpha -tubulin are shown as cell compartment controls (n = 3). Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24416446), licensed under a CC-BY license. Not internally tested by R&D Systems.

Reconstitution Calculator

Reconstitution Calculator

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Preparation and Storage

Reconstitution
Reconstitute at 0.2 mg/mL in sterile PBS.
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Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
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: Legumain/Asparaginyl Endopeptidase

Legumain is a lysosomal cysteine protease whose activity is found in several tissues tested (1, 2). Legumain plays a pivotal role in the endosomal/lysosomal degradation system because the Legumain deficiency causes the accumulation of pro cathepsins B, H and L, another group of lysosomal cysteine proteases (3). Over-expression of Legumain in tumors is significant for invasion/metastasis (4). Also known as Asparaginyl Endopeptidase, it specifically cleaves peptide bonds with Asn at the P1 position. Nevertheless, it also cleaves peptide bonds with Asp at the P1 position. Auto-activation of pro Legumain involves both types of the cleavage, which result in the removal of the pro peptides in both C- and N-termini (5). In addition, Legumain activates pro MMP-2 and processes bacterial antigens for MHC class II presentation and pro thymosin alpha to thymosin alpha 1 and thymosin alpha 11, two acidic peptides with immunoregulatory properties (6‑8). Human Legumain is synthesized as a 433 amino acid precursor with a signal peptide (residues 1‑17). The pro enzyme (residues 18‑433) was expressed with an N-terminal His tag. This activity of Legumain can be inhibited by recombinant human (rh) Cystatin C and rhCystatin E/M and recombinant mouse Cystatin C (R&D Systems, Catalog # 1196‑PI, 1286-PI and 1238-PI, respectively).

References
  1. Chen, J.-M. et al. (1997) J. Biol. Chem. 272:8090.
  2. Tanaka, T. et al. (1996) Cytogenet. Cell Genet. 74:120.
  3. Shirahama-Noda, K. et al. (2003) J. Biol. Chem. 278:33194.
  4. Liu, C. et al. (2003) Cancer Res. 63: 2957.
  5. Li D.N. et al. (2003) J. Biol. Chem. 278:38980.
  6. Chen, J.M. et al. (2001) Biol. Chem. 382:777.
  7. Schwarz, G. et al. (2002) Biol. Chem. 383:1813.
  8. Sarndeses, C.S. et al. (2003) J. Biol. Chem. 278:13286.
Entrez Gene IDs
5641 (Human); 19141 (Mouse)
Alternate Names
AEP; Asparaginyl Endopeptidase; cysteine protease 1; Legumain; LGMN; LGMN1; Protease, cysteine 1; protease, cysteine, 1 (legumain); PRSC1EC 3.4.22.34

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Citations for Human Legumain/Asparaginyl Endopeptidase Antibody

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

17 Citations: Showing 1 - 10
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  1. An infection-induced oxidation site regulates legumain processing and tumor growth
    Authors: Y Kovalyova, DW Bak, EM Gordon, C Fung, JHB Shuman, TL Cover, MR Amieva, E Weerapana, SK Hatzios
    Nature Chemical Biology, 2022-03-24;0(0):.
    Species: Gerbil
    Sample Types: Whole Tissue
    Applications: IHC
  2. Identification of the Cysteine Protease Legumain as a Potential Chronic Hypoxia-Specific Multiple Myeloma Target Gene
    Authors: AS Clees, V Stolp, B Häupl, DC Fuhrmann, F Wempe, M Seibert, S Weber, A Banning, R Tikkanen, R Williams, B Brüne, H Serve, F Schnütgen, I von Metzle, N Kurrle
    Cells, 2022-01-15;11(2):.
    Species: Human
    Sample Types: Cell Lysates, Whole Cells
    Applications: ICC, Western Blot
  3. Neuronal ApoE4 stimulates C/EBPbeta activation, promoting Alzheimer's disease pathology in a mouse model
    Authors: ZH Wang, Y Xia, Z Wu, SS Kang, JC Zhang, P Liu, X Liu, W Song, V Huin, CM Dhaenens, SP Yu, XC Wang, K Ye
    Progress in neurobiology, 2021-12-24;209(0):102212.
    Species: Human
    Sample Types: Cell Lysates, Whole Tissue
    Applications: ChIP, IHC
  4. Asparaginyl endopeptidase (AEP) regulates myocardial apoptosis in response to radiation exposure via alterations in NRF2 activation
    Authors: L Cao, C Xu, P Yi, H Li, Y Lin, G Cai, S Wang, D Ou, M Li, J Chen
    American journal of cancer research, 2021-04-15;11(4):1206-1225.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  5. ApoE4 activates C/EBP&beta/&delta-secretase with 27-hydroxycholesterol, driving the pathogenesis of Alzheimer's disease
    Authors: ZH Wang, Y Xia, P Liu, X Liu, L Edgington-, K Lei, SP Yu, XC Wang, K Ye
    Progress in neurobiology, 2021-03-11;0(0):102032.
    Species: Human
    Sample Types: Cell Lysates, Whole Cells, Whole Tissue
    Applications: ICC, IHC, Western Blot
  6. N368-Tau fragments generated by legumain are detected only in trace amount in the insoluble Tau aggregates isolated from AD brain
    Authors: K Schlegel, K Awwad, RG Heym, D Holzinger, A Doell, S Barghorn, TR Jahn, C Klein, Y Mordashova, M Schulz, L Gasparini
    Acta Neuropathol Commun, 2019-11-13;7(1):177.
    Species: Human, Mouse
    Sample Types: Tissue Homogenates
    Applications: Western Blot
  7. Deficiency in BDNF/TrkB Neurotrophic Activity Stimulates δ-Secretase by Upregulating C/EBP? in Alzheimer's Disease
    Authors: ZH Wang, J Xiang, X Liu, SP Yu, FP Manfredsso, IM Sandoval, S Wu, JZ Wang, K Ye
    Cell Rep, 2019-07-16;28(3):655-669.e5.
    Species: Human
    Sample Types: Tissue Homogenates
    Applications: Western Blot
  8. USP17 Suppresses Tumorigenesis and Tumor Growth through Deubiquitinating AEP
    Authors: X Chen, C Wang, K Liao, S Zhou, L Cao, J Chen, C Xu, Y Lin
    Int. J. Biol. Sci., 2019-01-29;15(4):738-748.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Immunoprecipitation, Western Blot
  9. Overexpression of asparaginyl endopeptidase is significant for esophageal carcinoma metastasis and predicts poor patient prognosis
    Authors: X Liu, Z Wang, G Zhang, Q Zhu, H Zeng, T Wang, F Gao, Z Qi, J Zhang, R Wang
    Oncol Lett, 2017-11-16;15(1):1229-1235.
    Species: Human
    Sample Types: Cell Lysates, Whole Tissue
    Applications: IHC, Western Blot
  10. Glycosylation is important for legumain localization and processing to active forms but not for cystatin E/M inhibitory functions
    Authors: NN Lunde, MH Haugen, KB Bodin Lars, I Damgaard, SJ Pettersen, R Kasem, W Rut, M Drag, M Poreba, HT Johansen, R Solberg
    Biochimie, 2017-05-17;0(0):.
    Species: Human
    Sample Types: Cell Lysates, Whole Cells
    Applications: ICC, Western Blot
  11. Suppression of asparaginyl endopeptidase attenuates breast cancer-induced bone pain through inhibition of neurotrophin receptors
    Authors: P Yao, Y Ding, Z Han, Y Mu, T Hong, Y Zhu, H Li
    Mol Pain, 2017-01-01;13(0):1.74481E+15.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  12. The Agaricus blazei-Based Mushroom Extract, Andosan�, Protects against Intestinal Tumorigenesis in the A/J Min/+ Mouse
    PLoS ONE, 2016-12-21;11(12):e0167754.
    Species: Mouse
    Sample Types: Whole Tissue
    Applications: IHC-P
  13. The asparaginyl endopeptidase legumain is essential for functional recovery after spinal cord injury in adult zebrafish.
    Authors: Ma, Liping, Shen, Yan-Qin, Khatri, Harsh P, Schachner, Melitta
    PLoS ONE, 2014-04-18;9(4):e95098.
    Species: Zebrafish
    Sample Types: Whole Tissue
    Applications: IHC-Fr
  14. Simvastatin inhibits glucose metabolism and legumain activity in human myotubes.
    Authors: Smith R, Solberg R, Jacobsen L, Voreland A, Rustan A, Thoresen G, Johansen H
    PLoS ONE, 2014-01-08;9(1):e85721.
    Species: Human
    Sample Types: Cell Lysates, Whole Cells
    Applications: ICC, Western Blot
  15. Differential secretome analysis reveals CST6 as a suppressor of breast cancer bone metastasis.
    Cell Res., 2012-06-12;22(9):1356-73.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  16. Cystatin E/M suppresses legumain activity and invasion of human melanoma.
    Authors: Briggs JJ, Haugen MH, Johansen HT, Riker AI, Abrahamson M, Fodstad O, Maelandsmo GM, Solberg R
    BMC Cancer, 2010-01-15;10(1):17.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  17. Expression of the cysteine protease legumain in vascular lesions and functional implications in atherogenesis.
    Authors: Clerin V, Shih HH, Deng N, Hebert G, Resmini C, Shields KM, Feldman JL, Winkler A, Albert L, Maganti V, Wong A, Paulsen JE, Keith JC, Vlasuk GP, Pittman DD
    Atherosclerosis, 2008-02-21;0(0):.
    Species: Human
    Sample Types: Cell Lysates, Whole Tissue
    Applications: IHC-P, Western Blot

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