cathepsin l inhibitor ii  (Millipore)


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    Name:
    Cathepsin L Inhibitor
    Description:

    Catalog Number:
    scp0110
    Price:
    None
    Applications:
    Cathepsin L is a lysosomal cysteine proteinase that metabolizes collagens and elastins. The roles and activity of Cathepsin L can be studied with the aid of peptide inhibitors such as the pentapeptide amide RKLLW-NH2 (Arg-Lys-Leu-Leu-Trp-NH2).
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    Structured Review

    Millipore cathepsin l inhibitor ii
    PGRN processing into GRNs is mediated by proper lysosome function and cysteine protease activity. A , HAP1 WT cells were treated for 24 h with the pan-lysosome inhibitors chloroquine (CQ; 50 µM), BafA1 (50 nM), or concanamycin A (ConA; 50 nM) and conditioned media were analyzed for secreted PGRN by ELISA. B , Lysates from HAP1 WT cells treated as in A were analyzed for PGRN and GRN-2,3 by immunoblot. C , D , Quantification of ( C ) PGRN and ( D ) GRN-2,3 from the experiment in B . E , HAP1 WT cells were treated with the indicated protease inhibitors for 24 h and analyzed for PGRN and GRN-2,3 by immunoblot. The inhibitors, their primary targets, and concentrations used are shown in the table at right. F , Time-dependent cleavage of C-TAP PGRN by recombinant <t>cathepsin</t> L in vitro . G , C-TAP PGRN was incubated with or without cathepsin L for 2 h in vitro and analyzed for multiple GRNs by immunoblot. H , HAP1 WT cells were treated with increasing concentrations of cathepsin L inhibitor II (Z-FY-CHO) for 40 h and lysates were analyzed for PGRN and GRN-2,3 by immunoblot. I , J , Quantification of ( I ) PGRN and ( J ) GRN-2,3 from the experiment in H . B , E , H , Arrows denote endogenous, intermediate PGRN cleavage products. For all immunoblots, PGRN and GRN-2,3 were detected with R D AF2420 and Sigma antibodies respectively. All immunoblot images are representative of at least three independent experiments and quantitative data are presented as mean ± SEM of three independent experiments; *differs from control p

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    Average 90 stars, based on 2 article reviews
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    cathepsin l inhibitor ii - by Bioz Stars, 2020-01
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    Images

    1) Product Images from "Intracellular Proteolysis of Progranulin Generates Stable, Lysosomal Granulins that Are Haploinsufficient in Patients with Frontotemporal Dementia Caused by GRN Mutations"

    Article Title: Intracellular Proteolysis of Progranulin Generates Stable, Lysosomal Granulins that Are Haploinsufficient in Patients with Frontotemporal Dementia Caused by GRN Mutations

    Journal: eNeuro

    doi: 10.1523/ENEURO.0100-17.2017

    PGRN processing into GRNs is mediated by proper lysosome function and cysteine protease activity. A , HAP1 WT cells were treated for 24 h with the pan-lysosome inhibitors chloroquine (CQ; 50 µM), BafA1 (50 nM), or concanamycin A (ConA; 50 nM) and conditioned media were analyzed for secreted PGRN by ELISA. B , Lysates from HAP1 WT cells treated as in A were analyzed for PGRN and GRN-2,3 by immunoblot. C , D , Quantification of ( C ) PGRN and ( D ) GRN-2,3 from the experiment in B . E , HAP1 WT cells were treated with the indicated protease inhibitors for 24 h and analyzed for PGRN and GRN-2,3 by immunoblot. The inhibitors, their primary targets, and concentrations used are shown in the table at right. F , Time-dependent cleavage of C-TAP PGRN by recombinant cathepsin L in vitro . G , C-TAP PGRN was incubated with or without cathepsin L for 2 h in vitro and analyzed for multiple GRNs by immunoblot. H , HAP1 WT cells were treated with increasing concentrations of cathepsin L inhibitor II (Z-FY-CHO) for 40 h and lysates were analyzed for PGRN and GRN-2,3 by immunoblot. I , J , Quantification of ( I ) PGRN and ( J ) GRN-2,3 from the experiment in H . B , E , H , Arrows denote endogenous, intermediate PGRN cleavage products. For all immunoblots, PGRN and GRN-2,3 were detected with R D AF2420 and Sigma antibodies respectively. All immunoblot images are representative of at least three independent experiments and quantitative data are presented as mean ± SEM of three independent experiments; *differs from control p
    Figure Legend Snippet: PGRN processing into GRNs is mediated by proper lysosome function and cysteine protease activity. A , HAP1 WT cells were treated for 24 h with the pan-lysosome inhibitors chloroquine (CQ; 50 µM), BafA1 (50 nM), or concanamycin A (ConA; 50 nM) and conditioned media were analyzed for secreted PGRN by ELISA. B , Lysates from HAP1 WT cells treated as in A were analyzed for PGRN and GRN-2,3 by immunoblot. C , D , Quantification of ( C ) PGRN and ( D ) GRN-2,3 from the experiment in B . E , HAP1 WT cells were treated with the indicated protease inhibitors for 24 h and analyzed for PGRN and GRN-2,3 by immunoblot. The inhibitors, their primary targets, and concentrations used are shown in the table at right. F , Time-dependent cleavage of C-TAP PGRN by recombinant cathepsin L in vitro . G , C-TAP PGRN was incubated with or without cathepsin L for 2 h in vitro and analyzed for multiple GRNs by immunoblot. H , HAP1 WT cells were treated with increasing concentrations of cathepsin L inhibitor II (Z-FY-CHO) for 40 h and lysates were analyzed for PGRN and GRN-2,3 by immunoblot. I , J , Quantification of ( I ) PGRN and ( J ) GRN-2,3 from the experiment in H . B , E , H , Arrows denote endogenous, intermediate PGRN cleavage products. For all immunoblots, PGRN and GRN-2,3 were detected with R D AF2420 and Sigma antibodies respectively. All immunoblot images are representative of at least three independent experiments and quantitative data are presented as mean ± SEM of three independent experiments; *differs from control p

    Techniques Used: Activity Assay, Enzyme-linked Immunosorbent Assay, Recombinant, In Vitro, Incubation, Western Blot

    Model of intracellular processing of PGRN into stable, lysosomal GRNs. Sortilin and other receptors target endocytosed and newly synthesized PGRN to lysosomes. Within lysosomes, PGRN is proteolytically cleaved, in part, by cysteine proteases (i.e., cathepsin L) into mature, stable GRN proteins. Ablation of sortilin results in the reduced production of GRNs. Further, lysosome dysfunction caused by alkalizing agents or TMEM106B overexpression inhibits the processing of PGRN into GRNs. FTD- GRN  patients are haploinsufficient for GRNs, which may drive lysosome dysfunction leading to neurodegeneration.
    Figure Legend Snippet: Model of intracellular processing of PGRN into stable, lysosomal GRNs. Sortilin and other receptors target endocytosed and newly synthesized PGRN to lysosomes. Within lysosomes, PGRN is proteolytically cleaved, in part, by cysteine proteases (i.e., cathepsin L) into mature, stable GRN proteins. Ablation of sortilin results in the reduced production of GRNs. Further, lysosome dysfunction caused by alkalizing agents or TMEM106B overexpression inhibits the processing of PGRN into GRNs. FTD- GRN patients are haploinsufficient for GRNs, which may drive lysosome dysfunction leading to neurodegeneration.

    Techniques Used: Synthesized, Over Expression

    2) Product Images from "Intracellular Proteolysis of Progranulin Generates Stable, Lysosomal Granulins that Are Haploinsufficient in Patients with Frontotemporal Dementia Caused by GRN Mutations"

    Article Title: Intracellular Proteolysis of Progranulin Generates Stable, Lysosomal Granulins that Are Haploinsufficient in Patients with Frontotemporal Dementia Caused by GRN Mutations

    Journal: eNeuro

    doi: 10.1523/ENEURO.0100-17.2017

    PGRN processing into GRNs is mediated by proper lysosome function and cysteine protease activity. A , HAP1 WT cells were treated for 24 h with the pan-lysosome inhibitors chloroquine (CQ; 50 µM), BafA1 (50 nM), or concanamycin A (ConA; 50 nM) and conditioned media were analyzed for secreted PGRN by ELISA. B , Lysates from HAP1 WT cells treated as in A were analyzed for PGRN and GRN-2,3 by immunoblot. C , D , Quantification of ( C ) PGRN and ( D ) GRN-2,3 from the experiment in B . E , HAP1 WT cells were treated with the indicated protease inhibitors for 24 h and analyzed for PGRN and GRN-2,3 by immunoblot. The inhibitors, their primary targets, and concentrations used are shown in the table at right. F , Time-dependent cleavage of C-TAP PGRN by recombinant cathepsin L in vitro . G , C-TAP PGRN was incubated with or without cathepsin L for 2 h in vitro and analyzed for multiple GRNs by immunoblot. H , HAP1 WT cells were treated with increasing concentrations of cathepsin L inhibitor II (Z-FY-CHO) for 40 h and lysates were analyzed for PGRN and GRN-2,3 by immunoblot. I , J , Quantification of ( I ) PGRN and ( J ) GRN-2,3 from the experiment in H . B , E , H , Arrows denote endogenous, intermediate PGRN cleavage products. For all immunoblots, PGRN and GRN-2,3 were detected with R D AF2420 and Sigma antibodies respectively. All immunoblot images are representative of at least three independent experiments and quantitative data are presented as mean ± SEM of three independent experiments; *differs from control p
    Figure Legend Snippet: PGRN processing into GRNs is mediated by proper lysosome function and cysteine protease activity. A , HAP1 WT cells were treated for 24 h with the pan-lysosome inhibitors chloroquine (CQ; 50 µM), BafA1 (50 nM), or concanamycin A (ConA; 50 nM) and conditioned media were analyzed for secreted PGRN by ELISA. B , Lysates from HAP1 WT cells treated as in A were analyzed for PGRN and GRN-2,3 by immunoblot. C , D , Quantification of ( C ) PGRN and ( D ) GRN-2,3 from the experiment in B . E , HAP1 WT cells were treated with the indicated protease inhibitors for 24 h and analyzed for PGRN and GRN-2,3 by immunoblot. The inhibitors, their primary targets, and concentrations used are shown in the table at right. F , Time-dependent cleavage of C-TAP PGRN by recombinant cathepsin L in vitro . G , C-TAP PGRN was incubated with or without cathepsin L for 2 h in vitro and analyzed for multiple GRNs by immunoblot. H , HAP1 WT cells were treated with increasing concentrations of cathepsin L inhibitor II (Z-FY-CHO) for 40 h and lysates were analyzed for PGRN and GRN-2,3 by immunoblot. I , J , Quantification of ( I ) PGRN and ( J ) GRN-2,3 from the experiment in H . B , E , H , Arrows denote endogenous, intermediate PGRN cleavage products. For all immunoblots, PGRN and GRN-2,3 were detected with R D AF2420 and Sigma antibodies respectively. All immunoblot images are representative of at least three independent experiments and quantitative data are presented as mean ± SEM of three independent experiments; *differs from control p

    Techniques Used: Activity Assay, Enzyme-linked Immunosorbent Assay, Recombinant, In Vitro, Incubation, Western Blot

    Model of intracellular processing of PGRN into stable, lysosomal GRNs. Sortilin and other receptors target endocytosed and newly synthesized PGRN to lysosomes. Within lysosomes, PGRN is proteolytically cleaved, in part, by cysteine proteases (i.e., cathepsin L) into mature, stable GRN proteins. Ablation of sortilin results in the reduced production of GRNs. Further, lysosome dysfunction caused by alkalizing agents or TMEM106B overexpression inhibits the processing of PGRN into GRNs. FTD- GRN  patients are haploinsufficient for GRNs, which may drive lysosome dysfunction leading to neurodegeneration.
    Figure Legend Snippet: Model of intracellular processing of PGRN into stable, lysosomal GRNs. Sortilin and other receptors target endocytosed and newly synthesized PGRN to lysosomes. Within lysosomes, PGRN is proteolytically cleaved, in part, by cysteine proteases (i.e., cathepsin L) into mature, stable GRN proteins. Ablation of sortilin results in the reduced production of GRNs. Further, lysosome dysfunction caused by alkalizing agents or TMEM106B overexpression inhibits the processing of PGRN into GRNs. FTD- GRN patients are haploinsufficient for GRNs, which may drive lysosome dysfunction leading to neurodegeneration.

    Techniques Used: Synthesized, Over Expression

    3) Product Images from "Intracellular Proteolysis of Progranulin Generates Stable, Lysosomal Granulins that Are Haploinsufficient in Patients with Frontotemporal Dementia Caused by GRN Mutations"

    Article Title: Intracellular Proteolysis of Progranulin Generates Stable, Lysosomal Granulins that Are Haploinsufficient in Patients with Frontotemporal Dementia Caused by GRN Mutations

    Journal: eNeuro

    doi: 10.1523/ENEURO.0100-17.2017

    PGRN processing into GRNs is mediated by proper lysosome function and cysteine protease activity. A , HAP1 WT cells were treated for 24 h with the pan-lysosome inhibitors chloroquine (CQ; 50 µM), BafA1 (50 nM), or concanamycin A (ConA; 50 nM) and conditioned media were analyzed for secreted PGRN by ELISA. B , Lysates from HAP1 WT cells treated as in A were analyzed for PGRN and GRN-2,3 by immunoblot. C , D , Quantification of ( C ) PGRN and ( D ) GRN-2,3 from the experiment in B . E , HAP1 WT cells were treated with the indicated protease inhibitors for 24 h and analyzed for PGRN and GRN-2,3 by immunoblot. The inhibitors, their primary targets, and concentrations used are shown in the table at right. F , Time-dependent cleavage of C-TAP PGRN by recombinant cathepsin L in vitro . G , C-TAP PGRN was incubated with or without cathepsin L for 2 h in vitro and analyzed for multiple GRNs by immunoblot. H , HAP1 WT cells were treated with increasing concentrations of cathepsin L inhibitor II (Z-FY-CHO) for 40 h and lysates were analyzed for PGRN and GRN-2,3 by immunoblot. I , J , Quantification of ( I ) PGRN and ( J ) GRN-2,3 from the experiment in H . B , E , H , Arrows denote endogenous, intermediate PGRN cleavage products. For all immunoblots, PGRN and GRN-2,3 were detected with R D AF2420 and Sigma antibodies respectively. All immunoblot images are representative of at least three independent experiments and quantitative data are presented as mean ± SEM of three independent experiments; *differs from control p
    Figure Legend Snippet: PGRN processing into GRNs is mediated by proper lysosome function and cysteine protease activity. A , HAP1 WT cells were treated for 24 h with the pan-lysosome inhibitors chloroquine (CQ; 50 µM), BafA1 (50 nM), or concanamycin A (ConA; 50 nM) and conditioned media were analyzed for secreted PGRN by ELISA. B , Lysates from HAP1 WT cells treated as in A were analyzed for PGRN and GRN-2,3 by immunoblot. C , D , Quantification of ( C ) PGRN and ( D ) GRN-2,3 from the experiment in B . E , HAP1 WT cells were treated with the indicated protease inhibitors for 24 h and analyzed for PGRN and GRN-2,3 by immunoblot. The inhibitors, their primary targets, and concentrations used are shown in the table at right. F , Time-dependent cleavage of C-TAP PGRN by recombinant cathepsin L in vitro . G , C-TAP PGRN was incubated with or without cathepsin L for 2 h in vitro and analyzed for multiple GRNs by immunoblot. H , HAP1 WT cells were treated with increasing concentrations of cathepsin L inhibitor II (Z-FY-CHO) for 40 h and lysates were analyzed for PGRN and GRN-2,3 by immunoblot. I , J , Quantification of ( I ) PGRN and ( J ) GRN-2,3 from the experiment in H . B , E , H , Arrows denote endogenous, intermediate PGRN cleavage products. For all immunoblots, PGRN and GRN-2,3 were detected with R D AF2420 and Sigma antibodies respectively. All immunoblot images are representative of at least three independent experiments and quantitative data are presented as mean ± SEM of three independent experiments; *differs from control p

    Techniques Used: Activity Assay, Enzyme-linked Immunosorbent Assay, Recombinant, In Vitro, Incubation, Western Blot

    Model of intracellular processing of PGRN into stable, lysosomal GRNs. Sortilin and other receptors target endocytosed and newly synthesized PGRN to lysosomes. Within lysosomes, PGRN is proteolytically cleaved, in part, by cysteine proteases (i.e., cathepsin L) into mature, stable GRN proteins. Ablation of sortilin results in the reduced production of GRNs. Further, lysosome dysfunction caused by alkalizing agents or TMEM106B overexpression inhibits the processing of PGRN into GRNs. FTD- GRN  patients are haploinsufficient for GRNs, which may drive lysosome dysfunction leading to neurodegeneration.
    Figure Legend Snippet: Model of intracellular processing of PGRN into stable, lysosomal GRNs. Sortilin and other receptors target endocytosed and newly synthesized PGRN to lysosomes. Within lysosomes, PGRN is proteolytically cleaved, in part, by cysteine proteases (i.e., cathepsin L) into mature, stable GRN proteins. Ablation of sortilin results in the reduced production of GRNs. Further, lysosome dysfunction caused by alkalizing agents or TMEM106B overexpression inhibits the processing of PGRN into GRNs. FTD- GRN patients are haploinsufficient for GRNs, which may drive lysosome dysfunction leading to neurodegeneration.

    Techniques Used: Synthesized, Over Expression

    4) Product Images from "Inhibition of cathepsin L sensitizes human glioma cells to ionizing radiation in vitro through NF-κB signaling pathway"

    Article Title: Inhibition of cathepsin L sensitizes human glioma cells to ionizing radiation in vitro through NF-κB signaling pathway

    Journal: Acta Pharmacologica Sinica

    doi: 10.1038/aps.2014.148

    Altered IR-induced nuclear translocation of NF-κB following selective inhibition of cathepsin L with Z-FY-CHO or cathepsin L shRNA. (A) p65 protein immunofluorescence in U251 cells treated with Z-FY-CHO, IR, or both. (B) p65 protein immunofluorescence
    Figure Legend Snippet: Altered IR-induced nuclear translocation of NF-κB following selective inhibition of cathepsin L with Z-FY-CHO or cathepsin L shRNA. (A) p65 protein immunofluorescence in U251 cells treated with Z-FY-CHO, IR, or both. (B) p65 protein immunofluorescence

    Techniques Used: Translocation Assay, Inhibition, shRNA, Immunofluorescence

    Reduced NF-κB-dependent transcriptional activation in U251 cells following IR with selective inhibition of cathepsin L with Z-FY-CHO (A) or cathepsin L shRNA (B). NF-κB activity was assessed using an NF-κB-RE firefly luciferase
    Figure Legend Snippet: Reduced NF-κB-dependent transcriptional activation in U251 cells following IR with selective inhibition of cathepsin L with Z-FY-CHO (A) or cathepsin L shRNA (B). NF-κB activity was assessed using an NF-κB-RE firefly luciferase

    Techniques Used: Activation Assay, Inhibition, shRNA, Activity Assay, Luciferase

    The inhibition of cathepsin L sensitized U251 cells to IR. (A) Clonogenic survival curves for U251 cells treated with radiation (0, 2, 4, 6, and 8 Gy) alone or in combination with Z-FY-CHO. (B) Western blot of CTSL in whole cell extracts from parental
    Figure Legend Snippet: The inhibition of cathepsin L sensitized U251 cells to IR. (A) Clonogenic survival curves for U251 cells treated with radiation (0, 2, 4, 6, and 8 Gy) alone or in combination with Z-FY-CHO. (B) Western blot of CTSL in whole cell extracts from parental

    Techniques Used: Inhibition, Western Blot

    Altered IR-induced nuclear translocation of NF-κB following selective inhibition of cathepsin L with Z-FY-CHO or cathepsin L shRNA. (A) Western blot analysis of IκBα expression and the nuclear translocation of NF-κB in
    Figure Legend Snippet: Altered IR-induced nuclear translocation of NF-κB following selective inhibition of cathepsin L with Z-FY-CHO or cathepsin L shRNA. (A) Western blot analysis of IκBα expression and the nuclear translocation of NF-κB in

    Techniques Used: Translocation Assay, Inhibition, shRNA, Western Blot, Expressing

    The expression of cathepsin L protein and nuclear translocation of NF-κB p65 and p50 in U251 cells following IR was analyzed by Western blot. (A) Western blot analysis of cathepsin L in U251 cells treated with IR (10 Gy). β-Actin was used
    Figure Legend Snippet: The expression of cathepsin L protein and nuclear translocation of NF-κB p65 and p50 in U251 cells following IR was analyzed by Western blot. (A) Western blot analysis of cathepsin L in U251 cells treated with IR (10 Gy). β-Actin was used

    Techniques Used: Expressing, Translocation Assay, Western Blot

    Effect of p65 shRNA on the expression of p65 and cathepsin L in U251 and MCF-7 cells. (A) Western blot of p65 whole cell extracts from normal U251, U251-consh and U251-p65sh cells. β-Actin was used as an internal control. c P
    Figure Legend Snippet: Effect of p65 shRNA on the expression of p65 and cathepsin L in U251 and MCF-7 cells. (A) Western blot of p65 whole cell extracts from normal U251, U251-consh and U251-p65sh cells. β-Actin was used as an internal control. c P

    Techniques Used: shRNA, Expressing, Western Blot

    5) Product Images from "Cathepsin L Mediates the Degradation of Novel APP C-Terminal Fragments"

    Article Title: Cathepsin L Mediates the Degradation of Novel APP C-Terminal Fragments

    Journal: Biochemistry

    doi: 10.1021/acs.biochem.5b00329

    Accumulation of novel APP CTFs was mediated by cathepsin L inhibition. (A) Naïve HEK293 cells were treated with vehicle or MDL28170 (20 or 40 μ M) for 24 h before being collected. The Western blot was probed with C1/6.1 and anti-actin antibody.
    Figure Legend Snippet: Accumulation of novel APP CTFs was mediated by cathepsin L inhibition. (A) Naïve HEK293 cells were treated with vehicle or MDL28170 (20 or 40 μ M) for 24 h before being collected. The Western blot was probed with C1/6.1 and anti-actin antibody.

    Techniques Used: Inhibition, Western Blot

    Related Articles

    Transduction:

    Article Title: Identification of a Broad-Spectrum Antiviral Small Molecule against Severe Acute Respiratory Syndrome Coronavirus and Ebola, Hendra, and Nipah Viruses by Using a Novel High-Throughput Screening Assay
    Article Snippet: Twenty-four hours later, the cells were transduced with the SARS-CoV-S pseudotyped virus. .. Camostat mesylate was used as a TMPRSS2 protease inhibitor (Tocris Bioscience, Bristol, United Kingdom) and Z-FY( t -Bu)-DMK as a CatL inhibitor (cathepsin L inhibitor III; Calbiochem catalog no. 219427).

    Article Title: Functional analysis of potential cleavage sites in the MERS-coronavirus spike protein
    Article Snippet: .. Protease inhibitors To test which proteases facilitate MERS-S activation, target cells were preincubated with cathepsin L inhibitor (MDL28170; Sigma-Aldrich) or serine protease inhibitor (Camostat, Sigma-Aldrich) for 2 h, before the cells were inoculated with transduction vectors. .. Antibodies The following antibodies were used as primary antibodies: Anti V5 (Invitrogen), anti β-actin (Sigma-Aldrich), anti VSV-M (Kerafast).

    Article Title: Functional analysis of potential cleavage sites in the MERS-coronavirus spike protein
    Article Snippet: .. To test which proteases facilitate MERS-S activation, target cells were preincubated with cathepsin L inhibitor (MDL28170; Sigma-Aldrich) or serine protease inhibitor (Camostat, Sigma-Aldrich) for 2 h, before the cells were inoculated with transduction vectors. .. The following antibodies were used as primary antibodies: Anti V5 (Invitrogen), anti β-actin (Sigma-Aldrich), anti VSV-M (Kerafast).

    Luciferase:

    Article Title: Identification of a Broad-Spectrum Antiviral Small Molecule against Severe Acute Respiratory Syndrome Coronavirus and Ebola, Hendra, and Nipah Viruses by Using a Novel High-Throughput Screening Assay
    Article Snippet: At 72 h postransduction, the entry was quantified by measuring the luciferase expression, as relative light units (RLU), in cell lysates. .. Camostat mesylate was used as a TMPRSS2 protease inhibitor (Tocris Bioscience, Bristol, United Kingdom) and Z-FY( t -Bu)-DMK as a CatL inhibitor (cathepsin L inhibitor III; Calbiochem catalog no. 219427).

    Cytometry:

    Article Title: Immuno-evasive tactics by schistosomes identifies an effective allergy preventative
    Article Snippet: Protease inhibitors including, leupeptin, CA-074, and cathepsin L inhibitor, were used in cultures at 5 µg/ml (Calbiochem-EMD Millipore, Billerica, MA). .. Following treatment, leukocytes were evaluated for surface levels of CD23 by flow cytometry (anti-CD23 PE) and cell-free supernatants were collected and stored at −20°C until evaluated.

    Blocking Assay:

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    Article Snippet: Protease inhibitors including, leupeptin, CA-074, and cathepsin L inhibitor, were used in cultures at 5 µg/ml (Calbiochem-EMD Millipore, Billerica, MA). .. To block schistosomal calpain activity, cultures were treated with heat-inactivated hyper-immune anti-Sm-p80 serum.

    Incubation:

    Article Title: Combined In Vivo Molecular and Anatomic Imaging for Detection of Ovarian Carcinoma-Associated Protease Activity and Integrin Expression in Mice 1 Molecular and Anatomic Imaging for Detection of Ovarian Carcinoma-Associated Protease Activity and Integrin Expression in Mice 1 2
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    Article Snippet: For cell migration and invasion assays, cells were seeded in the presence of the cathepsin D inhibitor (Pepstatin A, 100 μM, Calbiochem), the cathepsin L inhibitor (cathepsin L inhibitor II, Z-FY-CHO, 10 μM, Calbiochem) as well as two cathepsin B inhibitors: CA-074 (cell unpermeant) and CA-074Me (cell permeant) (both given at a concentration of 10 μM, Calbiochem, Nottingham, UK). .. Alternatively to these synthetic inhibitors, cells were also incubated with specific antibodies to cathepsin B, D (both Calbiochem) or L (Alexis) (60 μg/ml).

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    Article Snippet: Protein samples (25 μg) were incubated with 100 μM fluorogenic substrate (Z-Phe-Arg-AMC; Peptides International) in 100 mM sodium acetate buffer containing 1 mM EDTA and 1 mM DTT, pH 5.5. .. Released AMC was measured using a Fluoroskan Ascent fluorometer at an excitation wavelength of 390 nm and an emission wavelength of 460 nm for up to 120 min. Each assay was conducted in the absence and presence of a specific cathepsin L inhibitor (10 μM cathepsin L inhibitor I; Calbiochem) to determine cathepsin L-specific activity.

    Activity Assay:

    Article Title: Immuno-evasive tactics by schistosomes identifies an effective allergy preventative
    Article Snippet: Protease inhibitors including, leupeptin, CA-074, and cathepsin L inhibitor, were used in cultures at 5 µg/ml (Calbiochem-EMD Millipore, Billerica, MA). .. To block schistosomal calpain activity, cultures were treated with heat-inactivated hyper-immune anti-Sm-p80 serum.

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    Expressing:

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    Modification:

    Article Title: High-density lipoprotein inhibits serum amyloid A–mediated reactive oxygen species generation and NLRP3 inflammasome activation
    Article Snippet: Murine J774.2 macrophage-like cells were obtained from Sigma and maintained in complete medium (Dulbecco's modified Eagle's medium (DMEM), supplemented with 10% fetal bovine serum, 100 units/ml of penicillin/streptomycin, 2 m m l -glutamine). .. For LPS (Innaxon, catalog number IAX-100-012-5001) and ATP (Sigma) treatment, the cells were incubated with LPS (0.5 μg/ml) for 3 h in SFM followed by treatment with ATP (3 m m ) for 45 min. For LPS and alum (Imject alum adjuvant; a mixture of aluminum hydroxide and magnesium hydroxide, Pierce) treatment, the cells were incubated with LPS (0.5 μg/ml) for 3 h in SFM followed by treatment with alum (200 μg/ml) for 4 h. Cathepsin B inhibitor, CA-074-Me (Sigma), caspase-1 inhibitor, Z-YVAD-fmk (Vergent Bioscience), cathepsin L inhibitor (Calbiochem), cytochalasin D (Sigma), glyburide (Sigma), and the P2X7R antagonists, A-438079 hydrochloride (Sigma) and AZ-10606120 (TOCRIS) were dissolved in DMSO, and used at the indicated concentrations.

    Article Title: Immuno-evasive tactics by schistosomes identifies an effective allergy preventative
    Article Snippet: Live worms were cultured with human cells at concentrations of one to 20 worms to 1 million cells in the modified Basch media and both cells and worms remained viable for up to four days. .. Protease inhibitors including, leupeptin, CA-074, and cathepsin L inhibitor, were used in cultures at 5 µg/ml (Calbiochem-EMD Millipore, Billerica, MA).

    Derivative Assay:

    Article Title: Cathepsin B inhibition interferes with metastatic potential of human melanoma: an in vitro and in vivo study
    Article Snippet: The LP (PM2) cell line derived from a primary cutaneous melanoma (Clark's level V; Breslow 12 mm), while the LM cell line (MM2) from a supraclavicular lymph node metastasis of the same patient [ ]. .. For cell migration and invasion assays, cells were seeded in the presence of the cathepsin D inhibitor (Pepstatin A, 100 μM, Calbiochem), the cathepsin L inhibitor (cathepsin L inhibitor II, Z-FY-CHO, 10 μM, Calbiochem) as well as two cathepsin B inhibitors: CA-074 (cell unpermeant) and CA-074Me (cell permeant) (both given at a concentration of 10 μM, Calbiochem, Nottingham, UK).

    Transfection:

    Article Title: Identification of a Broad-Spectrum Antiviral Small Molecule against Severe Acute Respiratory Syndrome Coronavirus and Ebola, Hendra, and Nipah Viruses by Using a Novel High-Throughput Screening Assay
    Article Snippet: The entry of SARS-CoV-S pseudotyped virus into 293FT cells, transfected with either the receptor alone (ACE2 plasmid) or the receptor plus 10 ng of TMPRSS2 plasmid, was measured in the presence or absence of different inhibitors. .. Camostat mesylate was used as a TMPRSS2 protease inhibitor (Tocris Bioscience, Bristol, United Kingdom) and Z-FY( t -Bu)-DMK as a CatL inhibitor (cathepsin L inhibitor III; Calbiochem catalog no. 219427).

    Activation Assay:

    Article Title: Functional analysis of potential cleavage sites in the MERS-coronavirus spike protein
    Article Snippet: .. Protease inhibitors To test which proteases facilitate MERS-S activation, target cells were preincubated with cathepsin L inhibitor (MDL28170; Sigma-Aldrich) or serine protease inhibitor (Camostat, Sigma-Aldrich) for 2 h, before the cells were inoculated with transduction vectors. .. Antibodies The following antibodies were used as primary antibodies: Anti V5 (Invitrogen), anti β-actin (Sigma-Aldrich), anti VSV-M (Kerafast).

    Article Title: Functional analysis of potential cleavage sites in the MERS-coronavirus spike protein
    Article Snippet: .. To test which proteases facilitate MERS-S activation, target cells were preincubated with cathepsin L inhibitor (MDL28170; Sigma-Aldrich) or serine protease inhibitor (Camostat, Sigma-Aldrich) for 2 h, before the cells were inoculated with transduction vectors. .. The following antibodies were used as primary antibodies: Anti V5 (Invitrogen), anti β-actin (Sigma-Aldrich), anti VSV-M (Kerafast).

    Protease Inhibitor:

    Article Title: Identification of a Broad-Spectrum Antiviral Small Molecule against Severe Acute Respiratory Syndrome Coronavirus and Ebola, Hendra, and Nipah Viruses by Using a Novel High-Throughput Screening Assay
    Article Snippet: .. Camostat mesylate was used as a TMPRSS2 protease inhibitor (Tocris Bioscience, Bristol, United Kingdom) and Z-FY( t -Bu)-DMK as a CatL inhibitor (cathepsin L inhibitor III; Calbiochem catalog no. 219427). .. All inhibitors were used at a 10 μM concentration except the small molecule 5705213, which was used at a 50 μM final concentration.

    Article Title: Functional analysis of potential cleavage sites in the MERS-coronavirus spike protein
    Article Snippet: .. Protease inhibitors To test which proteases facilitate MERS-S activation, target cells were preincubated with cathepsin L inhibitor (MDL28170; Sigma-Aldrich) or serine protease inhibitor (Camostat, Sigma-Aldrich) for 2 h, before the cells were inoculated with transduction vectors. .. Antibodies The following antibodies were used as primary antibodies: Anti V5 (Invitrogen), anti β-actin (Sigma-Aldrich), anti VSV-M (Kerafast).

    Article Title: Functional analysis of potential cleavage sites in the MERS-coronavirus spike protein
    Article Snippet: .. To test which proteases facilitate MERS-S activation, target cells were preincubated with cathepsin L inhibitor (MDL28170; Sigma-Aldrich) or serine protease inhibitor (Camostat, Sigma-Aldrich) for 2 h, before the cells were inoculated with transduction vectors. .. The following antibodies were used as primary antibodies: Anti V5 (Invitrogen), anti β-actin (Sigma-Aldrich), anti VSV-M (Kerafast).

    Cell Culture:

    Article Title: Palmitate impairs angiogenesis via suppression of cathepsin activity
    Article Snippet: Paragraph title: Cell culture and incubation with fatty acids ... If not stated otherwise, cells were incubated for 1 h with 10 µΜ cathepsin L inhibitor (z-FF-FMK; cat. no. 219421; Calbiochem; EMD Millipore, Billerica, MA, USA) and cathepsin S inhibitor (z-FL-COCHO.H2 O; cat. no. 219393; Calbiochem; EMD Millipore) at 37°C, which was followed by incubation with palmitate or FFA-free BSA for 24 h at 37°C.

    Article Title: Immuno-evasive tactics by schistosomes identifies an effective allergy preventative
    Article Snippet: Live worms were cultured with human cells at concentrations of one to 20 worms to 1 million cells in the modified Basch media and both cells and worms remained viable for up to four days. .. Protease inhibitors including, leupeptin, CA-074, and cathepsin L inhibitor, were used in cultures at 5 µg/ml (Calbiochem-EMD Millipore, Billerica, MA).

    Article Title: Caspase-3-independent apoptotic pathways contribute to interleukin-32γ-mediated control of Mycobacterium tuberculosis infection in THP-1 cells
    Article Snippet: THP-1 cells were cultured in RPMI 1640 supplemented with 10% FBS and 2 mM glutamine and were maintained at a concentration of 2–10 ×105 cells/mL. .. The cathepsin B inhibitor [L-3-trans- (propylcarbamoyl) oxirane-2-carbonyl]-L-isoleucyl-L-proline methyl ester (CA-074-Me)], the cathepsin D inhibitor pepstatin A, and the cathepsin L inhibitor [benzyloxycarbonyl-Leu-Leu-Tyr-fluoromethylketone (z-LLY-fmk)] were purchased from Calbiochem-EMD Millipore (Billerica, Massachusetts).

    Generated:

    Article Title: Immuno-evasive tactics by schistosomes identifies an effective allergy preventative
    Article Snippet: Protease inhibitors including, leupeptin, CA-074, and cathepsin L inhibitor, were used in cultures at 5 µg/ml (Calbiochem-EMD Millipore, Billerica, MA). .. Recombinant (r) calpain/Sm-p80 and hyper-immune mouse sera to Sm-p80 were generated as previously described ( , ).

    other:

    Article Title: Cardiac Glycoside Glucoevatromonoside Induces Cancer Type-Specific Cell Death
    Article Snippet: Etoposide, 3-aminobenzamide (3-ABA), cathepsin L inhibitor, and bafilomycin A1 were from Sigma-Aldrich (Bornem, Belgium). z-VAD-FMK (z-VAD), necrostatin (Nec)-1, and calpain inhibitor PD150606 were from Calbiochem (Leuven, Belgium).

    Binding Assay:

    Article Title: Combined In Vivo Molecular and Anatomic Imaging for Detection of Ovarian Carcinoma-Associated Protease Activity and Integrin Expression in Mice 1 Molecular and Anatomic Imaging for Detection of Ovarian Carcinoma-Associated Protease Activity and Integrin Expression in Mice 1 2
    Article Snippet: Parallel cultures of MOVCAR 5009 cells were grown in the presence and absence of cathepsin protease inhibitors (cathepsin B inhibitor III [CA-074] or cathepsin L inhibitor [CAA0225]; EMD Chemicals, Inc, Gibbstown, NJ) and MMP inhibitors (ARP-100; Tocris Bioscience, Bristol, United Kingdom). .. For detection of both ProSense 680 and MMPSense 680, 3 x 104 cells were seeded on fibronectin-coated coverslips in 24-well plates for 4 hours and treated with 1 µg/ml saponin to facilitate binding and detection of hydrophobic probes.

    Flow Cytometry:

    Article Title: Immuno-evasive tactics by schistosomes identifies an effective allergy preventative
    Article Snippet: Protease inhibitors including, leupeptin, CA-074, and cathepsin L inhibitor, were used in cultures at 5 µg/ml (Calbiochem-EMD Millipore, Billerica, MA). .. Following treatment, leukocytes were evaluated for surface levels of CD23 by flow cytometry (anti-CD23 PE) and cell-free supernatants were collected and stored at −20°C until evaluated.

    Purification:

    Article Title: High-density lipoprotein inhibits serum amyloid A–mediated reactive oxygen species generation and NLRP3 inflammasome activation
    Article Snippet: For experiments with SAA, J774.2 cells were incubated with 5–50 μg/ml of purified mouse or human SAA in serum-free DMEM containing 0.2% fatty acid-free BSA (SFM) for 8–24 h. For experiments with SAA and HDL, SAA and HDL were co-incubated at a ratio of 1:2.8 (protein:protein) for 1 h at room temperature prior to treating the cells. .. For LPS (Innaxon, catalog number IAX-100-012-5001) and ATP (Sigma) treatment, the cells were incubated with LPS (0.5 μg/ml) for 3 h in SFM followed by treatment with ATP (3 m m ) for 45 min. For LPS and alum (Imject alum adjuvant; a mixture of aluminum hydroxide and magnesium hydroxide, Pierce) treatment, the cells were incubated with LPS (0.5 μg/ml) for 3 h in SFM followed by treatment with alum (200 μg/ml) for 4 h. Cathepsin B inhibitor, CA-074-Me (Sigma), caspase-1 inhibitor, Z-YVAD-fmk (Vergent Bioscience), cathepsin L inhibitor (Calbiochem), cytochalasin D (Sigma), glyburide (Sigma), and the P2X7R antagonists, A-438079 hydrochloride (Sigma) and AZ-10606120 (TOCRIS) were dissolved in DMSO, and used at the indicated concentrations.

    Article Title: Immuno-evasive tactics by schistosomes identifies an effective allergy preventative
    Article Snippet: To determine the effect of S. mansoni on CD23 levels, mixed mononuclear cells, purified B cells, or immortalized cell lines were treated with schistosomula E/S (1:1 E/S to cell culture media) or 2 µg/ml of schistosomula crude somatic antigens, SEA, or SWAP from 5 minutes to 48 hours. .. Protease inhibitors including, leupeptin, CA-074, and cathepsin L inhibitor, were used in cultures at 5 µg/ml (Calbiochem-EMD Millipore, Billerica, MA).

    Article Title: Rotaviruses Reach Late Endosomes and Require the Cation-Dependent Mannose-6-Phosphate Receptor and the Activity of Cathepsin Proteases To Enter the Cell
    Article Snippet: Rabbit hyperimmune sera to purified RV particles (anti-TLPs), RV nonstructural protein 5 (anti-NSP5), and purified reovirus particles (anti-Reo) were produced in our laboratory. .. Cathepsin L inhibitor (Z-FF-FMK) and dimethyl sulfoxide (DMSO) were purchased from Calbiochem, Merck KGaA (Darmstadt, Germany).

    Plasmid Preparation:

    Article Title: Identification of a Broad-Spectrum Antiviral Small Molecule against Severe Acute Respiratory Syndrome Coronavirus and Ebola, Hendra, and Nipah Viruses by Using a Novel High-Throughput Screening Assay
    Article Snippet: The entry of SARS-CoV-S pseudotyped virus into 293FT cells, transfected with either the receptor alone (ACE2 plasmid) or the receptor plus 10 ng of TMPRSS2 plasmid, was measured in the presence or absence of different inhibitors. .. Camostat mesylate was used as a TMPRSS2 protease inhibitor (Tocris Bioscience, Bristol, United Kingdom) and Z-FY( t -Bu)-DMK as a CatL inhibitor (cathepsin L inhibitor III; Calbiochem catalog no. 219427).

    In Vitro:

    Article Title: Combined In Vivo Molecular and Anatomic Imaging for Detection of Ovarian Carcinoma-Associated Protease Activity and Integrin Expression in Mice 1 Molecular and Anatomic Imaging for Detection of Ovarian Carcinoma-Associated Protease Activity and Integrin Expression in Mice 1 2
    Article Snippet: Paragraph title: In Vitro Detection of Optical Imaging Probes ... Parallel cultures of MOVCAR 5009 cells were grown in the presence and absence of cathepsin protease inhibitors (cathepsin B inhibitor III [CA-074] or cathepsin L inhibitor [CAA0225]; EMD Chemicals, Inc, Gibbstown, NJ) and MMP inhibitors (ARP-100; Tocris Bioscience, Bristol, United Kingdom).

    Article Title: Cathepsin B inhibition interferes with metastatic potential of human melanoma: an in vitro and in vivo study
    Article Snippet: Paragraph title: Melanoma cell cultures and in vitro treatments ... For cell migration and invasion assays, cells were seeded in the presence of the cathepsin D inhibitor (Pepstatin A, 100 μM, Calbiochem), the cathepsin L inhibitor (cathepsin L inhibitor II, Z-FY-CHO, 10 μM, Calbiochem) as well as two cathepsin B inhibitors: CA-074 (cell unpermeant) and CA-074Me (cell permeant) (both given at a concentration of 10 μM, Calbiochem, Nottingham, UK).

    Negative Control:

    Article Title: Identification of a Broad-Spectrum Antiviral Small Molecule against Severe Acute Respiratory Syndrome Coronavirus and Ebola, Hendra, and Nipah Viruses by Using a Novel High-Throughput Screening Assay
    Article Snippet: HIV/ΔE, which does not express the SARS-CoV-S protein, was used as a negative control. .. Camostat mesylate was used as a TMPRSS2 protease inhibitor (Tocris Bioscience, Bristol, United Kingdom) and Z-FY( t -Bu)-DMK as a CatL inhibitor (cathepsin L inhibitor III; Calbiochem catalog no. 219427).

    Recombinant:

    Article Title: Immuno-evasive tactics by schistosomes identifies an effective allergy preventative
    Article Snippet: Protease inhibitors including, leupeptin, CA-074, and cathepsin L inhibitor, were used in cultures at 5 µg/ml (Calbiochem-EMD Millipore, Billerica, MA). .. Recombinant (r) calpain/Sm-p80 and hyper-immune mouse sera to Sm-p80 were generated as previously described ( , ).

    Article Title: Caspase-3-independent apoptotic pathways contribute to interleukin-32γ-mediated control of Mycobacterium tuberculosis infection in THP-1 cells
    Article Snippet: Recombinant IL-32γ (confirmed to be LPS-free) and caspase-1 inhibitor (z-WEHD-fmk) were purchased from R & D System, Inc. (Minneapolis, MN). .. The cathepsin B inhibitor [L-3-trans- (propylcarbamoyl) oxirane-2-carbonyl]-L-isoleucyl-L-proline methyl ester (CA-074-Me)], the cathepsin D inhibitor pepstatin A, and the cathepsin L inhibitor [benzyloxycarbonyl-Leu-Leu-Tyr-fluoromethylketone (z-LLY-fmk)] were purchased from Calbiochem-EMD Millipore (Billerica, Massachusetts).

    In Situ:

    Article Title: Caspase-3-independent apoptotic pathways contribute to interleukin-32γ-mediated control of Mycobacterium tuberculosis infection in THP-1 cells
    Article Snippet: Apoptosis in Situ Detection Kit was purchased from Roche Diagnostic Systems (Indianapolis, IN). .. The cathepsin B inhibitor [L-3-trans- (propylcarbamoyl) oxirane-2-carbonyl]-L-isoleucyl-L-proline methyl ester (CA-074-Me)], the cathepsin D inhibitor pepstatin A, and the cathepsin L inhibitor [benzyloxycarbonyl-Leu-Leu-Tyr-fluoromethylketone (z-LLY-fmk)] were purchased from Calbiochem-EMD Millipore (Billerica, Massachusetts).

    Produced:

    Article Title: Rotaviruses Reach Late Endosomes and Require the Cation-Dependent Mannose-6-Phosphate Receptor and the Activity of Cathepsin Proteases To Enter the Cell
    Article Snippet: Rabbit hyperimmune sera to purified RV particles (anti-TLPs), RV nonstructural protein 5 (anti-NSP5), and purified reovirus particles (anti-Reo) were produced in our laboratory. .. Cathepsin L inhibitor (Z-FF-FMK) and dimethyl sulfoxide (DMSO) were purchased from Calbiochem, Merck KGaA (Darmstadt, Germany).

    Concentration Assay:

    Article Title: Identification of a Broad-Spectrum Antiviral Small Molecule against Severe Acute Respiratory Syndrome Coronavirus and Ebola, Hendra, and Nipah Viruses by Using a Novel High-Throughput Screening Assay
    Article Snippet: Camostat mesylate was used as a TMPRSS2 protease inhibitor (Tocris Bioscience, Bristol, United Kingdom) and Z-FY( t -Bu)-DMK as a CatL inhibitor (cathepsin L inhibitor III; Calbiochem catalog no. 219427). .. All inhibitors were used at a 10 μM concentration except the small molecule 5705213, which was used at a 50 μM final concentration.

    Article Title: Curcumin induces crosstalk between autophagy and apoptosis mediated by calcium release from the endoplasmic reticulum, lysosomal destabilization and mitochondrial events
    Article Snippet: .. The inhibitors used were cathepsin B inhibitor (CA074-Me, 50 μ M, Sigma-Aldrich SARL, Saint-Quentin Fallavier, France), cathepsin G inhibitor 1 (10 μ M, Calbiochem, Merck Chimie SAS, Fontenay sous Bois, France), cathepsin L inhibitor (10 μ M Z-FF-FMK, Calbiochem, Merck Chimie SAS), and cathepsin D inhibitor (10 μ M, pepstatin A, Sigma-Aldrich SARL) dissolved, in most cases, in DMSO (concentration±0.05%). .. In addition, 3-(4-Iodophenyl)-2-mercapto-(Z)-2-propenoic acid (PD 150606), a cell-permeable noncompetitive inhibitor of calpains 1 and 2, was provided by Calbiochem (Merck Chimie SAS).

    Article Title: High-density lipoprotein inhibits serum amyloid A–mediated reactive oxygen species generation and NLRP3 inflammasome activation
    Article Snippet: For LPS (Innaxon, catalog number IAX-100-012-5001) and ATP (Sigma) treatment, the cells were incubated with LPS (0.5 μg/ml) for 3 h in SFM followed by treatment with ATP (3 m m ) for 45 min. For LPS and alum (Imject alum adjuvant; a mixture of aluminum hydroxide and magnesium hydroxide, Pierce) treatment, the cells were incubated with LPS (0.5 μg/ml) for 3 h in SFM followed by treatment with alum (200 μg/ml) for 4 h. Cathepsin B inhibitor, CA-074-Me (Sigma), caspase-1 inhibitor, Z-YVAD-fmk (Vergent Bioscience), cathepsin L inhibitor (Calbiochem), cytochalasin D (Sigma), glyburide (Sigma), and the P2X7R antagonists, A-438079 hydrochloride (Sigma) and AZ-10606120 (TOCRIS) were dissolved in DMSO, and used at the indicated concentrations. .. Mito-TEMPO (Santa Cruz Biotechnology) was dissolved in water and used at the indicated concentration.

    Article Title: Cathepsin B inhibition interferes with metastatic potential of human melanoma: an in vitro and in vivo study
    Article Snippet: .. For cell migration and invasion assays, cells were seeded in the presence of the cathepsin D inhibitor (Pepstatin A, 100 μM, Calbiochem), the cathepsin L inhibitor (cathepsin L inhibitor II, Z-FY-CHO, 10 μM, Calbiochem) as well as two cathepsin B inhibitors: CA-074 (cell unpermeant) and CA-074Me (cell permeant) (both given at a concentration of 10 μM, Calbiochem, Nottingham, UK). ..

    Article Title: Cathepsin L suppression increases the radiosensitivity of human glioma U251 cells via G2/M cell cycle arrest and DNA damage
    Article Snippet: .. Reagents A specific cathepsin L inhibitor, Z-FY-CHO, was purchased from Calbiochem (San Diego, CA, USA) and dissolved in dimethyl sulfoxide (DMSO; Sigma Aldrich, St Louis, MO, USA) to obtain a stock concentration of 20 mmol/L, which was aliquoted, stored at −80 °C and then diluted to the desired final concentration in DMEM/F12 at the time of use. .. Antibodies The following antibodies were used in this study: cyclin B1 (1:2000, Abcam, Cambridge, UK), Rad51 (1:1000, Abcam, Cambridge, UK), cathepsin L (1:1000, Abcam, MA, USA), γ-H2AX (1:500, Abcam, Cambridge, UK), cyclin A (1:750, Abcam, Cambridge, UK), Ku70 (1:200, Cell Signaling Technology, MA, USA), β-actin (1:1000, MultiSciences, Nanjing, China), Bcl-2 (1:200, Millipore, MA, USA), and Bax (1:500, Millipore, Billerica, MA, USA).

    Article Title: Caspase-3-independent apoptotic pathways contribute to interleukin-32γ-mediated control of Mycobacterium tuberculosis infection in THP-1 cells
    Article Snippet: THP-1 cells were cultured in RPMI 1640 supplemented with 10% FBS and 2 mM glutamine and were maintained at a concentration of 2–10 ×105 cells/mL. .. The cathepsin B inhibitor [L-3-trans- (propylcarbamoyl) oxirane-2-carbonyl]-L-isoleucyl-L-proline methyl ester (CA-074-Me)], the cathepsin D inhibitor pepstatin A, and the cathepsin L inhibitor [benzyloxycarbonyl-Leu-Leu-Tyr-fluoromethylketone (z-LLY-fmk)] were purchased from Calbiochem-EMD Millipore (Billerica, Massachusetts).

    Migration:

    Article Title: Cathepsin B inhibition interferes with metastatic potential of human melanoma: an in vitro and in vivo study
    Article Snippet: .. For cell migration and invasion assays, cells were seeded in the presence of the cathepsin D inhibitor (Pepstatin A, 100 μM, Calbiochem), the cathepsin L inhibitor (cathepsin L inhibitor II, Z-FY-CHO, 10 μM, Calbiochem) as well as two cathepsin B inhibitors: CA-074 (cell unpermeant) and CA-074Me (cell permeant) (both given at a concentration of 10 μM, Calbiochem, Nottingham, UK). ..

    Optical Imaging:

    Article Title: Combined In Vivo Molecular and Anatomic Imaging for Detection of Ovarian Carcinoma-Associated Protease Activity and Integrin Expression in Mice 1 Molecular and Anatomic Imaging for Detection of Ovarian Carcinoma-Associated Protease Activity and Integrin Expression in Mice 1 2
    Article Snippet: Paragraph title: In Vitro Detection of Optical Imaging Probes ... Parallel cultures of MOVCAR 5009 cells were grown in the presence and absence of cathepsin protease inhibitors (cathepsin B inhibitor III [CA-074] or cathepsin L inhibitor [CAA0225]; EMD Chemicals, Inc, Gibbstown, NJ) and MMP inhibitors (ARP-100; Tocris Bioscience, Bristol, United Kingdom).

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    Millipore cathepsin l inhibitor
    Cathepsin activity and cystatin C amount in primary and metastatic human melanoma cells . ( A ) Quantitative cytofluorimetric analysis of plasma membrane cathepsin B, D and L in four different primary (white columns) and metastatic (black columns) melanoma cell lines. Numbers represent the mean values among four different primary or metastatic melanoma cell lines analyzed. ( B ) Fluorimetric assays for cathepsin B (black columns), cathepsin D (grey columns) and <t>cathepsin</t> L (white columns) activity in the cell medium of two representative primary and metastatic melanoma cell lines. Results are reported as fluorescence units. ( C ) Concentration of cystatin C in the growth medium of two representative primary and metastatic melanoma cell lines obtained by ELISA test. Results are reported as ng/ml. (*) p
    Cathepsin L Inhibitor, supplied by Millipore, used in various techniques. Bioz Stars score: 90/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cathepsin activity and cystatin C amount in primary and metastatic human melanoma cells . ( A ) Quantitative cytofluorimetric analysis of plasma membrane cathepsin B, D and L in four different primary (white columns) and metastatic (black columns) melanoma cell lines. Numbers represent the mean values among four different primary or metastatic melanoma cell lines analyzed. ( B ) Fluorimetric assays for cathepsin B (black columns), cathepsin D (grey columns) and cathepsin L (white columns) activity in the cell medium of two representative primary and metastatic melanoma cell lines. Results are reported as fluorescence units. ( C ) Concentration of cystatin C in the growth medium of two representative primary and metastatic melanoma cell lines obtained by ELISA test. Results are reported as ng/ml. (*) p

    Journal: Molecular Cancer

    Article Title: Cathepsin B inhibition interferes with metastatic potential of human melanoma: an in vitro and in vivo study

    doi: 10.1186/1476-4598-9-207

    Figure Lengend Snippet: Cathepsin activity and cystatin C amount in primary and metastatic human melanoma cells . ( A ) Quantitative cytofluorimetric analysis of plasma membrane cathepsin B, D and L in four different primary (white columns) and metastatic (black columns) melanoma cell lines. Numbers represent the mean values among four different primary or metastatic melanoma cell lines analyzed. ( B ) Fluorimetric assays for cathepsin B (black columns), cathepsin D (grey columns) and cathepsin L (white columns) activity in the cell medium of two representative primary and metastatic melanoma cell lines. Results are reported as fluorescence units. ( C ) Concentration of cystatin C in the growth medium of two representative primary and metastatic melanoma cell lines obtained by ELISA test. Results are reported as ng/ml. (*) p

    Article Snippet: For cell migration and invasion assays, cells were seeded in the presence of the cathepsin D inhibitor (Pepstatin A, 100 μM, Calbiochem), the cathepsin L inhibitor (cathepsin L inhibitor II, Z-FY-CHO, 10 μM, Calbiochem) as well as two cathepsin B inhibitors: CA-074 (cell unpermeant) and CA-074Me (cell permeant) (both given at a concentration of 10 μM, Calbiochem, Nottingham, UK).

    Techniques: Activity Assay, Fluorescence, Concentration Assay, Enzyme-linked Immunosorbent Assay

    Cathepsins and in vitro invasiveness: effect of chemical and biological inhibitors . ( A ) Invasion test on primary (white columns) and metastatic (black columns) melanoma cell lines. ( B ) Invasion test on two representative cell lines from primary (white columns) and metastatic (black columns) melanoma in the presence of CA-074 (left panel), Pepstatin A (central panel) or cathepsin L inhibitor (right panel). Dotted lines represent the mean of the results obtained in primary melanoma cell lines, and dashed lines indicate the mean of the results obtained in metastatic melanoma cell lines by using DMSO (vehicle of the cathepsin inhibitors). ( C ) Invasion test on two representative cell lines from primary (white columns) and metastatic (black columns) melanoma in the absence or in the presence of specific antibodies against cathepsin B (left panel), cathepsin D (central panel) or cathepsin L (right panel). Dotted lines represent the mean of the results obtained in primary melanoma cell lines, and dashed lines indicate the mean of the results obtained in metastatic melanoma cell lines by using IgG1 as positive control. ( D ) Histogram showing the results obtained from four different primary melanoma (white columns) or metastatic melanoma (black columns) cell lines in the absence or in the presence of CA-074 or antibodies against cathepsin B. Data are reported as mean ± SD of the percentage of invading cells. Student's t -test indicates: p = 0.0097 for untreated MM cells vs. CA-074-treated MM cells and p = 0.0030 for untreated MM cells vs. MM cells treated with anti-cathepsin B antibodies.

    Journal: Molecular Cancer

    Article Title: Cathepsin B inhibition interferes with metastatic potential of human melanoma: an in vitro and in vivo study

    doi: 10.1186/1476-4598-9-207

    Figure Lengend Snippet: Cathepsins and in vitro invasiveness: effect of chemical and biological inhibitors . ( A ) Invasion test on primary (white columns) and metastatic (black columns) melanoma cell lines. ( B ) Invasion test on two representative cell lines from primary (white columns) and metastatic (black columns) melanoma in the presence of CA-074 (left panel), Pepstatin A (central panel) or cathepsin L inhibitor (right panel). Dotted lines represent the mean of the results obtained in primary melanoma cell lines, and dashed lines indicate the mean of the results obtained in metastatic melanoma cell lines by using DMSO (vehicle of the cathepsin inhibitors). ( C ) Invasion test on two representative cell lines from primary (white columns) and metastatic (black columns) melanoma in the absence or in the presence of specific antibodies against cathepsin B (left panel), cathepsin D (central panel) or cathepsin L (right panel). Dotted lines represent the mean of the results obtained in primary melanoma cell lines, and dashed lines indicate the mean of the results obtained in metastatic melanoma cell lines by using IgG1 as positive control. ( D ) Histogram showing the results obtained from four different primary melanoma (white columns) or metastatic melanoma (black columns) cell lines in the absence or in the presence of CA-074 or antibodies against cathepsin B. Data are reported as mean ± SD of the percentage of invading cells. Student's t -test indicates: p = 0.0097 for untreated MM cells vs. CA-074-treated MM cells and p = 0.0030 for untreated MM cells vs. MM cells treated with anti-cathepsin B antibodies.

    Article Snippet: For cell migration and invasion assays, cells were seeded in the presence of the cathepsin D inhibitor (Pepstatin A, 100 μM, Calbiochem), the cathepsin L inhibitor (cathepsin L inhibitor II, Z-FY-CHO, 10 μM, Calbiochem) as well as two cathepsin B inhibitors: CA-074 (cell unpermeant) and CA-074Me (cell permeant) (both given at a concentration of 10 μM, Calbiochem, Nottingham, UK).

    Techniques: In Vitro, Positive Control

    Cathepsin B expression in cells from primary and metastatic human melanomas . Western blot analysis (left panels) of cathepsin B, cathepsin D, cathepsin L and tubulin in the primary melanoma cell lines PM1 ( A ) and PM2 ( B) and in the metastatic melanoma cell lines MM1 ( A ) and MM2 ( B ). Quantitative flow cytometry analysis (right panels) of plasma membrane cathepsin B in PM1 ( A ) and PM2 ( B ) cell lines and in MM1 ( A ) and MM2 ( B ) cell lines. The values of Western blot signals, reported in the bottom panels of ( A ) and ( B ), were obtained by densitometric analysis and expressed as arbitrary units (a.u.). Numbers in the right panels of ( A ) and ( B ) represent the median fluorescence values. In the right panels of ( A ) and ( B ) statistical analysis performed by non-parametric K/S test is reported.

    Journal: Molecular Cancer

    Article Title: Cathepsin B inhibition interferes with metastatic potential of human melanoma: an in vitro and in vivo study

    doi: 10.1186/1476-4598-9-207

    Figure Lengend Snippet: Cathepsin B expression in cells from primary and metastatic human melanomas . Western blot analysis (left panels) of cathepsin B, cathepsin D, cathepsin L and tubulin in the primary melanoma cell lines PM1 ( A ) and PM2 ( B) and in the metastatic melanoma cell lines MM1 ( A ) and MM2 ( B ). Quantitative flow cytometry analysis (right panels) of plasma membrane cathepsin B in PM1 ( A ) and PM2 ( B ) cell lines and in MM1 ( A ) and MM2 ( B ) cell lines. The values of Western blot signals, reported in the bottom panels of ( A ) and ( B ), were obtained by densitometric analysis and expressed as arbitrary units (a.u.). Numbers in the right panels of ( A ) and ( B ) represent the median fluorescence values. In the right panels of ( A ) and ( B ) statistical analysis performed by non-parametric K/S test is reported.

    Article Snippet: For cell migration and invasion assays, cells were seeded in the presence of the cathepsin D inhibitor (Pepstatin A, 100 μM, Calbiochem), the cathepsin L inhibitor (cathepsin L inhibitor II, Z-FY-CHO, 10 μM, Calbiochem) as well as two cathepsin B inhibitors: CA-074 (cell unpermeant) and CA-074Me (cell permeant) (both given at a concentration of 10 μM, Calbiochem, Nottingham, UK).

    Techniques: Expressing, Western Blot, Flow Cytometry, Cytometry, Fluorescence

    PGRN processing into GRNs is mediated by proper lysosome function and cysteine protease activity. A , HAP1 WT cells were treated for 24 h with the pan-lysosome inhibitors chloroquine (CQ; 50 µM), BafA1 (50 nM), or concanamycin A (ConA; 50 nM) and conditioned media were analyzed for secreted PGRN by ELISA. B , Lysates from HAP1 WT cells treated as in A were analyzed for PGRN and GRN-2,3 by immunoblot. C , D , Quantification of ( C ) PGRN and ( D ) GRN-2,3 from the experiment in B . E , HAP1 WT cells were treated with the indicated protease inhibitors for 24 h and analyzed for PGRN and GRN-2,3 by immunoblot. The inhibitors, their primary targets, and concentrations used are shown in the table at right. F , Time-dependent cleavage of C-TAP PGRN by recombinant cathepsin L in vitro . G , C-TAP PGRN was incubated with or without cathepsin L for 2 h in vitro and analyzed for multiple GRNs by immunoblot. H , HAP1 WT cells were treated with increasing concentrations of cathepsin L inhibitor II (Z-FY-CHO) for 40 h and lysates were analyzed for PGRN and GRN-2,3 by immunoblot. I , J , Quantification of ( I ) PGRN and ( J ) GRN-2,3 from the experiment in H . B , E , H , Arrows denote endogenous, intermediate PGRN cleavage products. For all immunoblots, PGRN and GRN-2,3 were detected with R D AF2420 and Sigma antibodies respectively. All immunoblot images are representative of at least three independent experiments and quantitative data are presented as mean ± SEM of three independent experiments; *differs from control p

    Journal: eNeuro

    Article Title: Intracellular Proteolysis of Progranulin Generates Stable, Lysosomal Granulins that Are Haploinsufficient in Patients with Frontotemporal Dementia Caused by GRN Mutations

    doi: 10.1523/ENEURO.0100-17.2017

    Figure Lengend Snippet: PGRN processing into GRNs is mediated by proper lysosome function and cysteine protease activity. A , HAP1 WT cells were treated for 24 h with the pan-lysosome inhibitors chloroquine (CQ; 50 µM), BafA1 (50 nM), or concanamycin A (ConA; 50 nM) and conditioned media were analyzed for secreted PGRN by ELISA. B , Lysates from HAP1 WT cells treated as in A were analyzed for PGRN and GRN-2,3 by immunoblot. C , D , Quantification of ( C ) PGRN and ( D ) GRN-2,3 from the experiment in B . E , HAP1 WT cells were treated with the indicated protease inhibitors for 24 h and analyzed for PGRN and GRN-2,3 by immunoblot. The inhibitors, their primary targets, and concentrations used are shown in the table at right. F , Time-dependent cleavage of C-TAP PGRN by recombinant cathepsin L in vitro . G , C-TAP PGRN was incubated with or without cathepsin L for 2 h in vitro and analyzed for multiple GRNs by immunoblot. H , HAP1 WT cells were treated with increasing concentrations of cathepsin L inhibitor II (Z-FY-CHO) for 40 h and lysates were analyzed for PGRN and GRN-2,3 by immunoblot. I , J , Quantification of ( I ) PGRN and ( J ) GRN-2,3 from the experiment in H . B , E , H , Arrows denote endogenous, intermediate PGRN cleavage products. For all immunoblots, PGRN and GRN-2,3 were detected with R D AF2420 and Sigma antibodies respectively. All immunoblot images are representative of at least three independent experiments and quantitative data are presented as mean ± SEM of three independent experiments; *differs from control p

    Article Snippet: Finally, treatment of HAP1 WT cells with the cathepsin L inhibitor II (Z-FY-CHO; Calbiochem) modestly increased endogenous PGRN , and significantly decreased production of GRN-2,3 ( ).

    Techniques: Activity Assay, Enzyme-linked Immunosorbent Assay, Recombinant, In Vitro, Incubation, Western Blot

    Model of intracellular processing of PGRN into stable, lysosomal GRNs. Sortilin and other receptors target endocytosed and newly synthesized PGRN to lysosomes. Within lysosomes, PGRN is proteolytically cleaved, in part, by cysteine proteases (i.e., cathepsin L) into mature, stable GRN proteins. Ablation of sortilin results in the reduced production of GRNs. Further, lysosome dysfunction caused by alkalizing agents or TMEM106B overexpression inhibits the processing of PGRN into GRNs. FTD- GRN  patients are haploinsufficient for GRNs, which may drive lysosome dysfunction leading to neurodegeneration.

    Journal: eNeuro

    Article Title: Intracellular Proteolysis of Progranulin Generates Stable, Lysosomal Granulins that Are Haploinsufficient in Patients with Frontotemporal Dementia Caused by GRN Mutations

    doi: 10.1523/ENEURO.0100-17.2017

    Figure Lengend Snippet: Model of intracellular processing of PGRN into stable, lysosomal GRNs. Sortilin and other receptors target endocytosed and newly synthesized PGRN to lysosomes. Within lysosomes, PGRN is proteolytically cleaved, in part, by cysteine proteases (i.e., cathepsin L) into mature, stable GRN proteins. Ablation of sortilin results in the reduced production of GRNs. Further, lysosome dysfunction caused by alkalizing agents or TMEM106B overexpression inhibits the processing of PGRN into GRNs. FTD- GRN patients are haploinsufficient for GRNs, which may drive lysosome dysfunction leading to neurodegeneration.

    Article Snippet: Finally, treatment of HAP1 WT cells with the cathepsin L inhibitor II (Z-FY-CHO; Calbiochem) modestly increased endogenous PGRN , and significantly decreased production of GRN-2,3 ( ).

    Techniques: Synthesized, Over Expression