proteinase k agarose  (Millipore)


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    Structured Review

    Millipore proteinase k agarose
    In vivo  activation of DCs after U-Omp19 injection. U-Omp19 either untreated or digested with proteinase K,  E. coli  LPS or PBS alone was injected i.v. to BALB/c mice. Splenic CD11c +  DCs were analyzed for their activation status 20 h after injection by assessing the surface expression of ( A ) CD40, ( B ) CD80 and ( C ) CD86 molecules by flow cytometry. This experiment was conducted three times with similar results. Histograms display results from one representative experiment.
    Proteinase K Agarose, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    proteinase k agarose - by Bioz Stars, 2020-04
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    Images

    1) Product Images from "An Oral Vaccine Based on U-Omp19 Induces Protection against B. abortus Mucosal Challenge by Inducing an Adaptive IL-17 Immune Response in Mice"

    Article Title: An Oral Vaccine Based on U-Omp19 Induces Protection against B. abortus Mucosal Challenge by Inducing an Adaptive IL-17 Immune Response in Mice

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0016203

    In vivo  activation of DCs after U-Omp19 injection. U-Omp19 either untreated or digested with proteinase K,  E. coli  LPS or PBS alone was injected i.v. to BALB/c mice. Splenic CD11c +  DCs were analyzed for their activation status 20 h after injection by assessing the surface expression of ( A ) CD40, ( B ) CD80 and ( C ) CD86 molecules by flow cytometry. This experiment was conducted three times with similar results. Histograms display results from one representative experiment.
    Figure Legend Snippet: In vivo activation of DCs after U-Omp19 injection. U-Omp19 either untreated or digested with proteinase K, E. coli LPS or PBS alone was injected i.v. to BALB/c mice. Splenic CD11c + DCs were analyzed for their activation status 20 h after injection by assessing the surface expression of ( A ) CD40, ( B ) CD80 and ( C ) CD86 molecules by flow cytometry. This experiment was conducted three times with similar results. Histograms display results from one representative experiment.

    Techniques Used: In Vivo, Activation Assay, Injection, Mouse Assay, Expressing, Flow Cytometry, Cytometry

    2) Product Images from "Unlipidated Outer Membrane Protein Omp16 (U-Omp16) from Brucella spp. as Nasal Adjuvant Induces a Th1 Immune Response and Modulates the Th2 Allergic Response to Cow's Milk Proteins"

    Article Title: Unlipidated Outer Membrane Protein Omp16 (U-Omp16) from Brucella spp. as Nasal Adjuvant Induces a Th1 Immune Response and Modulates the Th2 Allergic Response to Cow's Milk Proteins

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0069438

    U-Omp16 induces a T helper 1 immune response when administered as nasal adjuvant. C57BL/6 mice were immunized by the nasal route with: OVA plus (i) PBS or ii) U-Omp16 previously digested with proteinase K (U-Omp16PK) or plus (iii) U-Omp16. Three weeks after last immunization animals were sacrificed and spleen cells were stimulated in vitro with OVA 500 µg/ml or complete medium (RPMI). Culture supernatants were harvested 5 days after stimulation and cytokine concentration of ( A ) IFN-γ, ( B ) IL-17, ( C ) IL-4 and ( D ) IL-10 (pg/ml) were determined by ELISA. U-Omp16 when administered as nasal adjuvant stimulates the induction of CD4 + and CD8 + OVA-specific T cells that produce IFN-γ. Percentages are represented for spleen ( E ) CD8 + or CD4 + T lymphocytes, and lung CD8 + T cells ( F ) expressing IFN-γ. ( G ) Anti-OVA IgG was determined in sera from immunized animals on days 0 (pre-immune) and 30 (post-immune) by indirect ELISA. Data represent the mean ±SEM from each group of five mice; (*** P
    Figure Legend Snippet: U-Omp16 induces a T helper 1 immune response when administered as nasal adjuvant. C57BL/6 mice were immunized by the nasal route with: OVA plus (i) PBS or ii) U-Omp16 previously digested with proteinase K (U-Omp16PK) or plus (iii) U-Omp16. Three weeks after last immunization animals were sacrificed and spleen cells were stimulated in vitro with OVA 500 µg/ml or complete medium (RPMI). Culture supernatants were harvested 5 days after stimulation and cytokine concentration of ( A ) IFN-γ, ( B ) IL-17, ( C ) IL-4 and ( D ) IL-10 (pg/ml) were determined by ELISA. U-Omp16 when administered as nasal adjuvant stimulates the induction of CD4 + and CD8 + OVA-specific T cells that produce IFN-γ. Percentages are represented for spleen ( E ) CD8 + or CD4 + T lymphocytes, and lung CD8 + T cells ( F ) expressing IFN-γ. ( G ) Anti-OVA IgG was determined in sera from immunized animals on days 0 (pre-immune) and 30 (post-immune) by indirect ELISA. Data represent the mean ±SEM from each group of five mice; (*** P

    Techniques Used: Mouse Assay, In Vitro, Concentration Assay, Enzyme-linked Immunosorbent Assay, Expressing, Indirect ELISA

    3) Product Images from "Expression, purification and reconstitution of the 4-hydroxybenzoate transporter PcaK from Acinetobacter sp. ADP1"

    Article Title: Expression, purification and reconstitution of the 4-hydroxybenzoate transporter PcaK from Acinetobacter sp. ADP1

    Journal: Protein Expression and Purification

    doi: 10.1016/j.pep.2014.05.011

    Susceptibility of the C-terminal V5 tag to proteolysis as a measure of protein orientation. Western blotting shows that Proteinase K immediately digests the C-terminal V5 epitope of PcaK in detergent micelles ( Protein) and after reconstitution into proteoliposomes ( PL ), suggesting that the C-terminal of the reconstituted protein is exposed at the proteoliposome exterior. Time in minutes.
    Figure Legend Snippet: Susceptibility of the C-terminal V5 tag to proteolysis as a measure of protein orientation. Western blotting shows that Proteinase K immediately digests the C-terminal V5 epitope of PcaK in detergent micelles ( Protein) and after reconstitution into proteoliposomes ( PL ), suggesting that the C-terminal of the reconstituted protein is exposed at the proteoliposome exterior. Time in minutes.

    Techniques Used: Western Blot

    4) Product Images from "Mycobacterium tuberculosis Cpn60.2 and DnaK Are Located on the Bacterial Surface, Where Cpn60.2 Facilitates Efficient Bacterial Association with Macrophages ▿"

    Article Title: Mycobacterium tuberculosis Cpn60.2 and DnaK Are Located on the Bacterial Surface, Where Cpn60.2 Facilitates Efficient Bacterial Association with Macrophages ▿

    Journal: Infection and Immunity

    doi: 10.1128/IAI.00143-09

    The water-soluble inhibitory moiety within the M. tuberculosis capsule is a protein and not either of the two major capsular glycans. (A and B) Arabinomannan (ArMn) and glucan (Gn) purified from M. tuberculosis were added to peritoneal Mφ at a final concentration of 5 mg ml −1 . Control Mφ received binding medium alone (Con). The subsequent association of syringed M. tuberculosis to these Mφ was assessed microscopically. The mean (±SEM) percentage of the Mφ population able to bind ≥1 bacillus is shown for three experiments, each with two coverslips (A), or four experiments, each with two coverslips (B). No significant difference in binding was observed. (C) Five hundred μg ml −1 of capsule (Capsule) or the TCA-precipitated protein from that same amount of capsule (Protein) was added to peritoneal Mφ prior to the addition of M. tuberculosis . Control Mφ received binding medium alone (Control) or protein that had been treated with proteinase K (Digest). The subsequent association of syringed M. tuberculosis to these Mφ was assessed. The mean (±SEM) percentage of the Mφ population able to bind ≥1 or > 10 bacteria is shown for two experiments, each with three coverslips (*, P
    Figure Legend Snippet: The water-soluble inhibitory moiety within the M. tuberculosis capsule is a protein and not either of the two major capsular glycans. (A and B) Arabinomannan (ArMn) and glucan (Gn) purified from M. tuberculosis were added to peritoneal Mφ at a final concentration of 5 mg ml −1 . Control Mφ received binding medium alone (Con). The subsequent association of syringed M. tuberculosis to these Mφ was assessed microscopically. The mean (±SEM) percentage of the Mφ population able to bind ≥1 bacillus is shown for three experiments, each with two coverslips (A), or four experiments, each with two coverslips (B). No significant difference in binding was observed. (C) Five hundred μg ml −1 of capsule (Capsule) or the TCA-precipitated protein from that same amount of capsule (Protein) was added to peritoneal Mφ prior to the addition of M. tuberculosis . Control Mφ received binding medium alone (Control) or protein that had been treated with proteinase K (Digest). The subsequent association of syringed M. tuberculosis to these Mφ was assessed. The mean (±SEM) percentage of the Mφ population able to bind ≥1 or > 10 bacteria is shown for two experiments, each with three coverslips (*, P

    Techniques Used: Purification, Concentration Assay, Binding Assay

    5) Product Images from "REST/NRSF-Interacting LIM Domain Protein, a Putative Nuclear Translocation Receptor"

    Article Title: REST/NRSF-Interacting LIM Domain Protein, a Putative Nuclear Translocation Receptor

    Journal: Molecular and Cellular Biology

    doi: 10.1128/MCB.23.24.9025-9031.2003

    HeLa nuclear digestion with proteinase K. Nuclei (200 μl) from HeLa cells, prepared as described in Materials and Methods, were incubated with 10 μl of proteinase K-agarose (Sigma) at room temperature for 10 min. A protease inhibitor cocktail (P-2714; Sigma) and PMSF were added to stop the reaction, and the protease was removed by centrifugation. After sonication, the mixture was centrifuged (100,000 ×  g  for 30 min) and the supernatant was used as the nuclear extract. The pellet was resuspended in buffer containing Triton X-100, sonicated, and then centrifuged to yield solubilized membrane proteins. Alternatively, nuclei were treated in the same way, except that sonication was done before the incubation with proteinase K. An aliquot (10 μl) of each supernatant was subjected to SDS-PAGE (10% polyacrylamide gel) followed by Western blot analysis with anti-RILP and anti-Myc antibodies.
    Figure Legend Snippet: HeLa nuclear digestion with proteinase K. Nuclei (200 μl) from HeLa cells, prepared as described in Materials and Methods, were incubated with 10 μl of proteinase K-agarose (Sigma) at room temperature for 10 min. A protease inhibitor cocktail (P-2714; Sigma) and PMSF were added to stop the reaction, and the protease was removed by centrifugation. After sonication, the mixture was centrifuged (100,000 × g for 30 min) and the supernatant was used as the nuclear extract. The pellet was resuspended in buffer containing Triton X-100, sonicated, and then centrifuged to yield solubilized membrane proteins. Alternatively, nuclei were treated in the same way, except that sonication was done before the incubation with proteinase K. An aliquot (10 μl) of each supernatant was subjected to SDS-PAGE (10% polyacrylamide gel) followed by Western blot analysis with anti-RILP and anti-Myc antibodies.

    Techniques Used: Incubation, Protease Inhibitor, Centrifugation, Sonication, SDS Page, Western Blot

    6) Product Images from "The TLR2 is activated by sporozoites and suppresses intrahepatic rodent malaria parasite development"

    Article Title: The TLR2 is activated by sporozoites and suppresses intrahepatic rodent malaria parasite development

    Journal: Scientific Reports

    doi: 10.1038/srep18239

    Characterization of TLR2 agonist contained in sporozoites. ( A ), TLR2 -transfected cells were stimulated with the increased concentration of SPZ or NSG lysate, and the NF-κB gene reporter activity was then determined. ( B ), TLR2/CD14-, TLR2/1/CD14- or TLR2/6/CD14-transfected cells were stimulated with the indicated concentration of SPZ lysate or NSG lysate, and NF-κB gene reporter activity was determined. ( C ), TLR2/CD14-, TLR2/1/CD14- or TLR2/6/CD14-transfected cells were stimulated with boiled or unboiled SPZ or NSG lysate, and then NF-κB gene reporter activity was determined. ( D ), TLR2 -transfected cells (cell/sporozoites = 1:10)were stimulated with the SPZ or NSG lysate pretreated with or without protease K, and the NF-κB gene reporter activity was then determined. The NF-κB gene reporter activity was expressed as the ratio of Renilla luciferase activity to firefly luciferase activity. All the experiments were repeated three times, and all data were presented as the mean ± SD, * p 
    Figure Legend Snippet: Characterization of TLR2 agonist contained in sporozoites. ( A ), TLR2 -transfected cells were stimulated with the increased concentration of SPZ or NSG lysate, and the NF-κB gene reporter activity was then determined. ( B ), TLR2/CD14-, TLR2/1/CD14- or TLR2/6/CD14-transfected cells were stimulated with the indicated concentration of SPZ lysate or NSG lysate, and NF-κB gene reporter activity was determined. ( C ), TLR2/CD14-, TLR2/1/CD14- or TLR2/6/CD14-transfected cells were stimulated with boiled or unboiled SPZ or NSG lysate, and then NF-κB gene reporter activity was determined. ( D ), TLR2 -transfected cells (cell/sporozoites = 1:10)were stimulated with the SPZ or NSG lysate pretreated with or without protease K, and the NF-κB gene reporter activity was then determined. The NF-κB gene reporter activity was expressed as the ratio of Renilla luciferase activity to firefly luciferase activity. All the experiments were repeated three times, and all data were presented as the mean ± SD, * p 

    Techniques Used: Transfection, Concentration Assay, Activity Assay, Luciferase

    Related Articles

    Centrifugation:

    Article Title: IL-4 controls activated neutrophil FcγR2b expression and migration into inflamed joints
    Article Snippet: To degrade proteins in conditioned medium, the sample was incubated with proteinase K-agarose from Tritirachium album (P9290-10UN, Sigma-Aldrich). .. The treated samples were collected by centrifugation and assayed for its activity to up-regulate the IL-4Rα on neutrophils.

    IF-cells:

    Article Title: IL-4 controls activated neutrophil FcγR2b expression and migration into inflamed joints
    Article Snippet: Red blood cells (RBCs) were lysed using 1× RBC Lysis buffer (eBioscience), and cells stimulated overnight in 96-well plates (1:100 if cells from two femur were used per well in a 96-well plate). .. To degrade proteins in conditioned medium, the sample was incubated with proteinase K-agarose from Tritirachium album (P9290-10UN, Sigma-Aldrich).

    Blocking Assay:

    Article Title: Common cellular origin and diverging developmental programs for different sesamoid bones
    Article Snippet: In order to block non-specific binding of immunoglobulin, sections were incubated with 7% goat serum, 1% bovine serum albumin dissolved in PBST (PBS+0.1% Triton X-100+0.01% sodium azide). .. After staining for SOX9, slides were washed in PBST and fixed in 4% PFA at room temperature for 10 min. Then, slides were incubated with proteinase K (Sigma-Aldrich, P9290), washed and post-fixed again in 4% PFA.

    Incubation:

    Article Title: Saliva enhances infection of gingival fibroblasts by herpes simplex virus 1
    Article Snippet: Proteinase-K treatment: Proteinase-K agarose (P9290, Sigma-Aldrich) was hydrated in 10mM sodium phosphate buffer at pH 7.0. .. Fifty μl of packed beads were added to 100 μl saliva proteins at 6.2 mg/ml and incubated for 4h at 37°C.

    Article Title: Common cellular origin and diverging developmental programs for different sesamoid bones
    Article Snippet: .. After staining for SOX9, slides were washed in PBST and fixed in 4% PFA at room temperature for 10 min. Then, slides were incubated with proteinase K (Sigma-Aldrich, P9290), washed and post-fixed again in 4% PFA. .. Next, sections were washed and incubated overnight at 4°C with primary anti-COL2A1 antibody (1:50, II-II6B3, the Developmental Studies Hybridoma Bank).

    Article Title: Communication between viruses guides lysis-lysogeny decisions
    Article Snippet: Preparation of conditioned and control media Overnight cultures of B. subtilis 168 were diluted 1:50 in LB media supplemented with 0.1 mM MnCl2 and 5 mM MgCl2 , and incubated at 37°C with shaking until reaching O.D600 =0.5. .. For the Proteinase K assay 7.5 mg (per reaction) of Proteinase K–Agarose from Tritirachium album (Sigma, CAT# P9290) were washed twice with 750 µl of sterile water and then resuspended with 750 µl of LB supplemented with 0.1 mM MnCl2 and 5 mM MgCl2 .

    Article Title: Identification of Three Elicitins and a Galactan-Based Complex Polysaccharide from a Concentrated Culture Filtrate of Phytophthora infestans Efficient against Pectobacteriumatrosepticum
    Article Snippet: .. Polysaccharide Analysis The polysaccharide analysis was carried out after proteins were removed from fraction F1b by treatment with proteinase K. A 2.5 mL aliquot of a 10 mg·mL−1 proteinase K solution, prepared in 10 mM phosphate buffer, pH 7.6, from lyophilized enzyme fixed on an inert substrate (Sigma, P9290), was added to 5 mg F1b fraction and incubated for 8 h at 37 °C. .. Polysaccharides composition was determined according to a hydrolysis/reduction/acetylation sequence followed by mass spectrometry analysis of the resulting deuteriated residues.

    Article Title: IL-4 controls activated neutrophil FcγR2b expression and migration into inflamed joints
    Article Snippet: .. To degrade proteins in conditioned medium, the sample was incubated with proteinase K-agarose from Tritirachium album (P9290-10UN, Sigma-Aldrich). ..

    Activity Assay:

    Article Title: IL-4 controls activated neutrophil FcγR2b expression and migration into inflamed joints
    Article Snippet: To degrade proteins in conditioned medium, the sample was incubated with proteinase K-agarose from Tritirachium album (P9290-10UN, Sigma-Aldrich). .. The treated samples were collected by centrifugation and assayed for its activity to up-regulate the IL-4Rα on neutrophils.

    Mass Spectrometry:

    Article Title: Identification of Three Elicitins and a Galactan-Based Complex Polysaccharide from a Concentrated Culture Filtrate of Phytophthora infestans Efficient against Pectobacteriumatrosepticum
    Article Snippet: Polysaccharide Analysis The polysaccharide analysis was carried out after proteins were removed from fraction F1b by treatment with proteinase K. A 2.5 mL aliquot of a 10 mg·mL−1 proteinase K solution, prepared in 10 mM phosphate buffer, pH 7.6, from lyophilized enzyme fixed on an inert substrate (Sigma, P9290), was added to 5 mg F1b fraction and incubated for 8 h at 37 °C. .. Polysaccharides composition was determined according to a hydrolysis/reduction/acetylation sequence followed by mass spectrometry analysis of the resulting deuteriated residues.

    Ex Vivo:

    Article Title: IL-4 controls activated neutrophil FcγR2b expression and migration into inflamed joints
    Article Snippet: Paragraph title: Cell Lines and Ex Vivo Stimulations. ... To degrade proteins in conditioned medium, the sample was incubated with proteinase K-agarose from Tritirachium album (P9290-10UN, Sigma-Aldrich).

    Derivative Assay:

    Article Title: The Antistaphylococcal Lysin, CF-301, Activates Key Host Factors in Human Blood To Potentiate Methicillin-Resistant Staphylococcus aureus Bacteriolysis
    Article Snippet: The agents (and vendor sources) tested in combination with CF-301 were as follows: β-defensin-3, human (Anaspec); Leap-1, human (Anaspec); Leap-2, human (GenScript); LL-37, human (Anaspec); LL-18-37 (Anaspec); histatin-5, human (Anaspec); HNP-1, human (Anaspec); HNP-2, human (Anaspec); human platelet factor IV 18 (Anaspec); lactoferrin, human milk (Sigma-Aldrich); lactoferrin, bovine colostrum (Sigma-Aldrich); lactoferricin H, human (Anaspec); human lysozyme, recombinant expressed in rice (Sigma-Aldrich); lysozyme, hen egg (Sigma-Aldrich); lysozyme, human neutrophil derived (Aviva Biosciences); lysozyme, human neutrophil derived (RayBiotech); human serum albumin, recombinant expressed in rice (Sigma-Aldrich); human serum albumin, fraction V (Sigma-Aldrich); human serum albumin, fraction V, fatty acid free, globulin free (Sigma-Aldrich); human serum albumin, recombinant expressed in Saccharomyces cerevisiae (Albumin Bioscience); mouse serum albumin, recombinant expressed in yeast (Albumin Biosciences); rabbit serum albumin (Sigma-Aldrich); and rat serum albumin (Sigma-Aldrich). .. Sodium oleate, sodium palmitate, vancomycin hydrochloride, daptomycin, and proteinase K-agarose from Tritirachium album were obtained from Sigma-Aldrich.

    Transferring:

    Article Title: IL-4 controls activated neutrophil FcγR2b expression and migration into inflamed joints
    Article Snippet: To degrade proteins in conditioned medium, the sample was incubated with proteinase K-agarose from Tritirachium album (P9290-10UN, Sigma-Aldrich). .. Next, 400 µL of the beads were collected with a cut pipette tip, added to a spin column, and washed three times with sterile PBS.

    Infection:

    Article Title: Communication between viruses guides lysis-lysogeny decisions
    Article Snippet: For the Proteinase K assay 7.5 mg (per reaction) of Proteinase K–Agarose from Tritirachium album (Sigma, CAT# P9290) were washed twice with 750 µl of sterile water and then resuspended with 750 µl of LB supplemented with 0.1 mM MnCl2 and 5 mM MgCl2 . .. 1.5 ml of phi3T-derived conditioned medium or control medium was added to a tube containing the washed proteinase K. The media were incubated for 2 hours at 37°C with the proteinase K. The media were centrifuged and the supernatants were collected for the infection assay.

    other:

    Article Title: Host Iron Nutritional Immunity Induced by a Live Yersinia pestis Vaccine Strain Is Associated with Immediate Protection against Plague
    Article Snippet: Proteinase K digestion Proteolysis was performed by incubating the serum samples with proteinase K-conjugated beads (Sigma Aldrich P9290) for 2 h at 37°C according to the manufacturer's recommendations.

    Protein Concentration:

    Article Title: Saliva enhances infection of gingival fibroblasts by herpes simplex virus 1
    Article Snippet: Proteinase-K treatment: Proteinase-K agarose (P9290, Sigma-Aldrich) was hydrated in 10mM sodium phosphate buffer at pH 7.0. .. The supernatant was diluted in DMEM to an equivalent of 1 mg/ml protein concentration for cell stimulation.

    Sequencing:

    Article Title: Identification of Three Elicitins and a Galactan-Based Complex Polysaccharide from a Concentrated Culture Filtrate of Phytophthora infestans Efficient against Pectobacteriumatrosepticum
    Article Snippet: Polysaccharide Analysis The polysaccharide analysis was carried out after proteins were removed from fraction F1b by treatment with proteinase K. A 2.5 mL aliquot of a 10 mg·mL−1 proteinase K solution, prepared in 10 mM phosphate buffer, pH 7.6, from lyophilized enzyme fixed on an inert substrate (Sigma, P9290), was added to 5 mg F1b fraction and incubated for 8 h at 37 °C. .. Polysaccharides composition was determined according to a hydrolysis/reduction/acetylation sequence followed by mass spectrometry analysis of the resulting deuteriated residues.

    Injection:

    Article Title: IL-4 controls activated neutrophil FcγR2b expression and migration into inflamed joints
    Article Snippet: Stimulations include dilutions of serum from K/BxN mice (from mice with active disease), and mice injected with LPS (“LPS serum,” collected 20 to 24 h after intraperitoneal injection of 10 µg LPS into WT C57BL/6 mice), 20% conditioned medium from cell lines stimulated overnight ±0.2 µg/mL LPS, and cytokines (mouse: IL-4, IL-6, CSF3, LIF at 20 ng/mL; and human: CSF3 [10 and 25 ng/mL], and IL-6 [25 ng/mL]; all from Peprotech). .. To degrade proteins in conditioned medium, the sample was incubated with proteinase K-agarose from Tritirachium album (P9290-10UN, Sigma-Aldrich).

    Recombinant:

    Article Title: The Antistaphylococcal Lysin, CF-301, Activates Key Host Factors in Human Blood To Potentiate Methicillin-Resistant Staphylococcus aureus Bacteriolysis
    Article Snippet: The agents (and vendor sources) tested in combination with CF-301 were as follows: β-defensin-3, human (Anaspec); Leap-1, human (Anaspec); Leap-2, human (GenScript); LL-37, human (Anaspec); LL-18-37 (Anaspec); histatin-5, human (Anaspec); HNP-1, human (Anaspec); HNP-2, human (Anaspec); human platelet factor IV 18 (Anaspec); lactoferrin, human milk (Sigma-Aldrich); lactoferrin, bovine colostrum (Sigma-Aldrich); lactoferricin H, human (Anaspec); human lysozyme, recombinant expressed in rice (Sigma-Aldrich); lysozyme, hen egg (Sigma-Aldrich); lysozyme, human neutrophil derived (Aviva Biosciences); lysozyme, human neutrophil derived (RayBiotech); human serum albumin, recombinant expressed in rice (Sigma-Aldrich); human serum albumin, fraction V (Sigma-Aldrich); human serum albumin, fraction V, fatty acid free, globulin free (Sigma-Aldrich); human serum albumin, recombinant expressed in Saccharomyces cerevisiae (Albumin Bioscience); mouse serum albumin, recombinant expressed in yeast (Albumin Biosciences); rabbit serum albumin (Sigma-Aldrich); and rat serum albumin (Sigma-Aldrich). .. Sodium oleate, sodium palmitate, vancomycin hydrochloride, daptomycin, and proteinase K-agarose from Tritirachium album were obtained from Sigma-Aldrich.

    Article Title: Stimulation of TLR4 by recombinant HSP70 requires structural integrity of the HSP70 protein itself
    Article Snippet: Reagents Recombinant low-endotoxin human HSP70 (cat. # ADI-ESP-555-F; lot # 05040922) and ultrapure LPSs (cat. # ALX-581-007-L002 and ALX-581-013-L002) were from Enzo Life Sciences (Plymouth Meeting, PA). .. Polymyxin B solution, and Proteinase K-agarose from Tritirachium album, were from Sigma (St. Louis, MO).

    Immunofluorescence:

    Article Title: Common cellular origin and diverging developmental programs for different sesamoid bones
    Article Snippet: Paragraph title: Immunofluorescence staining ... After staining for SOX9, slides were washed in PBST and fixed in 4% PFA at room temperature for 10 min. Then, slides were incubated with proteinase K (Sigma-Aldrich, P9290), washed and post-fixed again in 4% PFA.

    Cell Stimulation:

    Article Title: Saliva enhances infection of gingival fibroblasts by herpes simplex virus 1
    Article Snippet: Proteinase-K treatment: Proteinase-K agarose (P9290, Sigma-Aldrich) was hydrated in 10mM sodium phosphate buffer at pH 7.0. .. The supernatant was diluted in DMEM to an equivalent of 1 mg/ml protein concentration for cell stimulation.

    Microscopy:

    Article Title: The Antistaphylococcal Lysin, CF-301, Activates Key Host Factors in Human Blood To Potentiate Methicillin-Resistant Staphylococcus aureus Bacteriolysis
    Article Snippet: Sodium oleate, sodium palmitate, vancomycin hydrochloride, daptomycin, and proteinase K-agarose from Tritirachium album were obtained from Sigma-Aldrich. .. For microscopy, DAPI (4′,6-diamidino-2-phenylindole), Alexa Fluor 488, and NHS-rhodamine were obtained from ThermoFisher Scientific.

    Mouse Assay:

    Article Title: IL-4 controls activated neutrophil FcγR2b expression and migration into inflamed joints
    Article Snippet: Stimulations include dilutions of serum from K/BxN mice (from mice with active disease), and mice injected with LPS (“LPS serum,” collected 20 to 24 h after intraperitoneal injection of 10 µg LPS into WT C57BL/6 mice), 20% conditioned medium from cell lines stimulated overnight ±0.2 µg/mL LPS, and cytokines (mouse: IL-4, IL-6, CSF3, LIF at 20 ng/mL; and human: CSF3 [10 and 25 ng/mL], and IL-6 [25 ng/mL]; all from Peprotech). .. To degrade proteins in conditioned medium, the sample was incubated with proteinase K-agarose from Tritirachium album (P9290-10UN, Sigma-Aldrich).

    Labeling:

    Article Title: The Antistaphylococcal Lysin, CF-301, Activates Key Host Factors in Human Blood To Potentiate Methicillin-Resistant Staphylococcus aureus Bacteriolysis
    Article Snippet: Sodium oleate, sodium palmitate, vancomycin hydrochloride, daptomycin, and proteinase K-agarose from Tritirachium album were obtained from Sigma-Aldrich. .. The labeling and purification of CF-301 conjugated to NHS-rhodamine and HuLYZ conjugated to Alexa Fluor 488 were performed as described by the manufacturer’s protocol.

    Polyacrylamide Gel Electrophoresis:

    Article Title: The Antistaphylococcal Lysin, CF-301, Activates Key Host Factors in Human Blood To Potentiate Methicillin-Resistant Staphylococcus aureus Bacteriolysis
    Article Snippet: The fractions of the elution peak were analyzed on a 4 to 12% PAGE gel to confirm the purity of the protein. .. Sodium oleate, sodium palmitate, vancomycin hydrochloride, daptomycin, and proteinase K-agarose from Tritirachium album were obtained from Sigma-Aldrich.

    Staining:

    Article Title: Common cellular origin and diverging developmental programs for different sesamoid bones
    Article Snippet: .. After staining for SOX9, slides were washed in PBST and fixed in 4% PFA at room temperature for 10 min. Then, slides were incubated with proteinase K (Sigma-Aldrich, P9290), washed and post-fixed again in 4% PFA. .. Next, sections were washed and incubated overnight at 4°C with primary anti-COL2A1 antibody (1:50, II-II6B3, the Developmental Studies Hybridoma Bank).

    Chloramphenicol Acetyltransferase Assay:

    Article Title: Communication between viruses guides lysis-lysogeny decisions
    Article Snippet: .. For the Proteinase K assay 7.5 mg (per reaction) of Proteinase K–Agarose from Tritirachium album (Sigma, CAT P9290) were washed twice with 750 µl of sterile water and then resuspended with 750 µl of LB supplemented with 0.1 mM MnCl2 and 5 mM MgCl2 . .. 1.5 ml of phi3T-derived conditioned medium or control medium was added to a tube containing the washed proteinase K. The media were incubated for 2 hours at 37°C with the proteinase K. The media were centrifuged and the supernatants were collected for the infection assay.

    Purification:

    Article Title: The Antistaphylococcal Lysin, CF-301, Activates Key Host Factors in Human Blood To Potentiate Methicillin-Resistant Staphylococcus aureus Bacteriolysis
    Article Snippet: Sodium oleate, sodium palmitate, vancomycin hydrochloride, daptomycin, and proteinase K-agarose from Tritirachium album were obtained from Sigma-Aldrich. .. The labeling and purification of CF-301 conjugated to NHS-rhodamine and HuLYZ conjugated to Alexa Fluor 488 were performed as described by the manufacturer’s protocol.

    Binding Assay:

    Article Title: Common cellular origin and diverging developmental programs for different sesamoid bones
    Article Snippet: In order to block non-specific binding of immunoglobulin, sections were incubated with 7% goat serum, 1% bovine serum albumin dissolved in PBST (PBS+0.1% Triton X-100+0.01% sodium azide). .. After staining for SOX9, slides were washed in PBST and fixed in 4% PFA at room temperature for 10 min. Then, slides were incubated with proteinase K (Sigma-Aldrich, P9290), washed and post-fixed again in 4% PFA.

    Lysis:

    Article Title: IL-4 controls activated neutrophil FcγR2b expression and migration into inflamed joints
    Article Snippet: Red blood cells (RBCs) were lysed using 1× RBC Lysis buffer (eBioscience), and cells stimulated overnight in 96-well plates (1:100 if cells from two femur were used per well in a 96-well plate). .. To degrade proteins in conditioned medium, the sample was incubated with proteinase K-agarose from Tritirachium album (P9290-10UN, Sigma-Aldrich).

    Affinity Column:

    Article Title: The Antistaphylococcal Lysin, CF-301, Activates Key Host Factors in Human Blood To Potentiate Methicillin-Resistant Staphylococcus aureus Bacteriolysis
    Article Snippet: The monoclonal antibody (MAb) was selectively bound to the affinity column; washed with 20 mM sodium phosphate, pH 7.0; and eluted in 100 mM glycine-hydrochloric acid, pH 2.7. .. Sodium oleate, sodium palmitate, vancomycin hydrochloride, daptomycin, and proteinase K-agarose from Tritirachium album were obtained from Sigma-Aldrich.

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  • 99
    Millipore proteinase k
    Proteinase K, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 610 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/proteinase k/product/Millipore
    Average 99 stars, based on 610 article reviews
    Price from $9.99 to $1999.99
    proteinase k - by Bioz Stars, 2020-04
    99/100 stars
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    94
    Millipore proteinase k inhibitor
    Detection of CDK2 in HBV capsids purified from HepG2 cells by Western blotting. HBV capsids (0.5 μg) were digested with <t>proteinase</t> K (PK) agarose beads as described in Materials and Methods. Proteinase K was then inactivated by the addition of the proteinase K inhibitor, and the sample was resolved by SDS-PAGE (lanes 8 and 10), along with the same amount of undigested capsids from the same fractions (lanes 7 and 9). Purified GST-CDK2 standards (lanes 1 to 3) and total lysate from HepG2 cells (lane 4) were loaded as controls for CDK2 detection. A sucrose fraction that did not contain HBV capsids was also treated with the proteinase K agarose beads to ensure that contaminating proteins were removed by the digestion (lane 6) or mock treated (lane 5). The capsids loaded in lanes 9 and 10 were from a capsid-positive sucrose fraction of higher purity than those loaded in lanes 7 and 8. CDK2 was detected by Western blotting using the anti-CDK2 antibody (top panel). HBc was visualized by Sypro ruby staining (bottom panel). C, core protein.
    Proteinase K Inhibitor, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/proteinase k inhibitor/product/Millipore
    Average 94 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    proteinase k inhibitor - by Bioz Stars, 2020-04
    94/100 stars
      Buy from Supplier

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    Detection of CDK2 in HBV capsids purified from HepG2 cells by Western blotting. HBV capsids (0.5 μg) were digested with proteinase K (PK) agarose beads as described in Materials and Methods. Proteinase K was then inactivated by the addition of the proteinase K inhibitor, and the sample was resolved by SDS-PAGE (lanes 8 and 10), along with the same amount of undigested capsids from the same fractions (lanes 7 and 9). Purified GST-CDK2 standards (lanes 1 to 3) and total lysate from HepG2 cells (lane 4) were loaded as controls for CDK2 detection. A sucrose fraction that did not contain HBV capsids was also treated with the proteinase K agarose beads to ensure that contaminating proteins were removed by the digestion (lane 6) or mock treated (lane 5). The capsids loaded in lanes 9 and 10 were from a capsid-positive sucrose fraction of higher purity than those loaded in lanes 7 and 8. CDK2 was detected by Western blotting using the anti-CDK2 antibody (top panel). HBc was visualized by Sypro ruby staining (bottom panel). C, core protein.

    Journal: Journal of Virology

    Article Title: Cyclin-Dependent Kinase 2 Phosphorylates S/T-P Sites in the Hepadnavirus Core Protein C-Terminal Domain and Is Incorporated into Viral Capsids

    doi: 10.1128/JVI.01218-12

    Figure Lengend Snippet: Detection of CDK2 in HBV capsids purified from HepG2 cells by Western blotting. HBV capsids (0.5 μg) were digested with proteinase K (PK) agarose beads as described in Materials and Methods. Proteinase K was then inactivated by the addition of the proteinase K inhibitor, and the sample was resolved by SDS-PAGE (lanes 8 and 10), along with the same amount of undigested capsids from the same fractions (lanes 7 and 9). Purified GST-CDK2 standards (lanes 1 to 3) and total lysate from HepG2 cells (lane 4) were loaded as controls for CDK2 detection. A sucrose fraction that did not contain HBV capsids was also treated with the proteinase K agarose beads to ensure that contaminating proteins were removed by the digestion (lane 6) or mock treated (lane 5). The capsids loaded in lanes 9 and 10 were from a capsid-positive sucrose fraction of higher purity than those loaded in lanes 7 and 8. CDK2 was detected by Western blotting using the anti-CDK2 antibody (top panel). HBc was visualized by Sypro ruby staining (bottom panel). C, core protein.

    Article Snippet: When proteinase K-treated capsids were resolved by SDS-PAGE, the proteinase K digestion was stopped by adding proteinase K inhibitor (Calbiochem) to 1 mM (final concentration) and incubating the sample at room temperature for 10 min ( ).

    Techniques: Purification, Western Blot, SDS Page, Staining

    Endogenous kinase reactions with virion-derived HBV capsids. HBV capsids were released from virions purified from the medium of HBV-transfected HepG2 cells by NP-40 treatment. One control aliquot was removed, and the remaining virion-derived capsids were digested by proteinase K. The untreated (lane 1) or proteinase K-treated (lanes 2 to 5), virion-derived capsids (ca. 6 ng per reaction) were phosphorylated by the encapsidated kinase in endogenous kinase reactions in the presence of the indicated kinase inhibitors (lanes 3 to 5) or DMSO control (lane 2). Kinase inhibitors used included CDK2 inhibitor III (lane 3, 250 μM, 500× IC 50 for CDK2), roscovitine (lane 4, 250 μM, 350× IC 50 for CDK2), and Bisindo (lane 3, 5 μM, 500× IC 50 for PKC). An equal amount of proteinase K alone was also loaded as an additional control (lane 6). In parallel, 6 ng (lane 7) or 12 ng (lane 8) of cytoplasmic HBV capsids from HepG2 cells (also treated with proteinase K; see Materials and Methods) were subjected to the endogenous kinase reaction. Half of each reaction product was resolved by agarose gel electrophoresis to visualize capsid particles (top), and proteinase K inhibitor was added to the other half to terminate the digestion before boiling in SDS sample buffer and resolving by SDS-PAGE to visualize the total core protein (bottom). The gels were dried, and labeled capsids or core proteins were detected by autoradiography. *, unknown labeled species. CA, capsids; C, core protein; PK, proteinase K; IC, intracellular.

    Journal: Journal of Virology

    Article Title: Cyclin-Dependent Kinase 2 Phosphorylates S/T-P Sites in the Hepadnavirus Core Protein C-Terminal Domain and Is Incorporated into Viral Capsids

    doi: 10.1128/JVI.01218-12

    Figure Lengend Snippet: Endogenous kinase reactions with virion-derived HBV capsids. HBV capsids were released from virions purified from the medium of HBV-transfected HepG2 cells by NP-40 treatment. One control aliquot was removed, and the remaining virion-derived capsids were digested by proteinase K. The untreated (lane 1) or proteinase K-treated (lanes 2 to 5), virion-derived capsids (ca. 6 ng per reaction) were phosphorylated by the encapsidated kinase in endogenous kinase reactions in the presence of the indicated kinase inhibitors (lanes 3 to 5) or DMSO control (lane 2). Kinase inhibitors used included CDK2 inhibitor III (lane 3, 250 μM, 500× IC 50 for CDK2), roscovitine (lane 4, 250 μM, 350× IC 50 for CDK2), and Bisindo (lane 3, 5 μM, 500× IC 50 for PKC). An equal amount of proteinase K alone was also loaded as an additional control (lane 6). In parallel, 6 ng (lane 7) or 12 ng (lane 8) of cytoplasmic HBV capsids from HepG2 cells (also treated with proteinase K; see Materials and Methods) were subjected to the endogenous kinase reaction. Half of each reaction product was resolved by agarose gel electrophoresis to visualize capsid particles (top), and proteinase K inhibitor was added to the other half to terminate the digestion before boiling in SDS sample buffer and resolving by SDS-PAGE to visualize the total core protein (bottom). The gels were dried, and labeled capsids or core proteins were detected by autoradiography. *, unknown labeled species. CA, capsids; C, core protein; PK, proteinase K; IC, intracellular.

    Article Snippet: When proteinase K-treated capsids were resolved by SDS-PAGE, the proteinase K digestion was stopped by adding proteinase K inhibitor (Calbiochem) to 1 mM (final concentration) and incubating the sample at room temperature for 10 min ( ).

    Techniques: Derivative Assay, Purification, Transfection, Agarose Gel Electrophoresis, SDS Page, Labeling, Autoradiography