polyubiquitin affinity resin  (Thermo Fisher)


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    Thermo Fisher polyubiquitin affinity resin
    Polyubiquitin Affinity Resin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polyubiquitin affinity resin/product/Thermo Fisher
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    polyubiquitin affinity resin - by Bioz Stars, 2023-06
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    polyubiquitin affinity resin a thermo scientific ubiquitin enrichment kit  (Thermo Fisher)


    Bioz Verified Symbol Thermo Fisher is a verified supplier
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    Thermo Fisher polyubiquitin affinity resin a thermo scientific ubiquitin enrichment kit
    Isolation of endogenously polyubiquitinated protein complexes by use of the <t>ubiquitin</t> enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the <t>polyubiquitin</t> affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.
    Polyubiquitin Affinity Resin A Thermo Scientific Ubiquitin Enrichment Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polyubiquitin affinity resin a thermo scientific ubiquitin enrichment kit/product/Thermo Fisher
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    polyubiquitin affinity resin a thermo scientific ubiquitin enrichment kit - by Bioz Stars, 2023-06
    86/100 stars

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    1) Product Images from "Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology"

    Article Title: Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology

    Journal: Antioxidants & Redox Signaling

    doi: 10.1089/ars.2016.6686

    Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the polyubiquitin affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.
    Figure Legend Snippet: Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the polyubiquitin affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.

    Techniques Used: Isolation, SDS Page, Staining, Expressing, Western Blot, Immunoprecipitation, Polyacrylamide Gel Electrophoresis

    Poly-Ub levels. (A) Total levels of poly-Ub-bound proteins in DS (n = 6/group) and DS/AD (n = 6/group) cases compared with their age-matched controls (n = 6/group). (B, C) Increased poly-Ub of two types of ubiquitin chains [K63 (B) and K48 (C)] in both DS (n = 6/group) and DS/AD (n = 6/group) cases compared with their age-matched controls (n = 6/group). Representative bands are shown and protein levels (A−C, upper bands) were normalized per total protein load, named total load (A−C, lower bands). Densitometric values are shown as percentage of Ctr Y set as 100%. Mean ± SEM (DS vs. Ctr y *p < 0.05; DS/AD vs. DS **p < 0.01; DS/AD vs. Ctr O ***p < 0.001; DS vs. Ctr Y and DS/AD vs. DS [for K48] *p < 0.05; DS vs. Ctr Y, DS/AD vs. Ctr O, and DS/AD vs. DS [for K63] **p < 0.01; Ctr O vs. Ctr Y #p < 0.05 one-way ANOVA). AD, Alzheimer's disease; ANOVA, analysis of variance; DS, Down syndrome; DS/AD, Down syndrome with Alzheimer's disease; SEM, standard error of the mean.
    Figure Legend Snippet: Poly-Ub levels. (A) Total levels of poly-Ub-bound proteins in DS (n = 6/group) and DS/AD (n = 6/group) cases compared with their age-matched controls (n = 6/group). (B, C) Increased poly-Ub of two types of ubiquitin chains [K63 (B) and K48 (C)] in both DS (n = 6/group) and DS/AD (n = 6/group) cases compared with their age-matched controls (n = 6/group). Representative bands are shown and protein levels (A−C, upper bands) were normalized per total protein load, named total load (A−C, lower bands). Densitometric values are shown as percentage of Ctr Y set as 100%. Mean ± SEM (DS vs. Ctr y *p < 0.05; DS/AD vs. DS **p < 0.01; DS/AD vs. Ctr O ***p < 0.001; DS vs. Ctr Y and DS/AD vs. DS [for K48] *p < 0.05; DS vs. Ctr Y, DS/AD vs. Ctr O, and DS/AD vs. DS [for K63] **p < 0.01; Ctr O vs. Ctr Y #p < 0.05 one-way ANOVA). AD, Alzheimer's disease; ANOVA, analysis of variance; DS, Down syndrome; DS/AD, Down syndrome with Alzheimer's disease; SEM, standard error of the mean.

    Techniques Used:

    List of Identified Polyubiquitinated Proteins by Mass Spectrometry
    Figure Legend Snippet: List of Identified Polyubiquitinated Proteins by Mass Spectrometry

    Techniques Used:

    MUbiSiDa Search
    Figure Legend Snippet: MUbiSiDa Search

    Techniques Used:

    UCH-L1. (A, B) All samples (n = 6/group) were immunoprecipitated with anti-UCH-L1. Immunoprecipitated proteins were separated on SDS-PAGE, transferred on nitrocellulose membranes, and probed with anti-poly-UbK63 (A, upper bands) and anti-poly-UbK48 (B, upper bands). (A) Shows increased levels of poly-UbK63 bound to UCH-L1 in DS and DS/AD compared with their matched controls. (B) Shows an increase in poly-UbK48 bound to UCH-L1 in DS/AD compared with DS. All the IP experiments were normalized on the total amount of UCH-L1 (indicated as Expr.; A, B, lower bands). Representative bands are shown. Densitometric values are shown as percentage of Ctr Y set as 100%. Mean ± SEM (DS vs. Ctr Y [for K63], DS/AD vs. DS [for K48] *p < 0.05; DS/AD vs. Ctr O [for K63] **p < 0.01 one-way ANOVA). UCH-L1, ubiquitin carboxyl-terminal hydrolase isozyme L1.
    Figure Legend Snippet: UCH-L1. (A, B) All samples (n = 6/group) were immunoprecipitated with anti-UCH-L1. Immunoprecipitated proteins were separated on SDS-PAGE, transferred on nitrocellulose membranes, and probed with anti-poly-UbK63 (A, upper bands) and anti-poly-UbK48 (B, upper bands). (A) Shows increased levels of poly-UbK63 bound to UCH-L1 in DS and DS/AD compared with their matched controls. (B) Shows an increase in poly-UbK48 bound to UCH-L1 in DS/AD compared with DS. All the IP experiments were normalized on the total amount of UCH-L1 (indicated as Expr.; A, B, lower bands). Representative bands are shown. Densitometric values are shown as percentage of Ctr Y set as 100%. Mean ± SEM (DS vs. Ctr Y [for K63], DS/AD vs. DS [for K48] *p < 0.05; DS/AD vs. Ctr O [for K63] **p < 0.01 one-way ANOVA). UCH-L1, ubiquitin carboxyl-terminal hydrolase isozyme L1.

    Techniques Used: Immunoprecipitation, SDS Page

    polyubiquitin affinity resin  (Thermo Fisher)


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    Thermo Fisher polyubiquitin affinity resin
    Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the <t>polyubiquitin</t> affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.
    Polyubiquitin Affinity Resin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polyubiquitin affinity resin/product/Thermo Fisher
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    polyubiquitin affinity resin - by Bioz Stars, 2023-06
    86/100 stars

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    1) Product Images from "Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology"

    Article Title: Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology

    Journal: Antioxidants & Redox Signaling

    doi: 10.1089/ars.2016.6686

    Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the polyubiquitin affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.
    Figure Legend Snippet: Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the polyubiquitin affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.

    Techniques Used: Isolation, SDS Page, Staining, Expressing, Western Blot, Immunoprecipitation, Polyacrylamide Gel Electrophoresis

    polyubiquitin affinity resin  (Thermo Fisher)


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    Thermo Fisher polyubiquitin affinity resin
    Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the <t>polyubiquitin</t> affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.
    Polyubiquitin Affinity Resin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polyubiquitin affinity resin/product/Thermo Fisher
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    polyubiquitin affinity resin - by Bioz Stars, 2023-06
    86/100 stars

    Images

    1) Product Images from "Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology"

    Article Title: Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology

    Journal: Antioxidants & Redox Signaling

    doi: 10.1089/ars.2016.6686

    Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the polyubiquitin affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.
    Figure Legend Snippet: Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the polyubiquitin affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.

    Techniques Used: Isolation, SDS Page, Staining, Expressing, Western Blot, Immunoprecipitation, Polyacrylamide Gel Electrophoresis

    polyubiquitin affinity resin  (Thermo Fisher)


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    Thermo Fisher polyubiquitin affinity resin
    Deletion of CKA2 reduces cellular levels of polyubiquitylated Cse4. A , levels of Ub n -Cse4-Myc were measured in WT, psh1 Δ, and cka2 Δ strains with a deletion in PDR5. Cse4-Myc was expressed from its native promoter. An untagged strain was used as a control. Cultures were grown to midlog phase and treated with either MG132 (100 μ m ) or DMSO for 3.5 h. Anti-ubiquitin and anti-Myc Western blots of cell lysates confirmed the efficiency of proteasome inhibition by MG132 and similar Cse4-Myc levels, respectively ( left panel ). Polyubiquitylated proteins were pulled down from 2 mg of total proteins (from MG132-treated samples) using <t>polyubiquitin</t> affinity resin. Final elutions were probed with anti-ubiquitin and anti-Myc antibodies after SDS-PAGE ( right panel ). No obvious difference was detected in Ub n -Cse4-Myc levels among WT, psh1 Δ, and cka2 Δ strains. A nonspecific band is marked with an asterisk. B , Ub n -Cse4 levels are reduced in the cka2 Δ strain. WT, psh1 Δ, and cka2 Δ strains from A were used to perform a Cse4 ubiquitylation assay. EV indicates the empty vector control. After Gal induction of Cse4 for 2 h, cells were treated with either MG132 (100 μ m ) or DMSO for another 2 h. Anti-ubiquitin and anti-Cse4 Western blots of cell lysates confirmed the efficiency of proteasome inhibition by MG132 and similar Cse4 levels, respectively ( left panel ). Polyubiquitylated proteins were pulled down, and final elutions were analyzed as in A ( right panel ).
    Polyubiquitin Affinity Resin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polyubiquitin affinity resin/product/Thermo Fisher
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    polyubiquitin affinity resin - by Bioz Stars, 2023-06
    86/100 stars

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    1) Product Images from "Phosphorylation by Casein Kinase 2 Facilitates Psh1 Protein-assisted Degradation of Cse4 Protein"

    Article Title: Phosphorylation by Casein Kinase 2 Facilitates Psh1 Protein-assisted Degradation of Cse4 Protein

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M114.580589

    Deletion of CKA2 reduces cellular levels of polyubiquitylated Cse4. A , levels of Ub n -Cse4-Myc were measured in WT, psh1 Δ, and cka2 Δ strains with a deletion in PDR5. Cse4-Myc was expressed from its native promoter. An untagged strain was used as a control. Cultures were grown to midlog phase and treated with either MG132 (100 μ m ) or DMSO for 3.5 h. Anti-ubiquitin and anti-Myc Western blots of cell lysates confirmed the efficiency of proteasome inhibition by MG132 and similar Cse4-Myc levels, respectively ( left panel ). Polyubiquitylated proteins were pulled down from 2 mg of total proteins (from MG132-treated samples) using polyubiquitin affinity resin. Final elutions were probed with anti-ubiquitin and anti-Myc antibodies after SDS-PAGE ( right panel ). No obvious difference was detected in Ub n -Cse4-Myc levels among WT, psh1 Δ, and cka2 Δ strains. A nonspecific band is marked with an asterisk. B , Ub n -Cse4 levels are reduced in the cka2 Δ strain. WT, psh1 Δ, and cka2 Δ strains from A were used to perform a Cse4 ubiquitylation assay. EV indicates the empty vector control. After Gal induction of Cse4 for 2 h, cells were treated with either MG132 (100 μ m ) or DMSO for another 2 h. Anti-ubiquitin and anti-Cse4 Western blots of cell lysates confirmed the efficiency of proteasome inhibition by MG132 and similar Cse4 levels, respectively ( left panel ). Polyubiquitylated proteins were pulled down, and final elutions were analyzed as in A ( right panel ).
    Figure Legend Snippet: Deletion of CKA2 reduces cellular levels of polyubiquitylated Cse4. A , levels of Ub n -Cse4-Myc were measured in WT, psh1 Δ, and cka2 Δ strains with a deletion in PDR5. Cse4-Myc was expressed from its native promoter. An untagged strain was used as a control. Cultures were grown to midlog phase and treated with either MG132 (100 μ m ) or DMSO for 3.5 h. Anti-ubiquitin and anti-Myc Western blots of cell lysates confirmed the efficiency of proteasome inhibition by MG132 and similar Cse4-Myc levels, respectively ( left panel ). Polyubiquitylated proteins were pulled down from 2 mg of total proteins (from MG132-treated samples) using polyubiquitin affinity resin. Final elutions were probed with anti-ubiquitin and anti-Myc antibodies after SDS-PAGE ( right panel ). No obvious difference was detected in Ub n -Cse4-Myc levels among WT, psh1 Δ, and cka2 Δ strains. A nonspecific band is marked with an asterisk. B , Ub n -Cse4 levels are reduced in the cka2 Δ strain. WT, psh1 Δ, and cka2 Δ strains from A were used to perform a Cse4 ubiquitylation assay. EV indicates the empty vector control. After Gal induction of Cse4 for 2 h, cells were treated with either MG132 (100 μ m ) or DMSO for another 2 h. Anti-ubiquitin and anti-Cse4 Western blots of cell lysates confirmed the efficiency of proteasome inhibition by MG132 and similar Cse4 levels, respectively ( left panel ). Polyubiquitylated proteins were pulled down, and final elutions were analyzed as in A ( right panel ).

    Techniques Used: Western Blot, Inhibition, SDS Page, Ubiquitin Assay, Plasmid Preparation

    polyubiquitin affinity resin  (Thermo Fisher)


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    Thermo Fisher polyubiquitin affinity resin
    Polyubiquitin Affinity Resin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polyubiquitin affinity resin/product/Thermo Fisher
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    polyubiquitin affinity resin - by Bioz Stars, 2023-06
    86/100 stars

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    polyubiquitin affinity resin  (Thermo Fisher)


    Bioz Verified Symbol Thermo Fisher is a verified supplier
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    Thermo Fisher polyubiquitin affinity resin
    Retroviral Gag proteins were tested for BCA2-induced ubiquitination by co-transfecting 293T cells with vectors expressing codon-optimized HIV-1 NL4-3, SIV mac 239 and Mo-MLV Gag proteins along with an expression vector coding for HA-BCA2 or empty vector. (A) Cell lysates were incubated with a <t>polyubiquitin</t> affinity resin and the bound fraction was eluted and analyzed by western blot. Membranes were probed with antibodies specific for p55/p24, MLV p30 and HA. Whole cell lysates were set aside for western blot analyses. (B) The BCA2-induced ubiquitination of Gag-GFP as well as native Gag was analyzed in side-by-side experiments. 293T cells were co-transfected in duplicate with each Gag construct, HA-BCA2, ΔRing BCA2 or empty vector. Cell lysates were set aside for regular western blotting, and the rest of the lysates were immunoprecipitated with an antibody anti-CA. Membranes were developed with an anti-Ubiquitin (Ub) antibody. (C) The ubiquitination of HIV-1 Gag, or HIV-1 Gag mutants, in cells expressing HA-BCA2 was also analyzed by immunoprecipitation. Cell lysates were set aside for western blot analyses and the rest of the samples were immunoprecipitated with an anti-CA specific antibody. Membranes were probed with an anti-Ubiquitin antibody. Similarly, the ubiquitination levels of HA-BCA2 were also determined. In this case, samples were immunoprecipitated using an anti-HA antibody. Results were confirmed in two additional independent assays. (D) A similar approach was used to analyze the ubiquitination levels of SIV Gag, or SIV Gag mutants, in the presence and absence of HA-BCA2. (E) Schematic representation of the putative lysine residues susceptible to become ubiquitinated by BCA2 in HIV-1 and SIV Gag (red). IP: immunoprecipitation. IB: immunoblot. WCL: whole cell lysate. V: empty vector.
    Polyubiquitin Affinity Resin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polyubiquitin affinity resin/product/Thermo Fisher
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    polyubiquitin affinity resin - by Bioz Stars, 2023-06
    86/100 stars

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    1) Product Images from "BCA2/Rabring7 Targets HIV-1 Gag for Lysosomal Degradation in a Tetherin-Independent Manner"

    Article Title: BCA2/Rabring7 Targets HIV-1 Gag for Lysosomal Degradation in a Tetherin-Independent Manner

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1004151

    Retroviral Gag proteins were tested for BCA2-induced ubiquitination by co-transfecting 293T cells with vectors expressing codon-optimized HIV-1 NL4-3, SIV mac 239 and Mo-MLV Gag proteins along with an expression vector coding for HA-BCA2 or empty vector. (A) Cell lysates were incubated with a polyubiquitin affinity resin and the bound fraction was eluted and analyzed by western blot. Membranes were probed with antibodies specific for p55/p24, MLV p30 and HA. Whole cell lysates were set aside for western blot analyses. (B) The BCA2-induced ubiquitination of Gag-GFP as well as native Gag was analyzed in side-by-side experiments. 293T cells were co-transfected in duplicate with each Gag construct, HA-BCA2, ΔRing BCA2 or empty vector. Cell lysates were set aside for regular western blotting, and the rest of the lysates were immunoprecipitated with an antibody anti-CA. Membranes were developed with an anti-Ubiquitin (Ub) antibody. (C) The ubiquitination of HIV-1 Gag, or HIV-1 Gag mutants, in cells expressing HA-BCA2 was also analyzed by immunoprecipitation. Cell lysates were set aside for western blot analyses and the rest of the samples were immunoprecipitated with an anti-CA specific antibody. Membranes were probed with an anti-Ubiquitin antibody. Similarly, the ubiquitination levels of HA-BCA2 were also determined. In this case, samples were immunoprecipitated using an anti-HA antibody. Results were confirmed in two additional independent assays. (D) A similar approach was used to analyze the ubiquitination levels of SIV Gag, or SIV Gag mutants, in the presence and absence of HA-BCA2. (E) Schematic representation of the putative lysine residues susceptible to become ubiquitinated by BCA2 in HIV-1 and SIV Gag (red). IP: immunoprecipitation. IB: immunoblot. WCL: whole cell lysate. V: empty vector.
    Figure Legend Snippet: Retroviral Gag proteins were tested for BCA2-induced ubiquitination by co-transfecting 293T cells with vectors expressing codon-optimized HIV-1 NL4-3, SIV mac 239 and Mo-MLV Gag proteins along with an expression vector coding for HA-BCA2 or empty vector. (A) Cell lysates were incubated with a polyubiquitin affinity resin and the bound fraction was eluted and analyzed by western blot. Membranes were probed with antibodies specific for p55/p24, MLV p30 and HA. Whole cell lysates were set aside for western blot analyses. (B) The BCA2-induced ubiquitination of Gag-GFP as well as native Gag was analyzed in side-by-side experiments. 293T cells were co-transfected in duplicate with each Gag construct, HA-BCA2, ΔRing BCA2 or empty vector. Cell lysates were set aside for regular western blotting, and the rest of the lysates were immunoprecipitated with an antibody anti-CA. Membranes were developed with an anti-Ubiquitin (Ub) antibody. (C) The ubiquitination of HIV-1 Gag, or HIV-1 Gag mutants, in cells expressing HA-BCA2 was also analyzed by immunoprecipitation. Cell lysates were set aside for western blot analyses and the rest of the samples were immunoprecipitated with an anti-CA specific antibody. Membranes were probed with an anti-Ubiquitin antibody. Similarly, the ubiquitination levels of HA-BCA2 were also determined. In this case, samples were immunoprecipitated using an anti-HA antibody. Results were confirmed in two additional independent assays. (D) A similar approach was used to analyze the ubiquitination levels of SIV Gag, or SIV Gag mutants, in the presence and absence of HA-BCA2. (E) Schematic representation of the putative lysine residues susceptible to become ubiquitinated by BCA2 in HIV-1 and SIV Gag (red). IP: immunoprecipitation. IB: immunoblot. WCL: whole cell lysate. V: empty vector.

    Techniques Used: Expressing, Plasmid Preparation, Incubation, Western Blot, Transfection, Construct, Immunoprecipitation

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    Thermo Fisher polyubiquitin affinity resin
    Polyubiquitin Affinity Resin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    polyubiquitin affinity resin a thermo scientific ubiquitin enrichment kit  (Thermo Fisher)
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    Thermo Fisher polyubiquitin affinity resin a thermo scientific ubiquitin enrichment kit
    Isolation of endogenously polyubiquitinated protein complexes by use of the <t>ubiquitin</t> enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the <t>polyubiquitin</t> affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.
    Polyubiquitin Affinity Resin A Thermo Scientific Ubiquitin Enrichment Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polyubiquitin affinity resin a thermo scientific ubiquitin enrichment kit/product/Thermo Fisher
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    polyubiquitin affinity resin a thermo scientific ubiquitin enrichment kit - by Bioz Stars, 2023-06
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    Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the polyubiquitin affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.

    Journal: Antioxidants & Redox Signaling

    Article Title: Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology

    doi: 10.1089/ars.2016.6686

    Figure Lengend Snippet: Isolation of endogenously polyubiquitinated protein complexes by use of the ubiquitin enrichment kit. A workflow for isolating endogenously ubiquitinylated protein complexes for proteomic analysis is shown. (A) Ubiquitinylated proteins were isolated from brain homogenates by IP. IP fraction (BF) and the supernatant (NBF) were loaded on an SDS-PAGE gel and stained for total protein expression (A1) and subsequently blotted on nitrocellulose membrane and stained with a polyclonal anti-Ub antibody (anti-Ub Ab) to determine enrichment efficiency (A2). (B) The IP fractions (BF and NBF) were further processed with the enrichment kit (Thermo) containing the polyubiquitin affinity resin (Poly-Ub AR). BF, NBF, and FT were loaded in SDS page (B1) and blotted with antiubiquitin antibody (B2). (C) Heat denaturation of high-affinity resin using high temperature (60°) was used to destroy the bond between resin and poly-Ub antibody. Poly-Ub was detected by performing a Western blot (agarose-only control; C1). BF, bound fraction; FT, flow-through; IP, immunoprecipitation; NBF, nonbound fraction; Poly-Ub, polymeric Ub; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Ub, ubiquitin.

    Article Snippet: Polyubiquitin affinity resin A Thermo Scientific Ubiquitin Enrichment Kit [89899] was used for the isolation and study of intracellular poly-Ub-modified proteins.

    Techniques: Isolation, SDS Page, Staining, Expressing, Western Blot, Immunoprecipitation, Polyacrylamide Gel Electrophoresis

    Poly-Ub levels. (A) Total levels of poly-Ub-bound proteins in DS (n = 6/group) and DS/AD (n = 6/group) cases compared with their age-matched controls (n = 6/group). (B, C) Increased poly-Ub of two types of ubiquitin chains [K63 (B) and K48 (C)] in both DS (n = 6/group) and DS/AD (n = 6/group) cases compared with their age-matched controls (n = 6/group). Representative bands are shown and protein levels (A−C, upper bands) were normalized per total protein load, named total load (A−C, lower bands). Densitometric values are shown as percentage of Ctr Y set as 100%. Mean ± SEM (DS vs. Ctr y *p < 0.05; DS/AD vs. DS **p < 0.01; DS/AD vs. Ctr O ***p < 0.001; DS vs. Ctr Y and DS/AD vs. DS [for K48] *p < 0.05; DS vs. Ctr Y, DS/AD vs. Ctr O, and DS/AD vs. DS [for K63] **p < 0.01; Ctr O vs. Ctr Y #p < 0.05 one-way ANOVA). AD, Alzheimer's disease; ANOVA, analysis of variance; DS, Down syndrome; DS/AD, Down syndrome with Alzheimer's disease; SEM, standard error of the mean.

    Journal: Antioxidants & Redox Signaling

    Article Title: Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology

    doi: 10.1089/ars.2016.6686

    Figure Lengend Snippet: Poly-Ub levels. (A) Total levels of poly-Ub-bound proteins in DS (n = 6/group) and DS/AD (n = 6/group) cases compared with their age-matched controls (n = 6/group). (B, C) Increased poly-Ub of two types of ubiquitin chains [K63 (B) and K48 (C)] in both DS (n = 6/group) and DS/AD (n = 6/group) cases compared with their age-matched controls (n = 6/group). Representative bands are shown and protein levels (A−C, upper bands) were normalized per total protein load, named total load (A−C, lower bands). Densitometric values are shown as percentage of Ctr Y set as 100%. Mean ± SEM (DS vs. Ctr y *p < 0.05; DS/AD vs. DS **p < 0.01; DS/AD vs. Ctr O ***p < 0.001; DS vs. Ctr Y and DS/AD vs. DS [for K48] *p < 0.05; DS vs. Ctr Y, DS/AD vs. Ctr O, and DS/AD vs. DS [for K63] **p < 0.01; Ctr O vs. Ctr Y #p < 0.05 one-way ANOVA). AD, Alzheimer's disease; ANOVA, analysis of variance; DS, Down syndrome; DS/AD, Down syndrome with Alzheimer's disease; SEM, standard error of the mean.

    Article Snippet: Polyubiquitin affinity resin A Thermo Scientific Ubiquitin Enrichment Kit [89899] was used for the isolation and study of intracellular poly-Ub-modified proteins.

    Techniques:

    List of Identified Polyubiquitinated Proteins by Mass Spectrometry

    Journal: Antioxidants & Redox Signaling

    Article Title: Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology

    doi: 10.1089/ars.2016.6686

    Figure Lengend Snippet: List of Identified Polyubiquitinated Proteins by Mass Spectrometry

    Article Snippet: Polyubiquitin affinity resin A Thermo Scientific Ubiquitin Enrichment Kit [89899] was used for the isolation and study of intracellular poly-Ub-modified proteins.

    Techniques:

    MUbiSiDa Search

    Journal: Antioxidants & Redox Signaling

    Article Title: Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology

    doi: 10.1089/ars.2016.6686

    Figure Lengend Snippet: MUbiSiDa Search

    Article Snippet: Polyubiquitin affinity resin A Thermo Scientific Ubiquitin Enrichment Kit [89899] was used for the isolation and study of intracellular poly-Ub-modified proteins.

    Techniques:

    UCH-L1. (A, B) All samples (n = 6/group) were immunoprecipitated with anti-UCH-L1. Immunoprecipitated proteins were separated on SDS-PAGE, transferred on nitrocellulose membranes, and probed with anti-poly-UbK63 (A, upper bands) and anti-poly-UbK48 (B, upper bands). (A) Shows increased levels of poly-UbK63 bound to UCH-L1 in DS and DS/AD compared with their matched controls. (B) Shows an increase in poly-UbK48 bound to UCH-L1 in DS/AD compared with DS. All the IP experiments were normalized on the total amount of UCH-L1 (indicated as Expr.; A, B, lower bands). Representative bands are shown. Densitometric values are shown as percentage of Ctr Y set as 100%. Mean ± SEM (DS vs. Ctr Y [for K63], DS/AD vs. DS [for K48] *p < 0.05; DS/AD vs. Ctr O [for K63] **p < 0.01 one-way ANOVA). UCH-L1, ubiquitin carboxyl-terminal hydrolase isozyme L1.

    Journal: Antioxidants & Redox Signaling

    Article Title: Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology

    doi: 10.1089/ars.2016.6686

    Figure Lengend Snippet: UCH-L1. (A, B) All samples (n = 6/group) were immunoprecipitated with anti-UCH-L1. Immunoprecipitated proteins were separated on SDS-PAGE, transferred on nitrocellulose membranes, and probed with anti-poly-UbK63 (A, upper bands) and anti-poly-UbK48 (B, upper bands). (A) Shows increased levels of poly-UbK63 bound to UCH-L1 in DS and DS/AD compared with their matched controls. (B) Shows an increase in poly-UbK48 bound to UCH-L1 in DS/AD compared with DS. All the IP experiments were normalized on the total amount of UCH-L1 (indicated as Expr.; A, B, lower bands). Representative bands are shown. Densitometric values are shown as percentage of Ctr Y set as 100%. Mean ± SEM (DS vs. Ctr Y [for K63], DS/AD vs. DS [for K48] *p < 0.05; DS/AD vs. Ctr O [for K63] **p < 0.01 one-way ANOVA). UCH-L1, ubiquitin carboxyl-terminal hydrolase isozyme L1.

    Article Snippet: Polyubiquitin affinity resin A Thermo Scientific Ubiquitin Enrichment Kit [89899] was used for the isolation and study of intracellular poly-Ub-modified proteins.

    Techniques: Immunoprecipitation, SDS Page