goat anti mis antibody  (Santa Cruz Biotechnology)

 
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    Name:
    MIS Antibody
    Description:
    Anti MIS Antibody B 11 is a mouse monoclonal IgG1 kappa light chain MIS antibody provided at 200 µg ml specific for an epitope mapping between amino acids 541 560 at the C terminus of MIS of human origin Anti MIS Antibody B 11 is recommended for detection of precursor and mature MIS of mouse rat and human origin by WB IP IF and ELISA also reactive with additional species including and equine canine bovine and porcine Anti MIS Antibody B 11 is available conjugated to agarose for IP HRP for WB IHC P and ELISA and to either phycoerythrin or FITC for IF IHC P and FCM also available conjugated to Alexa Fluor 488 Alexa Fluor 546 Alexa Fluor 594 or Alexa Fluor 647 for WB RGB IF IHC P and FCM and for use with RGB fluorescent imaging systems such as iBright FL1000 FluorChem Typhoon Azure and other comparable systems also available conjugated to Alexa Fluor 680 or Alexa Fluor 790 for WB NIR IF and FCM for use with Near Infrared NIR detection systems such as LI COR Odyssey iBright FL1000 FluorChem Typhoon Azure and other comparable systems blocking peptide sc 166752 P Contact our Technical Service Department or your local Distributor for more information on how to receive a FREE 10 µg sample of MIS B 11 sc 166752
    Catalog Number:
    SC-166752
    Price:
    None
    Category:
    Antibodies Primary Antibodies and ImmunoCruz Conjugates Growth Factors and Hormones MIS Antibodies MIS Antibody B 11
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    Structured Review

    Santa Cruz Biotechnology goat anti mis antibody
    Localization of <t>MIS,</t> Sox9, and <t>P450scc</t> proteins in DES-treated testis. IHC for MIS, Sox9, and P450scc proteins was performed on sections of fetal mouse testis at E12.5 (MIS) and E14.5 (MIS, Sox9, and P450scc) from mice exposed in utero to vehicle ( left panels ) or DES ( right panels ) (original magnification, ×132).
    Anti MIS Antibody B 11 is a mouse monoclonal IgG1 kappa light chain MIS antibody provided at 200 µg ml specific for an epitope mapping between amino acids 541 560 at the C terminus of MIS of human origin Anti MIS Antibody B 11 is recommended for detection of precursor and mature MIS of mouse rat and human origin by WB IP IF and ELISA also reactive with additional species including and equine canine bovine and porcine Anti MIS Antibody B 11 is available conjugated to agarose for IP HRP for WB IHC P and ELISA and to either phycoerythrin or FITC for IF IHC P and FCM also available conjugated to Alexa Fluor 488 Alexa Fluor 546 Alexa Fluor 594 or Alexa Fluor 647 for WB RGB IF IHC P and FCM and for use with RGB fluorescent imaging systems such as iBright FL1000 FluorChem Typhoon Azure and other comparable systems also available conjugated to Alexa Fluor 680 or Alexa Fluor 790 for WB NIR IF and FCM for use with Near Infrared NIR detection systems such as LI COR Odyssey iBright FL1000 FluorChem Typhoon Azure and other comparable systems blocking peptide sc 166752 P Contact our Technical Service Department or your local Distributor for more information on how to receive a FREE 10 µg sample of MIS B 11 sc 166752
    https://www.bioz.com/result/goat anti mis antibody/product/Santa Cruz Biotechnology
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    goat anti mis antibody - by Bioz Stars, 2021-09
    86/100 stars

    Images

    1) Product Images from "Effects of Gestational Diethylstilbestrol Treatment on Male and Female Gonads during Early Embryonic Development"

    Article Title: Effects of Gestational Diethylstilbestrol Treatment on Male and Female Gonads during Early Embryonic Development

    Journal: Endocrinology

    doi: 10.1210/en.2007-1599

    Localization of MIS, Sox9, and P450scc proteins in DES-treated testis. IHC for MIS, Sox9, and P450scc proteins was performed on sections of fetal mouse testis at E12.5 (MIS) and E14.5 (MIS, Sox9, and P450scc) from mice exposed in utero to vehicle ( left panels ) or DES ( right panels ) (original magnification, ×132).
    Figure Legend Snippet: Localization of MIS, Sox9, and P450scc proteins in DES-treated testis. IHC for MIS, Sox9, and P450scc proteins was performed on sections of fetal mouse testis at E12.5 (MIS) and E14.5 (MIS, Sox9, and P450scc) from mice exposed in utero to vehicle ( left panels ) or DES ( right panels ) (original magnification, ×132).

    Techniques Used: Immunohistochemistry, Mouse Assay, In Utero

    Localization of mRNAs of SF-1, MIS, and P450scc in DES-treated testis. In situ hybridization using 35 S-labeled cRNA probe was performed to analyze localization of mRNAs of SF-1, MIS, and P450scc on sections of fetal mouse testis at E14.5. Left and right panels show testis from mice treated in utero with vehicle and DES, respectively (original magnification, ×33).
    Figure Legend Snippet: Localization of mRNAs of SF-1, MIS, and P450scc in DES-treated testis. In situ hybridization using 35 S-labeled cRNA probe was performed to analyze localization of mRNAs of SF-1, MIS, and P450scc on sections of fetal mouse testis at E14.5. Left and right panels show testis from mice treated in utero with vehicle and DES, respectively (original magnification, ×33).

    Techniques Used: In Situ Hybridization, Labeling, Mouse Assay, In Utero

    Related Articles

    Mouse Assay:

    Article Title: ErbB4, a Receptor Tyrosine Kinase, Coordinates Organization of the Seminiferous Tubules in the Developing Testis
    Article Snippet: .. Testes were prepared from embryonic and adult mice, and the proteins were purified immediately and subjected to Western blotting using anti-ErbB4 (E200; Abcam), anti-Akt (sc-1618; Santa Cruz Biotechnology), anti-phospho-Akt (9271; Cell Signaling Technology), anti-Erk (9102; Cell Signaling Technology), anti-phospho-Erk (9101; Cell Signaling Technology), anti-MIS (sc-28912; Santa Cruz Biotechnology), or anti-Actin (sc-1616; Santa Cruz Biotechnology) antibodies as described previously ( ). ..

    Purification:

    Article Title: ErbB4, a Receptor Tyrosine Kinase, Coordinates Organization of the Seminiferous Tubules in the Developing Testis
    Article Snippet: .. Testes were prepared from embryonic and adult mice, and the proteins were purified immediately and subjected to Western blotting using anti-ErbB4 (E200; Abcam), anti-Akt (sc-1618; Santa Cruz Biotechnology), anti-phospho-Akt (9271; Cell Signaling Technology), anti-Erk (9102; Cell Signaling Technology), anti-phospho-Erk (9101; Cell Signaling Technology), anti-MIS (sc-28912; Santa Cruz Biotechnology), or anti-Actin (sc-1616; Santa Cruz Biotechnology) antibodies as described previously ( ). ..

    Western Blot:

    Article Title: ErbB4, a Receptor Tyrosine Kinase, Coordinates Organization of the Seminiferous Tubules in the Developing Testis
    Article Snippet: .. Testes were prepared from embryonic and adult mice, and the proteins were purified immediately and subjected to Western blotting using anti-ErbB4 (E200; Abcam), anti-Akt (sc-1618; Santa Cruz Biotechnology), anti-phospho-Akt (9271; Cell Signaling Technology), anti-Erk (9102; Cell Signaling Technology), anti-phospho-Erk (9101; Cell Signaling Technology), anti-MIS (sc-28912; Santa Cruz Biotechnology), or anti-Actin (sc-1616; Santa Cruz Biotechnology) antibodies as described previously ( ). ..

    Incubation:

    Article Title: Novel WT1 Missense Mutations in Han Chinese Women with Premature Ovarian Failure
    Article Snippet: .. The membranes were blocked, incubated with primary antibodies against ACTIN (catalog # 60008-1-lg, Proteintech, CH), WT1 (catalog # sc-192, Santa Cruz, USA), AMH (catalo # sc-166752, Santa Cruz), FSHR (catalog # 22665-1-AP, Proteintech), CYP19 (catalog # 16554-1-AP, Proteintech ) and CDH1 (catalog # 610182, BD Biosciences, USA ) overnight at 4 °C, washed, and then incubated with horseradish peroxidase-conjugated secondary antibodies. ..

    Immunoprecipitation:

    Article Title: Low paternal dietary folate alters the mouse sperm epigenome and is associated with negative pregnancy outcomes
    Article Snippet: .. Antibodies Primary antibodies used in this study were anti-H3K4-me1 (ab8895, Abcam, 1:1000), anti-H3K9-me1 (ab8896, 1:1000), anti-H3K9me3 (07-442, Millipore, 1:1000), anti-β-actin (A-1978, Sigma), anti-SYCP3 (ab15093; 1:100), anti-γH2AX (05-636; 1:100), anti-MIS (sc-6886, Santa Cruz; 1:50), anti-mouse VASA homologue (in house, 1:200), anti-3βHSD (sc-30820, 1:100) and anti-5-methylcytidine (BI-MECY-0100, Eurogentec, 10 μg per immunoprecipitation tube). ..

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    Santa Cruz Biotechnology goat anti mis antibody
    Localization of <t>MIS,</t> Sox9, and <t>P450scc</t> proteins in DES-treated testis. IHC for MIS, Sox9, and P450scc proteins was performed on sections of fetal mouse testis at E12.5 (MIS) and E14.5 (MIS, Sox9, and P450scc) from mice exposed in utero to vehicle ( left panels ) or DES ( right panels ) (original magnification, ×132).
    Goat Anti Mis Antibody, supplied by Santa Cruz Biotechnology, 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/goat anti mis antibody/product/Santa Cruz Biotechnology
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    goat anti mis antibody - by Bioz Stars, 2021-09
    86/100 stars
      Buy from Supplier

    99
    Santa Cruz Biotechnology goat anti mouse mis
    A model of <t>DKK3</t> mediated regulation of Sc maturation and spermatogenesis. In DKDM, LRP6 becomes available because of diminished DKK3 levels. Available LRP6 binds to Frizzled receptor forming Fz- LRP6 complex augmenting WNT signaling. As a result, GSK-3β is recruited to Frizzled receptor reducing its availability for binding to β-CATENIN, thereby minimizing phosphorylation and degradation of β-CATENIN. Accumulated cytoplasmic β-CATENIN translocates to nucleus and augments expression of different genes, including that of itself, WNT4 and <t>MIS</t> in Sc. Elevated levels of MIS interferes with the Sc maturation, hence, disturbing the balance between spermatogonial proliferation and differentiation leading to subfertility and/or infertility. Additionally, MIS is known to inhibit T production via suppression of CYP17 activity in Lc which might be the reason for reduced T levels in such mice. (eS-elongated spermatids, SSC-spermatogonial stem cells, Fz-Frizzled, rS-round spermatid, SpC-spermatocytes, PTc-peritubular cell, Tcf/Lcf -transcription factors).
    Goat Anti Mouse Mis, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/goat anti mouse mis/product/Santa Cruz Biotechnology
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    goat anti mouse mis - by Bioz Stars, 2021-09
    99/100 stars
      Buy from Supplier

    92
    Santa Cruz Biotechnology antigen purified goat anti mi 2 antibody
    The PYR complex contains SWI/SNF subunits, the NuRD subunit <t>Mi-2,</t> and the hematopoietic cell-restricted zinc finger protein Ikaros. Gel supershift assay was performed using MEL nuclear extract and labeled δ60ym DNA as a probe. Nuclear extract was preincubated for 1 h in the presence of antibody prior to the addition of probe. Supershifts are seen with anti-BAF57, anti-Mi-2, anti-SRG3, and anti-Ikaros antibodies.
    Antigen Purified Goat Anti Mi 2 Antibody, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/antigen purified goat anti mi 2 antibody/product/Santa Cruz Biotechnology
    Average 92 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    antigen purified goat anti mi 2 antibody - by Bioz Stars, 2021-09
    92/100 stars
      Buy from Supplier

    99
    Santa Cruz Biotechnology goat polyclonal anti mis amh c 20
    The PYR complex contains SWI/SNF subunits, the NuRD subunit <t>Mi-2,</t> and the hematopoietic cell-restricted zinc finger protein Ikaros. Gel supershift assay was performed using MEL nuclear extract and labeled δ60ym DNA as a probe. Nuclear extract was preincubated for 1 h in the presence of antibody prior to the addition of probe. Supershifts are seen with anti-BAF57, anti-Mi-2, anti-SRG3, and anti-Ikaros antibodies.
    Goat Polyclonal Anti Mis Amh C 20, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/goat polyclonal anti mis amh c 20/product/Santa Cruz Biotechnology
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    goat polyclonal anti mis amh c 20 - by Bioz Stars, 2021-09
    99/100 stars
      Buy from Supplier

    Image Search Results


    Localization of MIS, Sox9, and P450scc proteins in DES-treated testis. IHC for MIS, Sox9, and P450scc proteins was performed on sections of fetal mouse testis at E12.5 (MIS) and E14.5 (MIS, Sox9, and P450scc) from mice exposed in utero to vehicle ( left panels ) or DES ( right panels ) (original magnification, ×132).

    Journal: Endocrinology

    Article Title: Effects of Gestational Diethylstilbestrol Treatment on Male and Female Gonads during Early Embryonic Development

    doi: 10.1210/en.2007-1599

    Figure Lengend Snippet: Localization of MIS, Sox9, and P450scc proteins in DES-treated testis. IHC for MIS, Sox9, and P450scc proteins was performed on sections of fetal mouse testis at E12.5 (MIS) and E14.5 (MIS, Sox9, and P450scc) from mice exposed in utero to vehicle ( left panels ) or DES ( right panels ) (original magnification, ×132).

    Article Snippet: The sections were incubated in a blocking solution, 5% Block Ace (Dainippon Pharmaceutical Co., Ltd., Tokyo, Japan), at room temperature for 30 min and subsequently incubated with a diluted primary antibody overnight at 4 C. Rabbit anti-SF-1 antibody (17; 1:5000 dilution), rabbit anti-P450scc antibody (AB1244, Chemicon International, Inc., Temecula, CA; 1:5000 dilution), goat anti-MIS antibody (sc-6886; Santa Cruz Biotechnology, Inc., Santa Cruz, CA; 1:5000 dilution), and rabbit anti-Sox9 antibody (sc-20095; Santa Cruz; 1:500 dilution) were used as primary antibodies.

    Techniques: Immunohistochemistry, Mouse Assay, In Utero

    Localization of mRNAs of SF-1, MIS, and P450scc in DES-treated testis. In situ hybridization using 35 S-labeled cRNA probe was performed to analyze localization of mRNAs of SF-1, MIS, and P450scc on sections of fetal mouse testis at E14.5. Left and right panels show testis from mice treated in utero with vehicle and DES, respectively (original magnification, ×33).

    Journal: Endocrinology

    Article Title: Effects of Gestational Diethylstilbestrol Treatment on Male and Female Gonads during Early Embryonic Development

    doi: 10.1210/en.2007-1599

    Figure Lengend Snippet: Localization of mRNAs of SF-1, MIS, and P450scc in DES-treated testis. In situ hybridization using 35 S-labeled cRNA probe was performed to analyze localization of mRNAs of SF-1, MIS, and P450scc on sections of fetal mouse testis at E14.5. Left and right panels show testis from mice treated in utero with vehicle and DES, respectively (original magnification, ×33).

    Article Snippet: The sections were incubated in a blocking solution, 5% Block Ace (Dainippon Pharmaceutical Co., Ltd., Tokyo, Japan), at room temperature for 30 min and subsequently incubated with a diluted primary antibody overnight at 4 C. Rabbit anti-SF-1 antibody (17; 1:5000 dilution), rabbit anti-P450scc antibody (AB1244, Chemicon International, Inc., Temecula, CA; 1:5000 dilution), goat anti-MIS antibody (sc-6886; Santa Cruz Biotechnology, Inc., Santa Cruz, CA; 1:5000 dilution), and rabbit anti-Sox9 antibody (sc-20095; Santa Cruz; 1:500 dilution) were used as primary antibodies.

    Techniques: In Situ Hybridization, Labeling, Mouse Assay, In Utero

    A model of DKK3 mediated regulation of Sc maturation and spermatogenesis. In DKDM, LRP6 becomes available because of diminished DKK3 levels. Available LRP6 binds to Frizzled receptor forming Fz- LRP6 complex augmenting WNT signaling. As a result, GSK-3β is recruited to Frizzled receptor reducing its availability for binding to β-CATENIN, thereby minimizing phosphorylation and degradation of β-CATENIN. Accumulated cytoplasmic β-CATENIN translocates to nucleus and augments expression of different genes, including that of itself, WNT4 and MIS in Sc. Elevated levels of MIS interferes with the Sc maturation, hence, disturbing the balance between spermatogonial proliferation and differentiation leading to subfertility and/or infertility. Additionally, MIS is known to inhibit T production via suppression of CYP17 activity in Lc which might be the reason for reduced T levels in such mice. (eS-elongated spermatids, SSC-spermatogonial stem cells, Fz-Frizzled, rS-round spermatid, SpC-spermatocytes, PTc-peritubular cell, Tcf/Lcf -transcription factors).

    Journal: PLoS ONE

    Article Title: Dickkopf Homolog 3 (DKK3) Plays a Crucial Role Upstream of WNT/?-CATENIN Signaling for Sertoli Cell Mediated Regulation of Spermatogenesis

    doi: 10.1371/journal.pone.0063603

    Figure Lengend Snippet: A model of DKK3 mediated regulation of Sc maturation and spermatogenesis. In DKDM, LRP6 becomes available because of diminished DKK3 levels. Available LRP6 binds to Frizzled receptor forming Fz- LRP6 complex augmenting WNT signaling. As a result, GSK-3β is recruited to Frizzled receptor reducing its availability for binding to β-CATENIN, thereby minimizing phosphorylation and degradation of β-CATENIN. Accumulated cytoplasmic β-CATENIN translocates to nucleus and augments expression of different genes, including that of itself, WNT4 and MIS in Sc. Elevated levels of MIS interferes with the Sc maturation, hence, disturbing the balance between spermatogonial proliferation and differentiation leading to subfertility and/or infertility. Additionally, MIS is known to inhibit T production via suppression of CYP17 activity in Lc which might be the reason for reduced T levels in such mice. (eS-elongated spermatids, SSC-spermatogonial stem cells, Fz-Frizzled, rS-round spermatid, SpC-spermatocytes, PTc-peritubular cell, Tcf/Lcf -transcription factors).

    Article Snippet: Following primary and secondary antibodies were used: rabbit anti-mouse DKK3 (Santa Cruz Biotechnology, CA,USA), goat anti-mouse MIS (Santa Cruz Biotechnology, CA,USA), mouse anti-GFP (Abcam, Cambridge, MA,USA), β-CATENIN anti-mouse antibody (Santa Cruz Biotechnology, CA,USA), Alexa Fluor 488 anti-mouse secondary antibody (Invitrogen, Life Technologies, NY, USA), FITC anti-rabbit secondary antibody (Abcam, Cambridge, MA,USA), Cy5 anti-goat antibody F(ab)2 (The Jackson ImmunoResearch Laboratory, PA,USA).

    Techniques: Binding Assay, Expressing, Activity Assay, Mouse Assay

    The PYR complex contains SWI/SNF subunits, the NuRD subunit Mi-2, and the hematopoietic cell-restricted zinc finger protein Ikaros. Gel supershift assay was performed using MEL nuclear extract and labeled δ60ym DNA as a probe. Nuclear extract was preincubated for 1 h in the presence of antibody prior to the addition of probe. Supershifts are seen with anti-BAF57, anti-Mi-2, anti-SRG3, and anti-Ikaros antibodies.

    Journal: Molecular and Cellular Biology

    Article Title: An Ikaros-Containing Chromatin-Remodeling Complex in Adult-Type Erythroid Cells

    doi:

    Figure Lengend Snippet: The PYR complex contains SWI/SNF subunits, the NuRD subunit Mi-2, and the hematopoietic cell-restricted zinc finger protein Ikaros. Gel supershift assay was performed using MEL nuclear extract and labeled δ60ym DNA as a probe. Nuclear extract was preincubated for 1 h in the presence of antibody prior to the addition of probe. Supershifts are seen with anti-BAF57, anti-Mi-2, anti-SRG3, and anti-Ikaros antibodies.

    Article Snippet: An anti-Mi-2 immunoaffinity column was prepared using antigen-purified goat anti-Mi-2 antibody obtained commercially (Santa Cruz Biotechnology).

    Techniques: Labeling

    Ikaros functions as the DNA-binding subunit of the PYR complex. (A) PYR complex has a 60- to 70-kDa DNA-binding subunit as detected by photoactivated cross-linking using both MEL crude nuclear extract and Sephacryl S300 chromatography fractions containing peak PYR complex DNA-binding activity. The band corresponding to this activity is competed away with excess unlabeled δ99 DNA but not by consensus (YY1) or mutant (mut YY1) YY1-binding-site DNA. (B) The PYR complex and GST-Ikaros have similar DNA-binding specificities. Upper panel: Gel shift assay using labeled δ60ym DNA probe, GST-Ikaros protein (left lanes), and partially purified PYR complex (DNA-cellulose column peak fractions [DNA-Cell], right lanes). Both GST-Ikaros and the PYR complex bind δ60ym, and these binding activities are competed away with unlabeled δ60ym DNA and DNA from a previously described high-affinity Ikaros-binding site, IKBS4, but not with YY1-binding- site DNA. The various competitor DNAs were used at 0-, 10-, and 100-fold molar excesses. Lower panel: Gel shift assay using labeled IKBS4 DNA probe, GST-Ikaros protein (left), and DNA cellulose-purified PYR complex (right). Both GST-Ikaros and the PYR complex bind labeled IKBS4, and these binding activities are competed away with unlabeled IKBS4 and δ60ym, but not with YY1-binding-site, DNA. (C) Antibodies to Ikaros and Mi-2 supershift the activity in DNA-cellulose-purified fractions that binds δ60ym (left lanes) and IKBS4 (right lanes).

    Journal: Molecular and Cellular Biology

    Article Title: An Ikaros-Containing Chromatin-Remodeling Complex in Adult-Type Erythroid Cells

    doi:

    Figure Lengend Snippet: Ikaros functions as the DNA-binding subunit of the PYR complex. (A) PYR complex has a 60- to 70-kDa DNA-binding subunit as detected by photoactivated cross-linking using both MEL crude nuclear extract and Sephacryl S300 chromatography fractions containing peak PYR complex DNA-binding activity. The band corresponding to this activity is competed away with excess unlabeled δ99 DNA but not by consensus (YY1) or mutant (mut YY1) YY1-binding-site DNA. (B) The PYR complex and GST-Ikaros have similar DNA-binding specificities. Upper panel: Gel shift assay using labeled δ60ym DNA probe, GST-Ikaros protein (left lanes), and partially purified PYR complex (DNA-cellulose column peak fractions [DNA-Cell], right lanes). Both GST-Ikaros and the PYR complex bind δ60ym, and these binding activities are competed away with unlabeled δ60ym DNA and DNA from a previously described high-affinity Ikaros-binding site, IKBS4, but not with YY1-binding- site DNA. The various competitor DNAs were used at 0-, 10-, and 100-fold molar excesses. Lower panel: Gel shift assay using labeled IKBS4 DNA probe, GST-Ikaros protein (left), and DNA cellulose-purified PYR complex (right). Both GST-Ikaros and the PYR complex bind labeled IKBS4, and these binding activities are competed away with unlabeled IKBS4 and δ60ym, but not with YY1-binding-site, DNA. (C) Antibodies to Ikaros and Mi-2 supershift the activity in DNA-cellulose-purified fractions that binds δ60ym (left lanes) and IKBS4 (right lanes).

    Article Snippet: An anti-Mi-2 immunoaffinity column was prepared using antigen-purified goat anti-Mi-2 antibody obtained commercially (Santa Cruz Biotechnology).

    Techniques: Binding Assay, Chromatography, Activity Assay, Mutagenesis, Electrophoretic Mobility Shift Assay, Labeling, Purification

    Ikaros, NuRD, and SWI/SNF proteins coimmunopurify from chromatography fractions containing peak PYR complex DNA-binding activity. (A) Ikaros and Mi-2 coimmunoprecipitate from reactive yellow 3-agarose-fractionated MEL extract. (Left panel) Western blot analysis with rabbit polyclonal anti-Ikaros antibody. Mouse monoclonal antibodies used for immunoprecipitation (IP) are indicated above lanes 2 through 4. The sample for lane 1 was 1/10 the volume of reactive yellow 3-agarose-purified PYR complex (yellow 3 fraction 67 [Y3]) before precipitation as a positive control for the Ikaros signal (Ik-1, 65 kDa; Ik-2, 55 kDa). Weak cross-reaction of the anti-rabbit IgG secondary antibody with the mouse IgG heavy chain results in a faint band of approximately 50 kDa in lanes 2 through 4. (Right panel) Western blot analysis with an anti-Mi-2 rabbit polyclonal antibody. Lane 1, reactive yellow 3-agarose fraction 67; lane 2, anti-Aiolos precipitate; lane 3, anti-Mi-2 precipitate; lane 4, anti-Ikaros precipitate. (B) Ikaros, BRG1, and Mi-2 copurify with BAF57 off an anti-BAF57 immunoaffinity column. The start material (Start) was DNA-cellulose-purified PYR complex. The vast bulk of the protein loaded onto the column does not bind (FT). This was confirmed by silver staining (data not shown). The column was washed twice with 0.1 M NaCl buffer (0.1 A and B) and twice with 0.5 M NaCl buffer (0.5 A and B) and then eluted with 0.1 M glycine (pH 2.5) (Elute). BAF57 and Ikaros, particularly Ikaros isoform 2, bind very tightly to the column, withstanding the 0.5 M salt washes. Both BRG1 and Mi-2 bind to the column with somewhat lower affinity, eluting in the 0.5 M salt and pH 2.5 glycine fractions. (C) Ikaros, NuRD, and SWI/SNF proteins copurify with Mi-2 off an anti-Mi-2 immunoaffinity column. The start material (Start) was DNA-cellulose-purified PYR complex. Silver-stained gels showed that the vast bulk of the protein loaded onto the column does not bind (data not shown). The column was washed and eluted as described for panel B. Mi-2 binds very tightly to the column, with much of the Mi-2 remaining on the column after the pH 2.5 elution step. HDAC2, Ikaros isoforms 1 and 2, and SWI/SNF proteins (BAF57, BRG1, and BAF155) bind to the column with high affinity, eluting only in the pH 2.5 glycine fraction.

    Journal: Molecular and Cellular Biology

    Article Title: An Ikaros-Containing Chromatin-Remodeling Complex in Adult-Type Erythroid Cells

    doi:

    Figure Lengend Snippet: Ikaros, NuRD, and SWI/SNF proteins coimmunopurify from chromatography fractions containing peak PYR complex DNA-binding activity. (A) Ikaros and Mi-2 coimmunoprecipitate from reactive yellow 3-agarose-fractionated MEL extract. (Left panel) Western blot analysis with rabbit polyclonal anti-Ikaros antibody. Mouse monoclonal antibodies used for immunoprecipitation (IP) are indicated above lanes 2 through 4. The sample for lane 1 was 1/10 the volume of reactive yellow 3-agarose-purified PYR complex (yellow 3 fraction 67 [Y3]) before precipitation as a positive control for the Ikaros signal (Ik-1, 65 kDa; Ik-2, 55 kDa). Weak cross-reaction of the anti-rabbit IgG secondary antibody with the mouse IgG heavy chain results in a faint band of approximately 50 kDa in lanes 2 through 4. (Right panel) Western blot analysis with an anti-Mi-2 rabbit polyclonal antibody. Lane 1, reactive yellow 3-agarose fraction 67; lane 2, anti-Aiolos precipitate; lane 3, anti-Mi-2 precipitate; lane 4, anti-Ikaros precipitate. (B) Ikaros, BRG1, and Mi-2 copurify with BAF57 off an anti-BAF57 immunoaffinity column. The start material (Start) was DNA-cellulose-purified PYR complex. The vast bulk of the protein loaded onto the column does not bind (FT). This was confirmed by silver staining (data not shown). The column was washed twice with 0.1 M NaCl buffer (0.1 A and B) and twice with 0.5 M NaCl buffer (0.5 A and B) and then eluted with 0.1 M glycine (pH 2.5) (Elute). BAF57 and Ikaros, particularly Ikaros isoform 2, bind very tightly to the column, withstanding the 0.5 M salt washes. Both BRG1 and Mi-2 bind to the column with somewhat lower affinity, eluting in the 0.5 M salt and pH 2.5 glycine fractions. (C) Ikaros, NuRD, and SWI/SNF proteins copurify with Mi-2 off an anti-Mi-2 immunoaffinity column. The start material (Start) was DNA-cellulose-purified PYR complex. Silver-stained gels showed that the vast bulk of the protein loaded onto the column does not bind (data not shown). The column was washed and eluted as described for panel B. Mi-2 binds very tightly to the column, with much of the Mi-2 remaining on the column after the pH 2.5 elution step. HDAC2, Ikaros isoforms 1 and 2, and SWI/SNF proteins (BAF57, BRG1, and BAF155) bind to the column with high affinity, eluting only in the pH 2.5 glycine fraction.

    Article Snippet: An anti-Mi-2 immunoaffinity column was prepared using antigen-purified goat anti-Mi-2 antibody obtained commercially (Santa Cruz Biotechnology).

    Techniques: Chromatography, Binding Assay, Activity Assay, Western Blot, Immunoprecipitation, Purification, Positive Control, Silver Staining, Staining