rabbit polyclonal antibody  (Novus Biologicals)

 
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  • 99
    Name:
    Rabbit anti Golden Syrian Hamster IgM Secondary Antibody FITC
    Description:
    The Rabbit anti Golden Syrian Hamster IgM Secondary Antibody FITC from Novus Biologicals is a rabbit polyclonal antibody to IgM This antibody reacts with golden syrian hamster The Rabbit anti Golden Syrian Hamster IgM Secondary Antibody FITC has been validated for the following applications Flow Cytometry Immunocytochemistry Immunofluorescence Fluorophore linked immunosorbent assay
    Catalog Number:
    NBP1-73862-10mg
    Price:
    279
    Host:
    Rabbit
    Purity:
    Ion exchange chromatography
    Conjugate:
    FITC
    Immunogen:
    Hamster IgM mu heavy chain
    Size:
    10 mg
    Category:
    Secondary Antibodies
    Buy from Supplier


    Structured Review

    Novus Biologicals rabbit polyclonal antibody
    Association with hWDR79 is required for CB localization of a human scaRNA. ( A ) Subcellular localization of hWDR79 in HeLa cells using rabbit <t>polyclonal</t> antibody. ( B ) Mislocalization of a CAB box mutant ACA scaRNA. Either WT or L1+2mt ACA57 (see
    The Rabbit anti Golden Syrian Hamster IgM Secondary Antibody FITC from Novus Biologicals is a rabbit polyclonal antibody to IgM This antibody reacts with golden syrian hamster The Rabbit anti Golden Syrian Hamster IgM Secondary Antibody FITC has been validated for the following applications Flow Cytometry Immunocytochemistry Immunofluorescence Fluorophore linked immunosorbent assay
    https://www.bioz.com/result/rabbit polyclonal antibody/product/Novus Biologicals
    Average 99 stars, based on 36 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal antibody - by Bioz Stars, 2021-01
    99/100 stars

    Images

    1) Product Images from "A Conserved WD40 Protein Binds the Cajal Body Localization Signal of scaRNP Particles"

    Article Title: A Conserved WD40 Protein Binds the Cajal Body Localization Signal of scaRNP Particles

    Journal: Molecular cell

    doi: 10.1016/j.molcel.2009.02.020

    Association with hWDR79 is required for CB localization of a human scaRNA. ( A ) Subcellular localization of hWDR79 in HeLa cells using rabbit polyclonal antibody. ( B ) Mislocalization of a CAB box mutant ACA scaRNA. Either WT or L1+2mt ACA57 (see
    Figure Legend Snippet: Association with hWDR79 is required for CB localization of a human scaRNA. ( A ) Subcellular localization of hWDR79 in HeLa cells using rabbit polyclonal antibody. ( B ) Mislocalization of a CAB box mutant ACA scaRNA. Either WT or L1+2mt ACA57 (see

    Techniques Used: Mutagenesis

    2) Product Images from "Telomerase inhibition by siRNA causes senescence and apoptosis in Barrett's adenocarcinoma cells: mechanism and therapeutic potential"

    Article Title: Telomerase inhibition by siRNA causes senescence and apoptosis in Barrett's adenocarcinoma cells: mechanism and therapeutic potential

    Journal: Molecular Cancer

    doi: 10.1186/1476-4598-4-24

    Apoptosis following treatment of BEAC cells with telomerase siRNAs. SEG-1 cells, transfected with control (Cont) or telomerase specific (Tel) siRNAs were cultured for three weeks and analyzed for apoptosis by evaluating annexin labeling or cleavage of PARP. A. Cells were mixed with annexin V-BIOTIN and treated sequentially with streptavidin conjugated to fluorescein isothiocyanate (FITC) and propidium iodide (PI). Apoptotic cells within the same microscopic field were viewed and photographed by phase contrast ( PC ) or by fluorescence emitted at 518 nm ( FITC filter). Using the FITC filter, early apoptotic cells (positive for Annexin V-Biotin-FITC staining) appear bright green. B. SEG-1 cells were treated exactly as described for panel A and analysed for cleavage of poly(ADP-ribose) polymerase (PARP), a marker for apoptosis. PARP was identified by western blotting using a rabbit polyclonal antibody against PARP (Santa Cruz). C. Bar graph showing percentage of PARP found to be cleaved in cells treated with control or telomerase siRNAs.
    Figure Legend Snippet: Apoptosis following treatment of BEAC cells with telomerase siRNAs. SEG-1 cells, transfected with control (Cont) or telomerase specific (Tel) siRNAs were cultured for three weeks and analyzed for apoptosis by evaluating annexin labeling or cleavage of PARP. A. Cells were mixed with annexin V-BIOTIN and treated sequentially with streptavidin conjugated to fluorescein isothiocyanate (FITC) and propidium iodide (PI). Apoptotic cells within the same microscopic field were viewed and photographed by phase contrast ( PC ) or by fluorescence emitted at 518 nm ( FITC filter). Using the FITC filter, early apoptotic cells (positive for Annexin V-Biotin-FITC staining) appear bright green. B. SEG-1 cells were treated exactly as described for panel A and analysed for cleavage of poly(ADP-ribose) polymerase (PARP), a marker for apoptosis. PARP was identified by western blotting using a rabbit polyclonal antibody against PARP (Santa Cruz). C. Bar graph showing percentage of PARP found to be cleaved in cells treated with control or telomerase siRNAs.

    Techniques Used: Transfection, Cell Culture, Labeling, Fluorescence, Staining, Marker, Western Blot

    Induction of apoptosis proteins following treatment of SEG-1 cells to telomerase specific siRNAs. SEG-1 cells, transfected with control (Cont) or telomerase specific (Tel) siRNAs were cultured for two weeks and analyzed for protein expression by western blotting. The expression of p63, TNF-related apoptosis inducing ligand (TRAIL), TNF ligand superfamily, member 6 (FASL), and p16 was monitored using mouse monoclonal anti-p63 (Zymed), rabbit polyclonal anti-TRAIL (Santa Cruz), mouse monoclonal anti-FASL (Cell Science), and mouse monoclonal anti-p16 (Sigma) antibodies, respectively. A. Western blot showing expression of p63 and TRAIL in control (Cont) and telomerase (Tel) siRNA treated cells. B. Bar graph showing relative expression of p63 and TRAIL following normalization with glyceraldehyde-3-phosphate dehydrogenase (GAPDH). C. Western blot showing expression of FASL and p16 in control (Cont) and telomerase (Tel) siRNA treated cells. D. Bar graph showing relative expression of FASL and p16 following normalization with glyceraldehyde-3-phosphate dehydrogenase (GAPDH).
    Figure Legend Snippet: Induction of apoptosis proteins following treatment of SEG-1 cells to telomerase specific siRNAs. SEG-1 cells, transfected with control (Cont) or telomerase specific (Tel) siRNAs were cultured for two weeks and analyzed for protein expression by western blotting. The expression of p63, TNF-related apoptosis inducing ligand (TRAIL), TNF ligand superfamily, member 6 (FASL), and p16 was monitored using mouse monoclonal anti-p63 (Zymed), rabbit polyclonal anti-TRAIL (Santa Cruz), mouse monoclonal anti-FASL (Cell Science), and mouse monoclonal anti-p16 (Sigma) antibodies, respectively. A. Western blot showing expression of p63 and TRAIL in control (Cont) and telomerase (Tel) siRNA treated cells. B. Bar graph showing relative expression of p63 and TRAIL following normalization with glyceraldehyde-3-phosphate dehydrogenase (GAPDH). C. Western blot showing expression of FASL and p16 in control (Cont) and telomerase (Tel) siRNA treated cells. D. Bar graph showing relative expression of FASL and p16 following normalization with glyceraldehyde-3-phosphate dehydrogenase (GAPDH).

    Techniques Used: Transfection, Cell Culture, Expressing, Western Blot

    Effect of telomerase-specific siRNAs on telomerase expression and activity in SEG-1 cells. ( A ). Control (Cont) or telomerase specific (Tel) SiRNAs were transfected into Barrett's associated adenocarcinoma cells (SEG-1) and the cells were analyzed for protein levels of telomerase at day 7 by immunostaining, using a rabbit polyclonal antibody against telomerase. Antigen-antibody complex was located by incubation of cells with FITC-labeled anti-rabbit secondary antibody. Transfected cells within the same microscopic field were viewed and photographed by phase contrast (PC) or by fluorescence emitted at 518 nm (FITC filter). Using the FITC filter, telomerase positive cells appear bright green. ( B ). SEG-1 cells were treated exactly as described for panel
    Figure Legend Snippet: Effect of telomerase-specific siRNAs on telomerase expression and activity in SEG-1 cells. ( A ). Control (Cont) or telomerase specific (Tel) SiRNAs were transfected into Barrett's associated adenocarcinoma cells (SEG-1) and the cells were analyzed for protein levels of telomerase at day 7 by immunostaining, using a rabbit polyclonal antibody against telomerase. Antigen-antibody complex was located by incubation of cells with FITC-labeled anti-rabbit secondary antibody. Transfected cells within the same microscopic field were viewed and photographed by phase contrast (PC) or by fluorescence emitted at 518 nm (FITC filter). Using the FITC filter, telomerase positive cells appear bright green. ( B ). SEG-1 cells were treated exactly as described for panel "A" and telomerase expression was monitored by western blot analysis using a rabbit polyclonal anti-telomerase antibody. ( C ). Bar graph showing relative expression of telomerase in control (Cont) or telomerase (Tel) siRNA treated SEG-1 cells, as assessed by western blot analysis (shown in panel B). ( D ). SEG-1 cells were transfected with control (Cont) or telomerase specific (Tel) SiRNAs and evaluated for telomerase activity at days 1, 3, and 5 after transfection. Telomerase activity was evaluated using TRAPEZE R XL Telomerase Detection Kit, a highly sensitive, quantitative, and non-isotopic version of the original Telomeric Repeat Amplification Protocol. Briefly the cell lysates (1000 cell-equivalents) were mixed with TRAPEZE R XL reaction mix containing Amplifuor™ primers, incubated for 30 min at 30°C, and telomerase products generated by PCR were quantitated using a Fluorescence Plate Reader. Telomerase activity in SEG-1 cells treated with control siRNAs for 5 days and the cells treated with telomerase specific siRNAs for days 1, 3, and 5, is shown in TPG (total product generated) units.

    Techniques Used: Expressing, Activity Assay, Transfection, Immunostaining, Incubation, Labeling, Fluorescence, Western Blot, Non-Isotopic Labeling, Amplification, Generated, Polymerase Chain Reaction

    3) Product Images from "Regulated Expression of Rhox8 in the Mouse Ovary: Evidence for the Role of Progesterone and RHOX5 in Granulosa Cells 1"

    Article Title: Regulated Expression of Rhox8 in the Mouse Ovary: Evidence for the Role of Progesterone and RHOX5 in Granulosa Cells 1

    Journal: Biology of Reproduction

    doi: 10.1095/biolreprod.112.103267

    Immunolocalization of RHOX8 protein in mice. A ) Serial ovarian sections from eCG primed mice, after 8 h hCG-treated mice were incubated with either preimmune serum (1:250 dilution) or RHOX8 polyclonal antiserum (1:2500 dilution) ( B ) from Imgenex rabbit no.2223B immunized with a RHOX8 amino domain peptide. Visualization by diaminobenzidine staining reveals that most of the RHOX8 protein is localized to granulosa cells of large antral follicles (granulosa cells are shown at higher magnification in C−F ). Weak RHOX8 protein staining can be observed in some granulosa cells of preantral follicles ( C , arrowheads) and is absent in cumulus granulosa cells ( C and E , arrows) surrounding the oocyte (O). D ) Nuclei of thecal cells (arrowheads) are also positive for RHOX8. F ) RHOX8 expression remains in the corpus luteum (CL) of postovulatory follicles.
    Figure Legend Snippet: Immunolocalization of RHOX8 protein in mice. A ) Serial ovarian sections from eCG primed mice, after 8 h hCG-treated mice were incubated with either preimmune serum (1:250 dilution) or RHOX8 polyclonal antiserum (1:2500 dilution) ( B ) from Imgenex rabbit no.2223B immunized with a RHOX8 amino domain peptide. Visualization by diaminobenzidine staining reveals that most of the RHOX8 protein is localized to granulosa cells of large antral follicles (granulosa cells are shown at higher magnification in C−F ). Weak RHOX8 protein staining can be observed in some granulosa cells of preantral follicles ( C , arrowheads) and is absent in cumulus granulosa cells ( C and E , arrows) surrounding the oocyte (O). D ) Nuclei of thecal cells (arrowheads) are also positive for RHOX8. F ) RHOX8 expression remains in the corpus luteum (CL) of postovulatory follicles.

    Techniques Used: Mouse Assay, Incubation, Staining, Expressing

    4) Product Images from "TLR9-induced interferon ? is associated with protection from gammaherpesvirus-induced exacerbation of lung fibrosis"

    Article Title: TLR9-induced interferon ? is associated with protection from gammaherpesvirus-induced exacerbation of lung fibrosis

    Journal: Fibrogenesis & Tissue Repair

    doi: 10.1186/1755-1536-4-18

    Gamma herpesvirus 68 (γHV68) replicates in alveolar epithelial cells (AECs) . (A) Wild-type mice were infected with 5 × 10 4 PFU γHV68 on day 0. On day 7 after infection, frozen sections were prepared, and stained with a rabbit polyclonal antisera against γHV68, or with non-immune rabbit sera as control. The goat anti-rabbit secondary was linked to alkaline phosphatase. Vivid replication of γHV68 is visible in alveolar lining cells (original magnification × 100). Sections shown are representative of four mice examined. (B) AECs were isolated from lungs of Balb/c or TLR-9 -/- mice treated with bleomycin plus γHV68 on day 21, and were cultured on fibronectin-coated slides (TiterTek). Sections were stained with antibodies (M30 Cytodeath), and the number of positive cells per high power field (HPF; ×400) were calculated (n = 30 HPF per genotype). (C) AECs were isolated from Balb/c or TLR-9 -/- mice and cells were infected in vitro with 0.01 or 0.001 PFU γHV68 for 48 hours. Cell lysates were then analyzed for cleaved caspase 3 by western blotting. Data are from one experiment, representative of two.
    Figure Legend Snippet: Gamma herpesvirus 68 (γHV68) replicates in alveolar epithelial cells (AECs) . (A) Wild-type mice were infected with 5 × 10 4 PFU γHV68 on day 0. On day 7 after infection, frozen sections were prepared, and stained with a rabbit polyclonal antisera against γHV68, or with non-immune rabbit sera as control. The goat anti-rabbit secondary was linked to alkaline phosphatase. Vivid replication of γHV68 is visible in alveolar lining cells (original magnification × 100). Sections shown are representative of four mice examined. (B) AECs were isolated from lungs of Balb/c or TLR-9 -/- mice treated with bleomycin plus γHV68 on day 21, and were cultured on fibronectin-coated slides (TiterTek). Sections were stained with antibodies (M30 Cytodeath), and the number of positive cells per high power field (HPF; ×400) were calculated (n = 30 HPF per genotype). (C) AECs were isolated from Balb/c or TLR-9 -/- mice and cells were infected in vitro with 0.01 or 0.001 PFU γHV68 for 48 hours. Cell lysates were then analyzed for cleaved caspase 3 by western blotting. Data are from one experiment, representative of two.

    Techniques Used: Mouse Assay, Infection, Staining, Isolation, Cell Culture, In Vitro, Western Blot

    5) Product Images from "Carboxy-terminal deletion of the HDL receptor reduces receptor levels in liver and steroidogenic tissues, induces hypercholesterolemia, and causes fatal heart disease"

    Article Title: Carboxy-terminal deletion of the HDL receptor reduces receptor levels in liver and steroidogenic tissues, induces hypercholesterolemia, and causes fatal heart disease

    Journal: American Journal of Physiology - Heart and Circulatory Physiology

    doi: 10.1152/ajpheart.00463.2016

    Effects of the SR-BIΔCT mutation on the hepatic expression of SR-BI and SR-BII proteins. A and B : immunoblotting analyses of liver lysates (∼30 μg protein) from male WT and SR-BIΔCT (ΔCT) 6- to 10-wk-old mice. Results in female mice were virtually identical. Bands were visualized by chemiluminescence. A : proteins were detected with either a SR-BI-specific anti-carboxy-terminal peptide antibody [SR-BI (anti-C-term)] or a rabbit polyclonal antibody that recognizes the extracellular domains of both SR-BI and its minor splice isoform SR-BII [SR-BI/SR-BII (KKB-1), ∼82 kDa] or polyclonal rabbit anti-ε-COP (∼34 kDa) that was used as a loading control. B : immunoblotting analysis of the expression of SR-BII using a rabbit polyclonal, SR-BII-specific anti-carboxy-terminal peptide antibody [SR-BII (anti-C-term)] and anti-ε-COP. C : quantitative analyses of immunoblots was used to determine the relative expression of SR-BI/SR-BII (KKB-1) and SR-BII in the livers of WT and SR-BIΔCT (ΔCT) mice. *Significantly different [ P
    Figure Legend Snippet: Effects of the SR-BIΔCT mutation on the hepatic expression of SR-BI and SR-BII proteins. A and B : immunoblotting analyses of liver lysates (∼30 μg protein) from male WT and SR-BIΔCT (ΔCT) 6- to 10-wk-old mice. Results in female mice were virtually identical. Bands were visualized by chemiluminescence. A : proteins were detected with either a SR-BI-specific anti-carboxy-terminal peptide antibody [SR-BI (anti-C-term)] or a rabbit polyclonal antibody that recognizes the extracellular domains of both SR-BI and its minor splice isoform SR-BII [SR-BI/SR-BII (KKB-1), ∼82 kDa] or polyclonal rabbit anti-ε-COP (∼34 kDa) that was used as a loading control. B : immunoblotting analysis of the expression of SR-BII using a rabbit polyclonal, SR-BII-specific anti-carboxy-terminal peptide antibody [SR-BII (anti-C-term)] and anti-ε-COP. C : quantitative analyses of immunoblots was used to determine the relative expression of SR-BI/SR-BII (KKB-1) and SR-BII in the livers of WT and SR-BIΔCT (ΔCT) mice. *Significantly different [ P

    Techniques Used: Mutagenesis, Expressing, Mouse Assay, Western Blot

    6) Product Images from "The sumoylation pathway is dysregulated in multiple myeloma and is associated with adverse patient outcome"

    Article Title: The sumoylation pathway is dysregulated in multiple myeloma and is associated with adverse patient outcome

    Journal: Blood

    doi: 10.1182/blood-2009-03-211045

    Effect of UBE2I- dominant-negative mutant on growth, proliferation, and adhesion to BMSCs . (A) Myc-tagged fusion proteins were expressed in RPMI 8226 cells. Mo indicates mock transfected; Wt, UBE2I wild-type; and DN , UBE2I-DN (dominant negative). Lysates were probed with a Myc-tag specific mAb (top), Ube2I-specific mAb (middle), or Sumo-1 polyclonal Ab (bottom). (B) UBE2I-DN increased γ-radiation–induced apoptosis. Early apoptosis was detected by annexin V staining. Plots indicate percentage of RPMI 8226 cells in early apoptosis 24 hours after treatment with 10 Gy γ-radiation. Data are representative of 3 independent experiments. (C) UBE2I-wt and UBE2I-DN resulted in opposite effects on MM cell growth. Values represent the mean of triplicate measurements using the MTT assay. (D) Ube2I is necessary for MM adhesion to bone marrow stroma. Adhesion of GFP-labeled UBE2I transfectants to normal BMSCs is shown by fluorescence microscopy after 96 hours of incubation under standard conditions. Data are representative of 3 independent experiments. Images were viewed with a Zeiss Axiovert 200 inverted epifluorescence microscope using a 20× objective (37°C; cells were in PBS; FITC fluorescent filter). Images were acquired with a Zeiss AxioCam HRc 14-bit color CCD camera and were processed with Axio Vision software (Version 3.1). (E) UBE2I-DN decreased BMSCs-induced uptake of 3 H-thymidine. Values represent the mean of triplicate measurements of 3 H-thymidine after 96 hours of coculture.
    Figure Legend Snippet: Effect of UBE2I- dominant-negative mutant on growth, proliferation, and adhesion to BMSCs . (A) Myc-tagged fusion proteins were expressed in RPMI 8226 cells. Mo indicates mock transfected; Wt, UBE2I wild-type; and DN , UBE2I-DN (dominant negative). Lysates were probed with a Myc-tag specific mAb (top), Ube2I-specific mAb (middle), or Sumo-1 polyclonal Ab (bottom). (B) UBE2I-DN increased γ-radiation–induced apoptosis. Early apoptosis was detected by annexin V staining. Plots indicate percentage of RPMI 8226 cells in early apoptosis 24 hours after treatment with 10 Gy γ-radiation. Data are representative of 3 independent experiments. (C) UBE2I-wt and UBE2I-DN resulted in opposite effects on MM cell growth. Values represent the mean of triplicate measurements using the MTT assay. (D) Ube2I is necessary for MM adhesion to bone marrow stroma. Adhesion of GFP-labeled UBE2I transfectants to normal BMSCs is shown by fluorescence microscopy after 96 hours of incubation under standard conditions. Data are representative of 3 independent experiments. Images were viewed with a Zeiss Axiovert 200 inverted epifluorescence microscope using a 20× objective (37°C; cells were in PBS; FITC fluorescent filter). Images were acquired with a Zeiss AxioCam HRc 14-bit color CCD camera and were processed with Axio Vision software (Version 3.1). (E) UBE2I-DN decreased BMSCs-induced uptake of 3 H-thymidine. Values represent the mean of triplicate measurements of 3 H-thymidine after 96 hours of coculture.

    Techniques Used: Dominant Negative Mutation, Transfection, Staining, MTT Assay, Labeling, Fluorescence, Microscopy, Incubation, Inverted Epifluorescence, Software

    7) Product Images from "Amyloid Precursor-like Protein 2 and Sortilin Do Not Regulate the PCSK9 Convertase-mediated Low Density Lipoprotein Receptor Degradation but Interact with Each Other *"

    Article Title: Amyloid Precursor-like Protein 2 and Sortilin Do Not Regulate the PCSK9 Convertase-mediated Low Density Lipoprotein Receptor Degradation but Interact with Each Other *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M115.647180

    Co-expression of sortilin, APLP2, or both with PCSK9 has no major effect on LDLR degradation. Huh7 cells were transfected with a total of 3 μg using 1 μg of each vector encoding for either a control protein 7B2 (−), sortilin (+), APLP2 (+), or PCSK9 (+), as indicated. After 48 h, lysates were analyzed by Western blotting for expression of the LDLR, sortilin-Myc, APLP2-V5, intracellular pro- and mature-PCSK9-V5, and β-actin. Media were analyzed for secreted endogenous and overexpressed PCSK9-V5 using a rabbit polyclonal human PCSK9 antibody. Quantification of LDLR expression was normalized against that of β-actin. These data are representative of at least 3 different experiments showing similar results.
    Figure Legend Snippet: Co-expression of sortilin, APLP2, or both with PCSK9 has no major effect on LDLR degradation. Huh7 cells were transfected with a total of 3 μg using 1 μg of each vector encoding for either a control protein 7B2 (−), sortilin (+), APLP2 (+), or PCSK9 (+), as indicated. After 48 h, lysates were analyzed by Western blotting for expression of the LDLR, sortilin-Myc, APLP2-V5, intracellular pro- and mature-PCSK9-V5, and β-actin. Media were analyzed for secreted endogenous and overexpressed PCSK9-V5 using a rabbit polyclonal human PCSK9 antibody. Quantification of LDLR expression was normalized against that of β-actin. These data are representative of at least 3 different experiments showing similar results.

    Techniques Used: Expressing, Transfection, Plasmid Preparation, Western Blot

    8) Product Images from "Rhox8 Ablation in the Sertoli Cells Using a Tissue-Specific RNAi Approach Results in Impaired Male Fertility in Mice 1"

    Article Title: Rhox8 Ablation in the Sertoli Cells Using a Tissue-Specific RNAi Approach Results in Impaired Male Fertility in Mice 1

    Journal: Biology of Reproduction

    doi: 10.1095/biolreprod.114.124834

    Rhox8 encodes a Sertoli-specific homeobox factor. Serial testis sections from PND 12 mice were incubated with either preimmune serum (1:250 dilution; A ) or RHOX8 polyclonal antiserum (1:2500 dilution; B ) from IMGENEX rabbit #2223B immunized with a RHOX8
    Figure Legend Snippet: Rhox8 encodes a Sertoli-specific homeobox factor. Serial testis sections from PND 12 mice were incubated with either preimmune serum (1:250 dilution; A ) or RHOX8 polyclonal antiserum (1:2500 dilution; B ) from IMGENEX rabbit #2223B immunized with a RHOX8

    Techniques Used: Mouse Assay, Incubation

    9) Product Images from "Regulation of a rat VL30 element in human breast cancer cells in hypoxia and anoxia: role of HIF-1"

    Article Title: Regulation of a rat VL30 element in human breast cancer cells in hypoxia and anoxia: role of HIF-1

    Journal: British Journal of Cancer

    doi: 10.1038/sj.bjc.6600576

    ( a ) Electrophoretic mobility supershift assay showing binding of HIF-1α and HIF-1β in MCF-7 extracts to the trimerised SARE probe following exposure of cells for 16 hours to various O 2 tensions. Bands appear in 1, 0.5 and 0% O 2 . These inducible complexes were supershifted in the presence of a monoclonal antibody to HIF-1α or a polyclonal antibody to HIF-1β (C=binding of constitutive factors). ( b ) Electrophoretic mobility supershift assay showing the inducible HIF-1 band binding to the wt SARE or M1 trimers (the HIF-binding sequence of the latter being 5′-CACGTG-3′) in nuclear extracts of MCF-7 cells exposed for 16 h to various O 2 tensions. Bands appear in 1, 0.5 and 0% O 2 with both probes. These are super-shifted by addition of a monoclonal antibody to HIF-1α. More extensive binding of constitutive factors (‘C’) can be seen with M1 than with the wt SARE probe.
    Figure Legend Snippet: ( a ) Electrophoretic mobility supershift assay showing binding of HIF-1α and HIF-1β in MCF-7 extracts to the trimerised SARE probe following exposure of cells for 16 hours to various O 2 tensions. Bands appear in 1, 0.5 and 0% O 2 . These inducible complexes were supershifted in the presence of a monoclonal antibody to HIF-1α or a polyclonal antibody to HIF-1β (C=binding of constitutive factors). ( b ) Electrophoretic mobility supershift assay showing the inducible HIF-1 band binding to the wt SARE or M1 trimers (the HIF-binding sequence of the latter being 5′-CACGTG-3′) in nuclear extracts of MCF-7 cells exposed for 16 h to various O 2 tensions. Bands appear in 1, 0.5 and 0% O 2 with both probes. These are super-shifted by addition of a monoclonal antibody to HIF-1α. More extensive binding of constitutive factors (‘C’) can be seen with M1 than with the wt SARE probe.

    Techniques Used: Binding Assay, Sequencing

    10) Product Images from "Amyloid Precursor-like Protein 2 and Sortilin Do Not Regulate the PCSK9 Convertase-mediated Low Density Lipoprotein Receptor Degradation but Interact with Each Other *"

    Article Title: Amyloid Precursor-like Protein 2 and Sortilin Do Not Regulate the PCSK9 Convertase-mediated Low Density Lipoprotein Receptor Degradation but Interact with Each Other *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M115.647180

    Co-expression of sortilin, APLP2, or both with PCSK9 has no major effect on LDLR degradation. Huh7 cells were transfected with a total of 3 μg using 1 μg of each vector encoding for either a control protein 7B2 (−), sortilin (+), APLP2 (+), or PCSK9 (+), as indicated. After 48 h, lysates were analyzed by Western blotting for expression of the LDLR, sortilin-Myc, APLP2-V5, intracellular pro- and mature-PCSK9-V5, and β-actin. Media were analyzed for secreted endogenous and overexpressed PCSK9-V5 using a rabbit polyclonal human PCSK9 antibody. Quantification of LDLR expression was normalized against that of β-actin. These data are representative of at least 3 different experiments showing similar results.
    Figure Legend Snippet: Co-expression of sortilin, APLP2, or both with PCSK9 has no major effect on LDLR degradation. Huh7 cells were transfected with a total of 3 μg using 1 μg of each vector encoding for either a control protein 7B2 (−), sortilin (+), APLP2 (+), or PCSK9 (+), as indicated. After 48 h, lysates were analyzed by Western blotting for expression of the LDLR, sortilin-Myc, APLP2-V5, intracellular pro- and mature-PCSK9-V5, and β-actin. Media were analyzed for secreted endogenous and overexpressed PCSK9-V5 using a rabbit polyclonal human PCSK9 antibody. Quantification of LDLR expression was normalized against that of β-actin. These data are representative of at least 3 different experiments showing similar results.

    Techniques Used: Expressing, Transfection, Plasmid Preparation, Western Blot

    11) Product Images from "Cross-talk between IRAK-4 and the NADPH oxidase 1"

    Article Title: Cross-talk between IRAK-4 and the NADPH oxidase 1

    Journal: Biochemical Journal

    doi: 10.1042/BJ20061184

    IRAK-4 enhances the NADPH oxidase activity in granulocytes ( A ) HL-60 cells were transfected by nucleofection (Amaxa) with the IRAK-4 wild-type expression vector (●) or with the control empty vector (○). The cells were differentiated to granulocytes by incubation with DMSO for 48 h, harvested and incubated with LPS (100 ng/ml) for 30 min before stimulation with 1 μM fMLP. An example of a kinetic reaction in the absence of stimulation is also shown (▼). The production of ROS was measured by the detection of the luminol-dependent chemiluminescence after activation with fMLP at 37 °C as described in the Experimental section. ( B ) The oxidative response is significantly increased in granulocytes overexpressing IRAK-4. Chemiluminescence response to fMLP is expressed relative to the condition corresponding to cells transfected with the empty vector and in the absence of LPS treatment. Results are means±S.E.M. for three independent experiments. * P =0.05. ( C ) The cells were recovered and evaluated for the expression of IRAK-4 by Western blot (WB) using a specific anti-IRAK-4 polyclonal antibody (ProSci).
    Figure Legend Snippet: IRAK-4 enhances the NADPH oxidase activity in granulocytes ( A ) HL-60 cells were transfected by nucleofection (Amaxa) with the IRAK-4 wild-type expression vector (●) or with the control empty vector (○). The cells were differentiated to granulocytes by incubation with DMSO for 48 h, harvested and incubated with LPS (100 ng/ml) for 30 min before stimulation with 1 μM fMLP. An example of a kinetic reaction in the absence of stimulation is also shown (▼). The production of ROS was measured by the detection of the luminol-dependent chemiluminescence after activation with fMLP at 37 °C as described in the Experimental section. ( B ) The oxidative response is significantly increased in granulocytes overexpressing IRAK-4. Chemiluminescence response to fMLP is expressed relative to the condition corresponding to cells transfected with the empty vector and in the absence of LPS treatment. Results are means±S.E.M. for three independent experiments. * P =0.05. ( C ) The cells were recovered and evaluated for the expression of IRAK-4 by Western blot (WB) using a specific anti-IRAK-4 polyclonal antibody (ProSci).

    Techniques Used: Activity Assay, Transfection, Expressing, Plasmid Preparation, Incubation, Activation Assay, Western Blot

    12) Product Images from "Human Immunodeficiency Virus Impairs Reverse Cholesterol Transport from MacrophagesHIV-Cholesterol Connection Suggests a New Antiretroviral Strategy"

    Article Title: Human Immunodeficiency Virus Impairs Reverse Cholesterol Transport from MacrophagesHIV-Cholesterol Connection Suggests a New Antiretroviral Strategy

    Journal: PLoS Biology

    doi: 10.1371/journal.pbio.0040365

    Nef Interacts with ABCA1 (A) ABCA1 was immunoprecipitated using anti-FLAG M2 affinity gel from HeLa cells co-transfected with ABCA1-FLAG and an empty vector (mock), WT SF2 Nef (Nef.wt), or myristoylation-defective mutant Nef.G2A. Immunoprecipitates were analyzed by Wester n blotting for Nef (upper panel) or ABCA1 (middle panel) using specific antibodies. Bottom panel shows Nef expression in lysates of cells used for immunoprecipitation. (B) Experiment was performed as in Figure 4 E, except that cells were incubated with monoclonal anti-ABCA1 and polyclonal anti-Nef antibody, followed by FITC-conjugated anti-rabbit and Cy5-conjugated anti-mouse antibodies. Since all transfected cells show re-localization of ABCA1 ( Figure 4 E), a typical single cell is shown here. Images were analyzed using software on the Zeiss LSM 510 microscope. The scale bar represents 20 μm. (C) Fluorescence profile of the image in Figure 5 B was analyzed using the LSM 510 software. The top panel shows the distributions of the ABCA1 and HIV-1 Nef proteins in blue and green, respectively. The same analysis in the lower panel was performed for ABCA1 (blue) and Nef.G2A (green). The co-localization of WT Nef and ABCA1 at the plasma membrane is indicated by overlapping green and blue peaks at either end of the graph in the top panel.
    Figure Legend Snippet: Nef Interacts with ABCA1 (A) ABCA1 was immunoprecipitated using anti-FLAG M2 affinity gel from HeLa cells co-transfected with ABCA1-FLAG and an empty vector (mock), WT SF2 Nef (Nef.wt), or myristoylation-defective mutant Nef.G2A. Immunoprecipitates were analyzed by Wester n blotting for Nef (upper panel) or ABCA1 (middle panel) using specific antibodies. Bottom panel shows Nef expression in lysates of cells used for immunoprecipitation. (B) Experiment was performed as in Figure 4 E, except that cells were incubated with monoclonal anti-ABCA1 and polyclonal anti-Nef antibody, followed by FITC-conjugated anti-rabbit and Cy5-conjugated anti-mouse antibodies. Since all transfected cells show re-localization of ABCA1 ( Figure 4 E), a typical single cell is shown here. Images were analyzed using software on the Zeiss LSM 510 microscope. The scale bar represents 20 μm. (C) Fluorescence profile of the image in Figure 5 B was analyzed using the LSM 510 software. The top panel shows the distributions of the ABCA1 and HIV-1 Nef proteins in blue and green, respectively. The same analysis in the lower panel was performed for ABCA1 (blue) and Nef.G2A (green). The co-localization of WT Nef and ABCA1 at the plasma membrane is indicated by overlapping green and blue peaks at either end of the graph in the top panel.

    Techniques Used: Immunoprecipitation, Transfection, Plasmid Preparation, Mutagenesis, Expressing, Incubation, Software, Microscopy, Fluorescence

    The Effect of Nef on ABCA1 Localization (A–D) On day 5 after infection with VSV-G-pseudotyped Nef-expressing (B and D) or ΔNef (A and C) HIV-1 SF2, cells were co-stained with anti-p24 mouse monoclonal and anti-ABCA1 rabbit polyclonal antibodies, followed by FITC-conjugated anti-mouse (A and B) and Cy5-conjugated anti-rabbit IgG (C and D). Arrows point to cells with re-localized ABCA1. The scale bars represent 20 μm. (E–G) Distribution of ABCA1 revealed by staining with monoclonal anti-ABCA1 antibody and FITC-conjugated anti-mouse IgG in RAW 264.7 cells transfected with empty vector (E), WT Nef derived from SF2 HIV-1 ( Nef.wt, panel [F]), or SF2 Nef carrying a G2A mutation ( Nef.G2A , [G]). Insets in (E and F) show cross-section of the image reconstituted from serial sectioning. Scale bars represent 20 μm. (H) [ 125 -I]apoA-I binding (left panel) and internalization (right panel) in RAW 264.7 macrophages transfected with HIV-1 SF2-derived Nef. An asterisk (*) indicates p
    Figure Legend Snippet: The Effect of Nef on ABCA1 Localization (A–D) On day 5 after infection with VSV-G-pseudotyped Nef-expressing (B and D) or ΔNef (A and C) HIV-1 SF2, cells were co-stained with anti-p24 mouse monoclonal and anti-ABCA1 rabbit polyclonal antibodies, followed by FITC-conjugated anti-mouse (A and B) and Cy5-conjugated anti-rabbit IgG (C and D). Arrows point to cells with re-localized ABCA1. The scale bars represent 20 μm. (E–G) Distribution of ABCA1 revealed by staining with monoclonal anti-ABCA1 antibody and FITC-conjugated anti-mouse IgG in RAW 264.7 cells transfected with empty vector (E), WT Nef derived from SF2 HIV-1 ( Nef.wt, panel [F]), or SF2 Nef carrying a G2A mutation ( Nef.G2A , [G]). Insets in (E and F) show cross-section of the image reconstituted from serial sectioning. Scale bars represent 20 μm. (H) [ 125 -I]apoA-I binding (left panel) and internalization (right panel) in RAW 264.7 macrophages transfected with HIV-1 SF2-derived Nef. An asterisk (*) indicates p

    Techniques Used: Infection, Expressing, Staining, Transfection, Plasmid Preparation, Derivative Assay, Mutagenesis, Binding Assay

    Identification of HIV-1–Positive Macrophages in Atherosclerotic Plaques of HIV-Infected Subjects. Single (A–D) and double (E and F) immunostaining of aortic wall segments. (A) p24 staining. A low-magnification image showing the presence of p24 + cells in an area adjacent to the plaque lipid core. The scale bar represents 100 μm. (B) Detail of (A). p24 + cells show a characteristic morphology of foam cells. The scale bar represents 10 μm. (C) CD68 staining. CD68 + cells were identified in a parallel consecutive section to that shown in (A). The scale bar represents 100 μm. (D) Negative control (staining with an irrelevant primary antibody). The scale bar represents 100 μm. (E) Double immunostaining showing the co-localization of p24 (brown) with CD68 (rose). Immunostaining included a combination of a rabbit polyclonal anti-p24 antibody in the peroxidase–anti-peroxidase system with DAB chromogen yielding a brown reaction product, and a mouse monoclonal antibody to CD68 in the alkaline phosphatase–anti-alkaline phosphatase system with Fast Red chromogen, resulting in a rose precipitate. Counterstaining was with Mayer's hematoxylin. The scale bar represents 50 μm. (F) A detail of (E). The scale bar represents 15 μm.
    Figure Legend Snippet: Identification of HIV-1–Positive Macrophages in Atherosclerotic Plaques of HIV-Infected Subjects. Single (A–D) and double (E and F) immunostaining of aortic wall segments. (A) p24 staining. A low-magnification image showing the presence of p24 + cells in an area adjacent to the plaque lipid core. The scale bar represents 100 μm. (B) Detail of (A). p24 + cells show a characteristic morphology of foam cells. The scale bar represents 10 μm. (C) CD68 staining. CD68 + cells were identified in a parallel consecutive section to that shown in (A). The scale bar represents 100 μm. (D) Negative control (staining with an irrelevant primary antibody). The scale bar represents 100 μm. (E) Double immunostaining showing the co-localization of p24 (brown) with CD68 (rose). Immunostaining included a combination of a rabbit polyclonal anti-p24 antibody in the peroxidase–anti-peroxidase system with DAB chromogen yielding a brown reaction product, and a mouse monoclonal antibody to CD68 in the alkaline phosphatase–anti-alkaline phosphatase system with Fast Red chromogen, resulting in a rose precipitate. Counterstaining was with Mayer's hematoxylin. The scale bar represents 50 μm. (F) A detail of (E). The scale bar represents 15 μm.

    Techniques Used: Infection, Immunostaining, Staining, Negative Control, Double Immunostaining

    13) Product Images from "Human T-cell leukemia virus type 2 post-transcriptional control protein p28 is required for viral infectivity and persistence in vivo"

    Article Title: Human T-cell leukemia virus type 2 post-transcriptional control protein p28 is required for viral infectivity and persistence in vivo

    Journal: Retrovirology

    doi: 10.1186/1742-4690-5-38

    Expression of p19 Gag and Tax protein and p28 mRNA in permanent transfectants . (A) Four 729 stable transfectants (clone 1–4) were isolated for HTLV-2Δp28 as described in Materials and Methods. Our well-established 729pH6neo (729.HTLV-2) cell clone was used as the wtHTLV-2 stable producer cell line. p19 Gag was quantified by ELISA from the four independently isolated 729.HTLV-2Δp28 (Clones 1–4), 729.HTLV-2, and the 729 negative control. Each 729.HTLV-2 producer cell line displayed variable p19 production. (B) Clones indicated by asterisks, which have been shown to produce similar quantities of p19 Gag, were further characterized by Western blot for Tax protein expression using rabbit polyclonal antisera raised against Tax-2. β-actin was used as a loading control. Numbers below each lane are the copy number of p28 transcript per 10 6 copies of GAPDH determined by realtime RT PCR. The results show similar levels of p28 mRNA expression.
    Figure Legend Snippet: Expression of p19 Gag and Tax protein and p28 mRNA in permanent transfectants . (A) Four 729 stable transfectants (clone 1–4) were isolated for HTLV-2Δp28 as described in Materials and Methods. Our well-established 729pH6neo (729.HTLV-2) cell clone was used as the wtHTLV-2 stable producer cell line. p19 Gag was quantified by ELISA from the four independently isolated 729.HTLV-2Δp28 (Clones 1–4), 729.HTLV-2, and the 729 negative control. Each 729.HTLV-2 producer cell line displayed variable p19 production. (B) Clones indicated by asterisks, which have been shown to produce similar quantities of p19 Gag, were further characterized by Western blot for Tax protein expression using rabbit polyclonal antisera raised against Tax-2. β-actin was used as a loading control. Numbers below each lane are the copy number of p28 transcript per 10 6 copies of GAPDH determined by realtime RT PCR. The results show similar levels of p28 mRNA expression.

    Techniques Used: Expressing, Isolation, Enzyme-linked Immunosorbent Assay, Negative Control, Western Blot, Reverse Transcription Polymerase Chain Reaction

    14) Product Images from "The sumoylation pathway is dysregulated in multiple myeloma and is associated with adverse patient outcome"

    Article Title: The sumoylation pathway is dysregulated in multiple myeloma and is associated with adverse patient outcome

    Journal: Blood

    doi: 10.1182/blood-2009-03-211045

    Effect of UBE2I- dominant-negative mutant on growth, proliferation, and adhesion to BMSCs . (A) Myc-tagged fusion proteins were expressed in RPMI 8226 cells. Mo indicates mock transfected; Wt, UBE2I wild-type; and DN , UBE2I-DN (dominant negative). Lysates were probed with a Myc-tag specific mAb (top), Ube2I-specific mAb (middle), or Sumo-1 polyclonal Ab (bottom). (B) UBE2I-DN increased γ-radiation–induced apoptosis. Early apoptosis was detected by annexin V staining. Plots indicate percentage of RPMI 8226 cells in early apoptosis 24 hours after treatment with 10 Gy γ-radiation. Data are representative of 3 independent experiments. (C) UBE2I-wt and UBE2I-DN resulted in opposite effects on MM cell growth. Values represent the mean of triplicate measurements using the MTT assay. (D) Ube2I is necessary for MM adhesion to bone marrow stroma. Adhesion of GFP-labeled UBE2I transfectants to normal BMSCs is shown by fluorescence microscopy after 96 hours of incubation under standard conditions. Data are representative of 3 independent experiments. Images were viewed with a Zeiss Axiovert 200 inverted epifluorescence microscope using a 20× objective (37°C; cells were in PBS; FITC fluorescent filter). Images were acquired with a Zeiss AxioCam HRc 14-bit color CCD camera and were processed with Axio Vision software (Version 3.1). (E) UBE2I-DN decreased BMSCs-induced uptake of 3 H-thymidine. Values represent the mean of triplicate measurements of 3 H-thymidine after 96 hours of coculture.
    Figure Legend Snippet: Effect of UBE2I- dominant-negative mutant on growth, proliferation, and adhesion to BMSCs . (A) Myc-tagged fusion proteins were expressed in RPMI 8226 cells. Mo indicates mock transfected; Wt, UBE2I wild-type; and DN , UBE2I-DN (dominant negative). Lysates were probed with a Myc-tag specific mAb (top), Ube2I-specific mAb (middle), or Sumo-1 polyclonal Ab (bottom). (B) UBE2I-DN increased γ-radiation–induced apoptosis. Early apoptosis was detected by annexin V staining. Plots indicate percentage of RPMI 8226 cells in early apoptosis 24 hours after treatment with 10 Gy γ-radiation. Data are representative of 3 independent experiments. (C) UBE2I-wt and UBE2I-DN resulted in opposite effects on MM cell growth. Values represent the mean of triplicate measurements using the MTT assay. (D) Ube2I is necessary for MM adhesion to bone marrow stroma. Adhesion of GFP-labeled UBE2I transfectants to normal BMSCs is shown by fluorescence microscopy after 96 hours of incubation under standard conditions. Data are representative of 3 independent experiments. Images were viewed with a Zeiss Axiovert 200 inverted epifluorescence microscope using a 20× objective (37°C; cells were in PBS; FITC fluorescent filter). Images were acquired with a Zeiss AxioCam HRc 14-bit color CCD camera and were processed with Axio Vision software (Version 3.1). (E) UBE2I-DN decreased BMSCs-induced uptake of 3 H-thymidine. Values represent the mean of triplicate measurements of 3 H-thymidine after 96 hours of coculture.

    Techniques Used: Dominant Negative Mutation, Transfection, Staining, MTT Assay, Labeling, Fluorescence, Microscopy, Incubation, Inverted Epifluorescence, Software

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    Article Snippet: .. For immunoprecipitation of Myc- and FLAG-tagged proteins or of human WDR79, monoclonal 9E10 and M2 antibodies (Sigma) or rabbit polyclonal antibody (Novus Biologicals) were used, respectively. ..

    Mouse Assay:

    Article Title: Carboxy-terminal deletion of the HDL receptor reduces receptor levels in liver and steroidogenic tissues, induces hypercholesterolemia, and causes fatal heart disease
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    Concentration Assay:

    Article Title: The sumoylation pathway is dysregulated in multiple myeloma and is associated with adverse patient outcome
    Article Snippet: .. Rabbit polyclonal antibody to human NSE2 (BC100-2506) was obtained from Novus Biologicals and was used at a final concentration of 1:500 and rabbit polyclonal antibody to PIAS1 was from Abcam. .. Mouse monoclonal antibody to GAPDH (Ab9484) was from Abcam.

    Incubation:

    Article Title: Telomerase inhibition by siRNA causes senescence and apoptosis in Barrett's adenocarcinoma cells: mechanism and therapeutic potential
    Article Snippet: .. Fixed cells were rinsed, rehydrated in PBS, and incubated for 2 h at RT with rabbit polyclonal antibody to telomerase (Novus Biologicals, Inc., Littleton, CO). .. Antigen-antibody complex was detected by incubation with a fluorescein isothiocyanate (FITC)-labeled secondary antibody.

    Article Title: Carboxy-terminal deletion of the HDL receptor reduces receptor levels in liver and steroidogenic tissues, induces hypercholesterolemia, and causes fatal heart disease
    Article Snippet: .. Protein samples (∼30 μg, as determined with a Bio-Rad DC protein assay) from total tissue lysates of both male and female mice (7 livers, 7 adrenal glands, and 3 ovaries per group) were fractionated by 5–20% gradient SDS-PAGE, transferred to nitrocellulose membranes, and incubated with a rabbit polyclonal anti-SR-BI antibody (mSR-BI495-112 ) raised against a carboxy-terminal peptide of the protein, the anti-SR-BI KKB-1 antibody , a rabbit polyclonal antibody that recognizes both SR-BI and SR-BII , or a specific rabbit anti-SR-BII ( ) antibody (Novus Biologicals) (all used at 1:500 dilution), followed by an anti-rabbit IgG conjugated to horseradish peroxidase (Invitrogen, 1:10,000), and visualized by ECL chemiluminescence (GE Healthcare). ..

    Article Title: TLR9-induced interferon ? is associated with protection from gammaherpesvirus-induced exacerbation of lung fibrosis
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    Recombinant:

    Article Title: Expression Pattern of the Aspartyl-tRNA Synthetase DARS in the Human Brain
    Article Snippet: .. The second antibody was a polyclonal rabbit antibody from Novus Biologicals (rbαDARS), which was developed against recombinant protein corresponding to amino acids 1–135 of human DARS. .. HEK-293 cells were either transfected with a control plasmid or with a DARS plasmid containing an N-terminal FLAG-HA-tag, lysed after 3 days, and analyzed using Western-blotting.

    SDS Page:

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    DC Protein Assay:

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    Novus Biologicals polyclonal rabbit ab
    Immunohistochemical and subcellular localization of transgenic survivin expression. ( a ) Five-micrometer tissue sections were cut from fresh-frozen skin of K14-survivin transgenic mice (K14-survivin) or control nontransgenic (non-TG) littermates, fixed in ice-cold acetone, and analyzed by immunohistochemistry with Ab against survivin or a control Ab to EGFP. Binding of the primary Ab’s was visualized with a goat anti-rabbit <t>polyclonal</t> Ab using Vectastain Elite ABC and AEC peroxidase substrate kits. ( b ) Subcellular localization of transgenic survivin. Keratinocytes isolated from K14-survivin transgenic mice (K14-survivin) or nontransgenic littermates (non-TG) were adhered to glass coverslips, fixed in methanol-acetone, and incubated with an Ab to survivin, followed by Texas red–conjugated goat anti-rabbit Ab. Nuclei were stained with Hoechst 33342. Image merging analysis is shown.
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    Novus Biologicals rabbit polyclonal anti hif 1α
    Expression of specific OATP genes and <t>HIF-1α-target</t> genes in tumor tissues and cells qPCR analysis of VEGF-A and specific OATP gene expression in canine cancer tissues (A) human cancer tissues (B) and canine cancer cell lines (C) . Cancer-adjacent normal tissues derived from the same canine (A) or human (B) patients were used as normal controls for respective cancer tissues (N=5 patients for each type of cancer). Primary cultures of normal canine prostate and mammary gland epithelial cells were used as normal controls for ACE1 and CHMp-5b cells, respectively (C). Either human GAPDH or canine Gapdh was used as an internal control for normalization. Data represent the mean ± SD from three independent experiments. * P
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    Novus Biologicals ccl2
    The adipocyte mTORC2 /Rictor mediate the effect of FGF1 on regulating <t>Ccl2</t> expression. A and B, Western blot analyses of the mTORC2/Rictor and inflammation signalling in epididymal adipose tissue of db/db mice after chronic treatment with FGF1, and (B) the densitometric quantification by ImageJ software. Data are presented as mean ± SEM. * P
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    Novus Biologicals nox4 antibody
    Diabetic atrophied muscles exhibited a state of heightened oxidative stress (HSOS). (a and b) Superoxide generation was measured in frozen muscle sections of control and diabetic using dihydroethidium-based confocal microscopic staining technique. (c and d) NADPH oxidase in a membrane fraction was assessed according to procedure involving the substrate NADPH and lucigenin chemiluminescence or the Amplex Red/horseradish peroxidase fluorescence-based assays. (e and f) Muscle NADPH oxidase-related isoforms including NOX2 and <t>NOX4</t> were determined at the mRNA (e) and protein levels (f) using RT-PCR and Western blotting-based techniques. (g) Mitochondrial H 2 O 2 generation at the steady state level and in the presence of added glutamate/malate substrates was measured using the Amplex Red/horseradish peroxidase fluorescence-based assay (g). Activities of complexes I (h) and III (i) of the electron transport chain were measured using spectrophotometric-based assay. Abbreviation: C: control; D: diabetic. Values are means ± SEM for at least 6 animals/group. ∗ Significantly different from corresponding control values at P ≤ 0.05.
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    Immunohistochemical and subcellular localization of transgenic survivin expression. ( a ) Five-micrometer tissue sections were cut from fresh-frozen skin of K14-survivin transgenic mice (K14-survivin) or control nontransgenic (non-TG) littermates, fixed in ice-cold acetone, and analyzed by immunohistochemistry with Ab against survivin or a control Ab to EGFP. Binding of the primary Ab’s was visualized with a goat anti-rabbit polyclonal Ab using Vectastain Elite ABC and AEC peroxidase substrate kits. ( b ) Subcellular localization of transgenic survivin. Keratinocytes isolated from K14-survivin transgenic mice (K14-survivin) or nontransgenic littermates (non-TG) were adhered to glass coverslips, fixed in methanol-acetone, and incubated with an Ab to survivin, followed by Texas red–conjugated goat anti-rabbit Ab. Nuclei were stained with Hoechst 33342. Image merging analysis is shown.

    Journal: Journal of Clinical Investigation

    Article Title: Transgenic expression of survivin in keratinocytes counteracts UVB-induced apoptosis and cooperates with loss of p53

    doi:

    Figure Lengend Snippet: Immunohistochemical and subcellular localization of transgenic survivin expression. ( a ) Five-micrometer tissue sections were cut from fresh-frozen skin of K14-survivin transgenic mice (K14-survivin) or control nontransgenic (non-TG) littermates, fixed in ice-cold acetone, and analyzed by immunohistochemistry with Ab against survivin or a control Ab to EGFP. Binding of the primary Ab’s was visualized with a goat anti-rabbit polyclonal Ab using Vectastain Elite ABC and AEC peroxidase substrate kits. ( b ) Subcellular localization of transgenic survivin. Keratinocytes isolated from K14-survivin transgenic mice (K14-survivin) or nontransgenic littermates (non-TG) were adhered to glass coverslips, fixed in methanol-acetone, and incubated with an Ab to survivin, followed by Texas red–conjugated goat anti-rabbit Ab. Nuclei were stained with Hoechst 33342. Image merging analysis is shown.

    Article Snippet: Analysis for survivin expression by fluorescence microscopy was performed using the polyclonal rabbit Ab to full-length survivin (NOVUS Biologicals Inc.).

    Techniques: Immunohistochemistry, Transgenic Assay, Expressing, Mouse Assay, Binding Assay, Isolation, Incubation, Staining

    Expression of specific OATP genes and HIF-1α-target genes in tumor tissues and cells qPCR analysis of VEGF-A and specific OATP gene expression in canine cancer tissues (A) human cancer tissues (B) and canine cancer cell lines (C) . Cancer-adjacent normal tissues derived from the same canine (A) or human (B) patients were used as normal controls for respective cancer tissues (N=5 patients for each type of cancer). Primary cultures of normal canine prostate and mammary gland epithelial cells were used as normal controls for ACE1 and CHMp-5b cells, respectively (C). Either human GAPDH or canine Gapdh was used as an internal control for normalization. Data represent the mean ± SD from three independent experiments. * P

    Journal: Oncotarget

    Article Title: Heptamethine carbocyanine dye-mediated near-infrared imaging of canine and human cancers through the HIF-1α/OATPs signaling axis

    doi:

    Figure Lengend Snippet: Expression of specific OATP genes and HIF-1α-target genes in tumor tissues and cells qPCR analysis of VEGF-A and specific OATP gene expression in canine cancer tissues (A) human cancer tissues (B) and canine cancer cell lines (C) . Cancer-adjacent normal tissues derived from the same canine (A) or human (B) patients were used as normal controls for respective cancer tissues (N=5 patients for each type of cancer). Primary cultures of normal canine prostate and mammary gland epithelial cells were used as normal controls for ACE1 and CHMp-5b cells, respectively (C). Either human GAPDH or canine Gapdh was used as an internal control for normalization. Data represent the mean ± SD from three independent experiments. * P

    Article Snippet: Rabbit polyclonal anti-HIF-1α and anti-OATP2B1 antibodies both predicted to react with canine species were purchased from Novus Biologicals (Littleton, CO).

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Derivative Assay

    Specific uptake of MHI-148 dye by canine cancer but not normal cells (A) Representative NIR fluorescence microscopic images show the significant uptake of MHI-148 dye by canine 5b, 13a and ACE1 and human PC-3 cancer cells but not by normal canine MDCK and human HEK293 cells (magnification, 200×). (B) Continued documentation revealed preferential uptake and retention of MHI-148 dye in canine tumor xenografts. Top panels: nude mice bearing subcutaneous xenograft tumors were subjected to NIR fluorescence optical imaging consecutively on days 8, 15, and 22 after implantation. Bottom panels: a histogram summarizes signal intensity detected from xenografts (N=3) at each measurement time point. (C) Mice bearing dually luciferase- and GFP-labeled canine cancer cells were subjected to whole-body bioluminescence (Luciferase), NIR (MHI-148) and GFP fluorescence imaging. Signal intensity ratio of canine 13a to ACE1 tumor xenografts with different labels is shown. (D) Uptake of the NIR dye by human clinical prostate tumor xenografts implanted under the subrenal capsule of nude mice. (E) Examination of dissected xenografts for HIF-1α and OATP2B1 expression following IHC staining (magnification, 200×).

    Journal: Oncotarget

    Article Title: Heptamethine carbocyanine dye-mediated near-infrared imaging of canine and human cancers through the HIF-1α/OATPs signaling axis

    doi:

    Figure Lengend Snippet: Specific uptake of MHI-148 dye by canine cancer but not normal cells (A) Representative NIR fluorescence microscopic images show the significant uptake of MHI-148 dye by canine 5b, 13a and ACE1 and human PC-3 cancer cells but not by normal canine MDCK and human HEK293 cells (magnification, 200×). (B) Continued documentation revealed preferential uptake and retention of MHI-148 dye in canine tumor xenografts. Top panels: nude mice bearing subcutaneous xenograft tumors were subjected to NIR fluorescence optical imaging consecutively on days 8, 15, and 22 after implantation. Bottom panels: a histogram summarizes signal intensity detected from xenografts (N=3) at each measurement time point. (C) Mice bearing dually luciferase- and GFP-labeled canine cancer cells were subjected to whole-body bioluminescence (Luciferase), NIR (MHI-148) and GFP fluorescence imaging. Signal intensity ratio of canine 13a to ACE1 tumor xenografts with different labels is shown. (D) Uptake of the NIR dye by human clinical prostate tumor xenografts implanted under the subrenal capsule of nude mice. (E) Examination of dissected xenografts for HIF-1α and OATP2B1 expression following IHC staining (magnification, 200×).

    Article Snippet: Rabbit polyclonal anti-HIF-1α and anti-OATP2B1 antibodies both predicted to react with canine species were purchased from Novus Biologicals (Littleton, CO).

    Techniques: Fluorescence, Mouse Assay, Optical Imaging, Luciferase, Labeling, Imaging, Expressing, Immunohistochemistry, Staining

    Aberrant expression of HIF-1α and selected OATP genes in cancers Double QDL analysis was used to quantify HIF-1α and OATP2B1 protein expression in paired cancer and normal (derived from cancer-adjacent normal tissue from the same patient) tissue specimens from either canine (A) or human (B) cancer patients (N=5 patients for each type of cancer). Average signal intensity counts from 1,000 cells were quantified using inForm software. * P

    Journal: Oncotarget

    Article Title: Heptamethine carbocyanine dye-mediated near-infrared imaging of canine and human cancers through the HIF-1α/OATPs signaling axis

    doi:

    Figure Lengend Snippet: Aberrant expression of HIF-1α and selected OATP genes in cancers Double QDL analysis was used to quantify HIF-1α and OATP2B1 protein expression in paired cancer and normal (derived from cancer-adjacent normal tissue from the same patient) tissue specimens from either canine (A) or human (B) cancer patients (N=5 patients for each type of cancer). Average signal intensity counts from 1,000 cells were quantified using inForm software. * P

    Article Snippet: Rabbit polyclonal anti-HIF-1α and anti-OATP2B1 antibodies both predicted to react with canine species were purchased from Novus Biologicals (Littleton, CO).

    Techniques: Expressing, Derivative Assay, Software

    Uptake of the MHI-148 dye through the activation of HIF-1α/OATPs signaling axis (A) Uptake of NIR dye by canine cancer cells was regulated by HIF-1α. Canine cancer cells pre-treated with HIF-1α stabilizer DMOG or cobalt chloride showed increased uptake of NIR dye. Canine cancer cells pre-treated with OATP inhibitor RIF or BSP showed reduced NIR dye uptake. All the images are shown at 400× magnification. (B) ACE1 and 5b cells were subjected to qPCR analysis of VEGF-A , Glut1 , and selected OATPs following treatment with HIF-1α stabilizer DMOG or cobalt chloride. Data represent the mean ± SD from three independent experiments. * P

    Journal: Oncotarget

    Article Title: Heptamethine carbocyanine dye-mediated near-infrared imaging of canine and human cancers through the HIF-1α/OATPs signaling axis

    doi:

    Figure Lengend Snippet: Uptake of the MHI-148 dye through the activation of HIF-1α/OATPs signaling axis (A) Uptake of NIR dye by canine cancer cells was regulated by HIF-1α. Canine cancer cells pre-treated with HIF-1α stabilizer DMOG or cobalt chloride showed increased uptake of NIR dye. Canine cancer cells pre-treated with OATP inhibitor RIF or BSP showed reduced NIR dye uptake. All the images are shown at 400× magnification. (B) ACE1 and 5b cells were subjected to qPCR analysis of VEGF-A , Glut1 , and selected OATPs following treatment with HIF-1α stabilizer DMOG or cobalt chloride. Data represent the mean ± SD from three independent experiments. * P

    Article Snippet: Rabbit polyclonal anti-HIF-1α and anti-OATP2B1 antibodies both predicted to react with canine species were purchased from Novus Biologicals (Littleton, CO).

    Techniques: Activation Assay, Real-time Polymerase Chain Reaction

    The adipocyte mTORC2 /Rictor mediate the effect of FGF1 on regulating Ccl2 expression. A and B, Western blot analyses of the mTORC2/Rictor and inflammation signalling in epididymal adipose tissue of db/db mice after chronic treatment with FGF1, and (B) the densitometric quantification by ImageJ software. Data are presented as mean ± SEM. * P

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: Fibroblast growth factor 1 ameliorates adipose tissue inflammation and systemic insulin resistance via enhancing adipocyte mTORC2/Rictor signal. Fibroblast growth factor 1 ameliorates adipose tissue inflammation and systemic insulin resistance via enhancing adipocyte mTORC2/Rictor signal

    doi: 10.1111/jcmm.15872

    Figure Lengend Snippet: The adipocyte mTORC2 /Rictor mediate the effect of FGF1 on regulating Ccl2 expression. A and B, Western blot analyses of the mTORC2/Rictor and inflammation signalling in epididymal adipose tissue of db/db mice after chronic treatment with FGF1, and (B) the densitometric quantification by ImageJ software. Data are presented as mean ± SEM. * P

    Article Snippet: Protein blots were probed with antibodies against CD68 (Abcam; Cat#: ab31630), CCL2 (Novus biological; Cat#: NBP1‐07035), Phospho‐IKKα/β (Cat#: ab59195), IKKα/β (Santa, Cat#: sc‐7607), Phospho‐IKBα (Santa, Cat#: sc‐8404) and IKBα (Santa, Cat#: sc‐371).

    Techniques: Expressing, Western Blot, Mouse Assay, Software

    FGF1 inhibits macrophages migration by negatively controlling CCL2 transcription and expression. A, A schematic diagram showing the co‐culture of adipocytes and macrophages in vitro. B and C, The representative image of the effect of FGF1 (100 ng/mL) and INCB3344 (specific inhibitor of CCR2, 100 nmol/L) on 3T3‐L1 CM‐induced chemotaxis of macrophages (B) and the quantitative analysis (C). The image was representative of similar results from three independent experiments. Scale bar represents 1000 μm for 100×. D, The relative mRNA level of chemokines in eWAT of db/db mice after chronic administration of FGF1. E and F, Representative confocal merged images of CCL2 from epididymal adipose tissue of db/db mice, stained with anti‐CCL2 (red) and DAPI (blue), and the data of quantitative analysis of staining of db/db mice was conducted by ImageJ software. G, Western blot analysis of CCL2 in epididymal adipose tissue of db/db mice after treatment with FGF1. H, The plasma CCL2 level of db/db mice after treatment with FGF1. * P

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: Fibroblast growth factor 1 ameliorates adipose tissue inflammation and systemic insulin resistance via enhancing adipocyte mTORC2/Rictor signal. Fibroblast growth factor 1 ameliorates adipose tissue inflammation and systemic insulin resistance via enhancing adipocyte mTORC2/Rictor signal

    doi: 10.1111/jcmm.15872

    Figure Lengend Snippet: FGF1 inhibits macrophages migration by negatively controlling CCL2 transcription and expression. A, A schematic diagram showing the co‐culture of adipocytes and macrophages in vitro. B and C, The representative image of the effect of FGF1 (100 ng/mL) and INCB3344 (specific inhibitor of CCR2, 100 nmol/L) on 3T3‐L1 CM‐induced chemotaxis of macrophages (B) and the quantitative analysis (C). The image was representative of similar results from three independent experiments. Scale bar represents 1000 μm for 100×. D, The relative mRNA level of chemokines in eWAT of db/db mice after chronic administration of FGF1. E and F, Representative confocal merged images of CCL2 from epididymal adipose tissue of db/db mice, stained with anti‐CCL2 (red) and DAPI (blue), and the data of quantitative analysis of staining of db/db mice was conducted by ImageJ software. G, Western blot analysis of CCL2 in epididymal adipose tissue of db/db mice after treatment with FGF1. H, The plasma CCL2 level of db/db mice after treatment with FGF1. * P

    Article Snippet: Protein blots were probed with antibodies against CD68 (Abcam; Cat#: ab31630), CCL2 (Novus biological; Cat#: NBP1‐07035), Phospho‐IKKα/β (Cat#: ab59195), IKKα/β (Santa, Cat#: sc‐7607), Phospho‐IKBα (Santa, Cat#: sc‐8404) and IKBα (Santa, Cat#: sc‐371).

    Techniques: Migration, Expressing, Co-Culture Assay, In Vitro, Chemotaxis Assay, Mouse Assay, Staining, Software, Western Blot

    Diabetic atrophied muscles exhibited a state of heightened oxidative stress (HSOS). (a and b) Superoxide generation was measured in frozen muscle sections of control and diabetic using dihydroethidium-based confocal microscopic staining technique. (c and d) NADPH oxidase in a membrane fraction was assessed according to procedure involving the substrate NADPH and lucigenin chemiluminescence or the Amplex Red/horseradish peroxidase fluorescence-based assays. (e and f) Muscle NADPH oxidase-related isoforms including NOX2 and NOX4 were determined at the mRNA (e) and protein levels (f) using RT-PCR and Western blotting-based techniques. (g) Mitochondrial H 2 O 2 generation at the steady state level and in the presence of added glutamate/malate substrates was measured using the Amplex Red/horseradish peroxidase fluorescence-based assay (g). Activities of complexes I (h) and III (i) of the electron transport chain were measured using spectrophotometric-based assay. Abbreviation: C: control; D: diabetic. Values are means ± SEM for at least 6 animals/group. ∗ Significantly different from corresponding control values at P ≤ 0.05.

    Journal: Oxidative Medicine and Cellular Longevity

    Article Title: Hydrogen Sulfide Donor NaHS Improves Metabolism and Reduces Muscle Atrophy in Type 2 Diabetes: Implication for Understanding Sarcopenic Pathophysiology

    doi: 10.1155/2018/6825452

    Figure Lengend Snippet: Diabetic atrophied muscles exhibited a state of heightened oxidative stress (HSOS). (a and b) Superoxide generation was measured in frozen muscle sections of control and diabetic using dihydroethidium-based confocal microscopic staining technique. (c and d) NADPH oxidase in a membrane fraction was assessed according to procedure involving the substrate NADPH and lucigenin chemiluminescence or the Amplex Red/horseradish peroxidase fluorescence-based assays. (e and f) Muscle NADPH oxidase-related isoforms including NOX2 and NOX4 were determined at the mRNA (e) and protein levels (f) using RT-PCR and Western blotting-based techniques. (g) Mitochondrial H 2 O 2 generation at the steady state level and in the presence of added glutamate/malate substrates was measured using the Amplex Red/horseradish peroxidase fluorescence-based assay (g). Activities of complexes I (h) and III (i) of the electron transport chain were measured using spectrophotometric-based assay. Abbreviation: C: control; D: diabetic. Values are means ± SEM for at least 6 animals/group. ∗ Significantly different from corresponding control values at P ≤ 0.05.

    Article Snippet: Antibodies Primary antibodies used in the current study included the following: rabbit anti-PTEN, 9552; rabbit ant-phospho S6 (Ser235/236), 4858; S6, 2217, rabbit anti-phospho-4E-BP1 (Thr37/46), 2855; rabbit anti-phospho-Akt (Ser473), 4060; rabbit anti-phospho-Akt (Thr308), 4056; rabbit anti-Akt, 9272; rabbit anti-FoxO1, 2880; rabbit anti-phospho-FoxO1 (Ser256), 8419; rabbit anti-Smad2, 5339; rabbit anti-phospho-Smad2 (Ser465/467), 18338 (Cell Signaling Technology, Bever, USA); rabbit anti-NOX2, NB2-41291 and -NOX4, NB110-58849 (Novus Biologicals, USA); rabbit ant-CSE (MyBioSource, USA MBS 769567); rabbit anti-PAX7, ab92317; and the housekeeping genes rabbit GAPDH, ab181602, and histone H3, ab201456 (Abcam, USA).

    Techniques: Staining, Fluorescence, Reverse Transcription Polymerase Chain Reaction, Western Blot, Spectrophotometric Assay