vectastain abc reagent  (Vector Laboratories)


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    Name:
    VECTASTAIN ABC AP Staining KIT Alkaline Phosphatase Standard
    Description:
    VECTASTAIN ABC AP Staining Kits remains widely popular with more than 50 000 citations to its credit Based on the versatile avidin biotin complex interaction the system is modular and along with our selection of secondary antibodies can accommodate a wide array of primary antibody and tissue species Our ABC kits are economical and continue to be a staple product in any immunohistochemistry IHC and immunocytochemistry ICC laboratory The sensitivity of the VECTASTAIN ABC AP system is comparable to that of the peroxidase VECTASTAIN Elite ABC system The VECTASTAIN ABC AP Kits may be preferred for tissues that have high endogenous peroxidase activity The system also offers additional substrate color choices VECTASTAIN ABC AP Kit Components 2 ml Reagent A2 ml Reagent BReagent A and B when mixed form the ABC AP complex Kit is sufficient to stain approximately 1000 2000 sections or fifty 100 cm2 blots
    Catalog Number:
    AK-5000
    Price:
    None
    Category:
    Protein chromogenic detection reagents or kits or substrates
    Reactivity:
    No antibody included
    Size:
    1 Kit
    Buy from Supplier


    Structured Review

    Vector Laboratories vectastain abc reagent
    VECTASTAIN ABC AP Staining KIT Alkaline Phosphatase Standard
    VECTASTAIN ABC AP Staining Kits remains widely popular with more than 50 000 citations to its credit Based on the versatile avidin biotin complex interaction the system is modular and along with our selection of secondary antibodies can accommodate a wide array of primary antibody and tissue species Our ABC kits are economical and continue to be a staple product in any immunohistochemistry IHC and immunocytochemistry ICC laboratory The sensitivity of the VECTASTAIN ABC AP system is comparable to that of the peroxidase VECTASTAIN Elite ABC system The VECTASTAIN ABC AP Kits may be preferred for tissues that have high endogenous peroxidase activity The system also offers additional substrate color choices VECTASTAIN ABC AP Kit Components 2 ml Reagent A2 ml Reagent BReagent A and B when mixed form the ABC AP complex Kit is sufficient to stain approximately 1000 2000 sections or fifty 100 cm2 blots
    https://www.bioz.com/result/vectastain abc reagent/product/Vector Laboratories
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    vectastain abc reagent - by Bioz Stars, 2021-06
    99/100 stars

    Images

    1) Product Images from "Upregulation of GRAIL Is Associated with Impaired CD4 T-Cell Proliferation in Sepsis"

    Article Title: Upregulation of GRAIL Is Associated with Impaired CD4 T-Cell Proliferation in Sepsis

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    doi: 10.4049/jimmunol.1302160

    Upregulation of GRAIL in CD4 T-cells following sepsis (A) After 24 h of CLP, spleen tissues were harvested from shams and septic mice and fixed in 10% formalin and embedded in paraffin. Tissue blocks were sectioned at a thickness of 5 μm and incubated with rabbit anti-mouse GRAIL Ab, followed by incubation with biotinylated anti-rabbit IgG. Staining was developed by Vectastain ABC reagent and a diaminobenzidine kit. Representative immunostaing images at original magnification of ×100 and ×200 ( inset ) are shown. (B) Cells were isolated from spleens of sham and CLP animals; a total of 2 ×10 6 cells were used to extract mRNA and subsequent realtime PCR analysis using GRAIL primer. The expression of β-actin serves as an internal control. The data is expressed as fold induction in comparison to the shams. (C) Splenocytes (1 × 10 6 cells) from the shams and septic mice were stained for surface CD4 antigen and intracellular GRAIL using PerCP/Cy5.5-CD4 Ab, and rabbit anti-GRAIL primary Ab, respectively. A FITC-labeled anti-rabbit secondary Ab was used subsequently and then events were acquired in flow cytometer. (D) Bar diagram representing the mean fluoresce intensities of sham and septic samples are shown. Data are expressed as means ± SE (n=5 mice/group). * P
    Figure Legend Snippet: Upregulation of GRAIL in CD4 T-cells following sepsis (A) After 24 h of CLP, spleen tissues were harvested from shams and septic mice and fixed in 10% formalin and embedded in paraffin. Tissue blocks were sectioned at a thickness of 5 μm and incubated with rabbit anti-mouse GRAIL Ab, followed by incubation with biotinylated anti-rabbit IgG. Staining was developed by Vectastain ABC reagent and a diaminobenzidine kit. Representative immunostaing images at original magnification of ×100 and ×200 ( inset ) are shown. (B) Cells were isolated from spleens of sham and CLP animals; a total of 2 ×10 6 cells were used to extract mRNA and subsequent realtime PCR analysis using GRAIL primer. The expression of β-actin serves as an internal control. The data is expressed as fold induction in comparison to the shams. (C) Splenocytes (1 × 10 6 cells) from the shams and septic mice were stained for surface CD4 antigen and intracellular GRAIL using PerCP/Cy5.5-CD4 Ab, and rabbit anti-GRAIL primary Ab, respectively. A FITC-labeled anti-rabbit secondary Ab was used subsequently and then events were acquired in flow cytometer. (D) Bar diagram representing the mean fluoresce intensities of sham and septic samples are shown. Data are expressed as means ± SE (n=5 mice/group). * P

    Techniques Used: Mouse Assay, Incubation, Staining, Isolation, Polymerase Chain Reaction, Expressing, Labeling, Flow Cytometry, Cytometry

    Related Articles

    Staining:

    Article Title: Immunohistochemical analysis of macroautophagy
    Article Snippet: In case the primary antibody was biotinylated, blocking with normal serum and incubation with biotinylated secondary antibodies were omitted. .. In some experiments, frozen sections were stained using a Vectastain ABC-AP kit (Vector Laboratories, AK-5000) containing biotinylated alkaline phosphatase instead of biotinylated horseradish peroxidase. .. The procedure is similar with the staining steps for frozen sections described above, except quenching of endogenous peroxidase is no longer required.

    Article Title: Induction of cyclooxygenase-2 in monocyte/macrophage by mucins secreted from colon cancer cells
    Article Snippet: .. Immunohistochemical staining was performed with the Vectastain avidin–biotin peroxidase complex kit (Vector Laboratories) as described ( ). .. Double immunostaining was carried out according to Kawahito et al. ( ).

    Article Title: Diet-induced obesity links to ER positive breast cancer progression via LPA/PKD-1-CD36 signaling-mediated microvascular remodeling
    Article Snippet: E0771 cells (1×106 cells/mouse) were implanted into mice close to the fourth mammary pad [ ] and tumor volume was measured using calipers in two dimensions and calculated using the formula: (width2 × length)/2 [ ]. .. Immunofluorescence and immunohistochemical assays Tissue immunohistochemical and immunofluorescence staining was performed using antibodies and methods as described in our previous studies [ , , ] or by using the Vectastain® ABC kit (Vector Laboratories). .. Mouse angiogenesis profiling A Proteome Profiler™ antibody array (R & D system® ) was performed to determine relative protein levels of angiogenic factors in the plasma according to the manufacturer's instruction.

    Article Title: Dynamic Populations of Dendritic Cell-Specific ICAM-3 Grabbing Nonintegrin-Positive Immature Dendritic Cells and Liver/Lymph Node-Specific ICAM-3 Grabbing Nonintegrin-Positive Endothelial Cells in the Outer Zones of the Paracortex of Human Lymph Nodes
    Article Snippet: Paraffin sections were rehydrated and subjected to antigen-retrieval by boiling in 0.01 mol/L citric acid (pH 6.0) for 10 minutes before incubation with antibodies. .. Staining was performed with the ABC-AP Vectastain kit (Vector Laboratories) or ABC-PO and diaminobenzidine tetrahydrochloride (0.5 mg/ml) and sections were counterstained with hematoxylin according to Pappanicolau. .. For immunofluorescence, Alexa 488 (Molecular Probes, Eugene, OR)- and Texas Red (Jackson, West Grove, PA)-conjugated secondary antibodies were used.

    Labeling:

    Article Title: Analysis of Pathogen-Host Cell Interactions in Purpura Fulminans: Expression of Capsule, Type IV Pili, and PorA by Neisseria meningitidis In Vivo
    Article Snippet: Briefly, sections were incubated in 0.3% aqueous hydrogen peroxide to quench endogenous peroxidase activity and blocked with normal mouse or rabbit serum before incubation with the monoclonal antibodies for 1 h at room temperature. .. Labeling was detected with an immunoperoxidase kit (Vectastain Elite ABC kit; Vector Laboratories, Burlingame, Calif.), and sections were counterstained in hematoxylin. .. Antigen retrieval treatment of the sections was required for optimal staining with all of the meningococcal monoclonal antibodies and the anti-CD31 and -CD68 antibodies ( ).

    Incubation:

    Article Title: Luteolin Inhibits Human Prostate Tumor Growth by Suppressing Vascular Endothelial Growth Factor Receptor 2-Mediated Angiogenesis
    Article Snippet: Histology and Immunohistochemistry Tumor tissues were fixed in 10% neutral-buffered formalin for 24 hours, processed, and embedded in paraffin blocks. .. The sections (5 µm) were blocked with 10% goat serum and incubated with a rabbit anti-CD31 (1∶100; Novus Biologicals Inc, Littleton, CO) and mouse ant-CD34 (1∶100; BD Pharmingen Inc, San Diego, CA) antibodies for 24 h. The slides were subsequently incubated for 30 min with biotinylated anti-rabbit/anti-mouse secondary antibody (Vector laboratories, Burlingame, CA) and followed by incubation of Vectastain ABC Kit (Vector Laboratories). .. Diaminobenzidine (Sigma) was used as the chromagen and methyl green (Sigma) as the counterstain.

    Immunohistochemistry:

    Article Title: Induction of cyclooxygenase-2 in monocyte/macrophage by mucins secreted from colon cancer cells
    Article Snippet: .. Immunohistochemical staining was performed with the Vectastain avidin–biotin peroxidase complex kit (Vector Laboratories) as described ( ). .. Double immunostaining was carried out according to Kawahito et al. ( ).

    Article Title: Diet-induced obesity links to ER positive breast cancer progression via LPA/PKD-1-CD36 signaling-mediated microvascular remodeling
    Article Snippet: E0771 cells (1×106 cells/mouse) were implanted into mice close to the fourth mammary pad [ ] and tumor volume was measured using calipers in two dimensions and calculated using the formula: (width2 × length)/2 [ ]. .. Immunofluorescence and immunohistochemical assays Tissue immunohistochemical and immunofluorescence staining was performed using antibodies and methods as described in our previous studies [ , , ] or by using the Vectastain® ABC kit (Vector Laboratories). .. Mouse angiogenesis profiling A Proteome Profiler™ antibody array (R & D system® ) was performed to determine relative protein levels of angiogenic factors in the plasma according to the manufacturer's instruction.

    Avidin-Biotin Assay:

    Article Title: Induction of cyclooxygenase-2 in monocyte/macrophage by mucins secreted from colon cancer cells
    Article Snippet: .. Immunohistochemical staining was performed with the Vectastain avidin–biotin peroxidase complex kit (Vector Laboratories) as described ( ). .. Double immunostaining was carried out according to Kawahito et al. ( ).

    Article Title: Bone Matrix Proteins: Isolation and Characterization of a Novel Cell-binding Keratan Sulfate Proteoglycan (Osteoadherin) from Bovine Bone
    Article Snippet: The sections were incubated with primary antibody against osteoadherin (diluted to 1:1,000 in PBS-0.01% BSA) or the preimmune serum (diluted to 1:1,000 in PBS 0.01% BSA) at 4°C overnight in a moist chamber. .. The sections were then treated with biotinylated secondary antibody (diluted 1:200) and avidin–peroxidase conjugate using the Vectastain ABC kit™ (Vector Labs, Burlingame, CA), following the recommendations of the manufacturer. .. Cell Attachment to Osteoadherin Primary osteoblasts isolated with the method of Robey et al. ( ) were grown to near confluency in Ham's F12 medium supplemented with 10% FBS, 50 UI penicillin, and 50 μg/ml streptomycin.

    Immunofluorescence:

    Article Title: Diet-induced obesity links to ER positive breast cancer progression via LPA/PKD-1-CD36 signaling-mediated microvascular remodeling
    Article Snippet: E0771 cells (1×106 cells/mouse) were implanted into mice close to the fourth mammary pad [ ] and tumor volume was measured using calipers in two dimensions and calculated using the formula: (width2 × length)/2 [ ]. .. Immunofluorescence and immunohistochemical assays Tissue immunohistochemical and immunofluorescence staining was performed using antibodies and methods as described in our previous studies [ , , ] or by using the Vectastain® ABC kit (Vector Laboratories). .. Mouse angiogenesis profiling A Proteome Profiler™ antibody array (R & D system® ) was performed to determine relative protein levels of angiogenic factors in the plasma according to the manufacturer's instruction.

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    Vector Laboratories vectastain abc kit
    Confirmation of virus production with truncated NA RNA segments. ( A ) MDCK cells were infected with NA(−) ( a and d ), NAFLAG ( b and e ), or NAFLAGMet(−) ( c and f ) viruses and overlaid with 0.6% agarose. After incubation for 48 h at 37°C, the cells were fixed and permeated with 0.1% Triton X-100 in 3% formaldehyde solution. The viral proteins or FLAG epitope were detected by immunostaining with antiserum to influenza WSN strain ( a – c ) or anti-FLAG monoclonal antibody ( d – f ) as the primary antibody and biotinylated secondary antibody with the <t>VECTASTAIN</t> <t>ABC</t> kit (Vector Laboratories). ( B ) MDCK cells infected with NA(−), NAFLAG, or NAFLAGMet(−) viruses were incubated, fixed, and permeated as described above. The FLAG sequence in mRNA was detected by in situ hybridization with a digoxigenin-labeled probe specific for the sequence and visualized with the DIG nucleic acid-detection kit (Roche).
    Vectastain Abc Kit, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 98/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/vectastain abc kit/product/Vector Laboratories
    Average 98 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    vectastain abc kit - by Bioz Stars, 2021-06
    98/100 stars
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    98
    Vector Laboratories streptavadin horseradish peroxidase
    Focused ultrasound delivery of BAM-10 to the brain reduces Aβ plaque pathology in TgCRND8 mice. Coronal sections were stained with <t>streptavadin-Cy2</t> for ( A ) biotinylated BAM-10 and ( B ) anti-Aβ antibody 6F3D for plaques to demonstrate that 6F3D binds to plaques which are strongly (arrows) and weakly (arrowheads) positive for BAM-10 ( C ). Once it was confirmed that BAM-10 does not interfere with 6F3D plaque detection, sections for mice in each treatment group were stained with 6F3D and the stereology software was used to draw contours outlining the FUS-targeted region (determined from MRI post-treatment scans) on the right side of the brain and an equivalent region on the contralateral side ( D ). Plaques were counted and measured at high magnification ( E ), using stereological methods. In 4 days, the mean ( F ) count, ( G ) size and ( H ) surface area of Aβ plaques on the right, MRIgFUS-targeted side of the brain was consistently reduced in comparison to the left side of the brain only for the BAM-10/FUS-treated mice ( F–H ; n = 6, paired t-tests, p = 0.008, p = 0.048 and p = 0.003, respectively). This difference is unique to BAM-10/FUS treatment as there was no significant difference between right and left side of the brain in mice from other treatment groups ( F′–H′ : BAM-10 treated group, n = 6, paired t-tests, p = 0.294, p = 0.941 and p = 0.402; F″–H″ : Untreated group, n = 6, paired t-tests, p = 0.502, p = 0.690, p = 0.610). Scale bars: A–C = 100 µm (inset = 20 µm); D = 1 mm; E = 50 µm.
    Streptavadin Horseradish Peroxidase, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 98/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/streptavadin horseradish peroxidase/product/Vector Laboratories
    Average 98 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    streptavadin horseradish peroxidase - by Bioz Stars, 2021-06
    98/100 stars
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    Image Search Results


    Confirmation of virus production with truncated NA RNA segments. ( A ) MDCK cells were infected with NA(−) ( a and d ), NAFLAG ( b and e ), or NAFLAGMet(−) ( c and f ) viruses and overlaid with 0.6% agarose. After incubation for 48 h at 37°C, the cells were fixed and permeated with 0.1% Triton X-100 in 3% formaldehyde solution. The viral proteins or FLAG epitope were detected by immunostaining with antiserum to influenza WSN strain ( a – c ) or anti-FLAG monoclonal antibody ( d – f ) as the primary antibody and biotinylated secondary antibody with the VECTASTAIN ABC kit (Vector Laboratories). ( B ) MDCK cells infected with NA(−), NAFLAG, or NAFLAGMet(−) viruses were incubated, fixed, and permeated as described above. The FLAG sequence in mRNA was detected by in situ hybridization with a digoxigenin-labeled probe specific for the sequence and visualized with the DIG nucleic acid-detection kit (Roche).

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Selective incorporation of influenza virus RNA segments into virions

    doi: 10.1073/pnas.0437772100

    Figure Lengend Snippet: Confirmation of virus production with truncated NA RNA segments. ( A ) MDCK cells were infected with NA(−) ( a and d ), NAFLAG ( b and e ), or NAFLAGMet(−) ( c and f ) viruses and overlaid with 0.6% agarose. After incubation for 48 h at 37°C, the cells were fixed and permeated with 0.1% Triton X-100 in 3% formaldehyde solution. The viral proteins or FLAG epitope were detected by immunostaining with antiserum to influenza WSN strain ( a – c ) or anti-FLAG monoclonal antibody ( d – f ) as the primary antibody and biotinylated secondary antibody with the VECTASTAIN ABC kit (Vector Laboratories). ( B ) MDCK cells infected with NA(−), NAFLAG, or NAFLAGMet(−) viruses were incubated, fixed, and permeated as described above. The FLAG sequence in mRNA was detected by in situ hybridization with a digoxigenin-labeled probe specific for the sequence and visualized with the DIG nucleic acid-detection kit (Roche).

    Article Snippet: Antigens were detected with anti-FLAG monoclonal antibody M2 (Sigma) or rabbit antiserum against influenza WSN virus used as the primary antibody and biotinylated anti-mouse IgG for the FLAG epitope or biotinylated anti-rabbit IgG for viral antigens used as the secondary antibody (VECTASTAIN ABC kit, Vector Laboratories).

    Techniques: Infection, Incubation, FLAG-tag, Immunostaining, Plasmid Preparation, Sequencing, In Situ Hybridization, Labeling

    Figure 6. Immunohistochemical detection of LC3B is feasible in liver from starved Gfp-Lc3 tg/tg mice using Vectastain ABC. ( A ) Western blot analysis of LC3B in liver from wild-type mice, heterozygous Gfp-Lc3 tg/+ or homozygous Gfp-Lc3 tg/tg transgenic mice. The GFP-LC3 expression in each group was quantified. **p

    Journal: Autophagy

    Article Title: Immunohistochemical analysis of macroautophagy

    doi: 10.4161/auto.22968

    Figure Lengend Snippet: Figure 6. Immunohistochemical detection of LC3B is feasible in liver from starved Gfp-Lc3 tg/tg mice using Vectastain ABC. ( A ) Western blot analysis of LC3B in liver from wild-type mice, heterozygous Gfp-Lc3 tg/+ or homozygous Gfp-Lc3 tg/tg transgenic mice. The GFP-LC3 expression in each group was quantified. **p

    Article Snippet: Next, the slides were incubated for 30 min with biotinylated anti-rabbit or anti-mouse immunoglobulins (Vectastain ABC Kit, Vector Laboratories), 1:200 diluted in staining buffer.

    Techniques: Immunohistochemistry, Mouse Assay, Western Blot, Transgenic Assay, Expressing

    Figure 15. SQSTM1 accumulates in cytoplasmic inclusions of liver from autophagy-deficient mice. Liver samples were isolated from fed Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice (control) or from mice that underwent starvation for 48 h. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for SQSTM1 using rabbit polyclonal anti-SQSTM1 (Sigma, 1:5,000) and Vectastain ABC. Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar, 20 μm. The positive area of SQSTM1 inclusions was quantified. ***p

    Journal: Autophagy

    Article Title: Immunohistochemical analysis of macroautophagy

    doi: 10.4161/auto.22968

    Figure Lengend Snippet: Figure 15. SQSTM1 accumulates in cytoplasmic inclusions of liver from autophagy-deficient mice. Liver samples were isolated from fed Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice (control) or from mice that underwent starvation for 48 h. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for SQSTM1 using rabbit polyclonal anti-SQSTM1 (Sigma, 1:5,000) and Vectastain ABC. Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar, 20 μm. The positive area of SQSTM1 inclusions was quantified. ***p

    Article Snippet: Next, the slides were incubated for 30 min with biotinylated anti-rabbit or anti-mouse immunoglobulins (Vectastain ABC Kit, Vector Laboratories), 1:200 diluted in staining buffer.

    Techniques: Mouse Assay, Isolation, Staining

    Figure 5. The highest expression of LC3A and LC3B in nonstarved control mice was found in brain tissue. ( A ) Western blot analysis of LC3A and LC3B in different tissue lysates. GAPDH served as a loading control. ( B ) Immunohistochemical detection of LC3A and LC3B in mouse brain using the Vectastain ABC system. Tissue samples were isolated from fed control mice. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for LC3A ( A ) and LC3B ( B ) using rabbit polyclonal anti-LC3A (Abgent, 1:3000) and biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:100). Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar, 20 μm.

    Journal: Autophagy

    Article Title: Immunohistochemical analysis of macroautophagy

    doi: 10.4161/auto.22968

    Figure Lengend Snippet: Figure 5. The highest expression of LC3A and LC3B in nonstarved control mice was found in brain tissue. ( A ) Western blot analysis of LC3A and LC3B in different tissue lysates. GAPDH served as a loading control. ( B ) Immunohistochemical detection of LC3A and LC3B in mouse brain using the Vectastain ABC system. Tissue samples were isolated from fed control mice. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for LC3A ( A ) and LC3B ( B ) using rabbit polyclonal anti-LC3A (Abgent, 1:3000) and biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:100). Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar, 20 μm.

    Article Snippet: Next, the slides were incubated for 30 min with biotinylated anti-rabbit or anti-mouse immunoglobulins (Vectastain ABC Kit, Vector Laboratories), 1:200 diluted in staining buffer.

    Techniques: Expressing, Mouse Assay, Western Blot, Immunohistochemistry, Isolation, Staining

    Figure 13. ATG5 and CTSD are not suitable targets for the immunohistochemical detection of autophagy in liver. Liver samples were isolated from fed Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice (control) or from mice that underwent starvation for 48 h. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for ATG5 ( A ) or CTSD ( B ) using rabbit polyclonal anti-ATG5 (Abcam, 1:100) and rabbit monoclonal anti-CTSD (clone EPR3057Y, Abcam, 1:1000), respectively, in combination with Vectastain ABC. Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar, 20 μm. The ATG5- and CTSD-positive area was quantified. Neither the effect of starvation nor the results between Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice were statistically significant.

    Journal: Autophagy

    Article Title: Immunohistochemical analysis of macroautophagy

    doi: 10.4161/auto.22968

    Figure Lengend Snippet: Figure 13. ATG5 and CTSD are not suitable targets for the immunohistochemical detection of autophagy in liver. Liver samples were isolated from fed Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice (control) or from mice that underwent starvation for 48 h. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for ATG5 ( A ) or CTSD ( B ) using rabbit polyclonal anti-ATG5 (Abcam, 1:100) and rabbit monoclonal anti-CTSD (clone EPR3057Y, Abcam, 1:1000), respectively, in combination with Vectastain ABC. Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar, 20 μm. The ATG5- and CTSD-positive area was quantified. Neither the effect of starvation nor the results between Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice were statistically significant.

    Article Snippet: Next, the slides were incubated for 30 min with biotinylated anti-rabbit or anti-mouse immunoglobulins (Vectastain ABC Kit, Vector Laboratories), 1:200 diluted in staining buffer.

    Techniques: Immunohistochemistry, Isolation, Mouse Assay, Staining

    Figure 7. Optimal immunohistochemical detection of LC3B requires processing of tissue samples in a suitable fixative. Liver samples were isolated from Gfp-Lc3 tg/tg mice that underwent starvation for 48 h. After fixation in different fixatives for 24 h, tissues were paraffin-embedded and stained for LC3B using biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:100) and Vectastain ABC. For formalin-fixed samples, heat-mediated antigen retrieval was performed either in citrate buffer (pH 6.0) or in EDTA buffer (pH 8.0). Scale bar, 20 μm.

    Journal: Autophagy

    Article Title: Immunohistochemical analysis of macroautophagy

    doi: 10.4161/auto.22968

    Figure Lengend Snippet: Figure 7. Optimal immunohistochemical detection of LC3B requires processing of tissue samples in a suitable fixative. Liver samples were isolated from Gfp-Lc3 tg/tg mice that underwent starvation for 48 h. After fixation in different fixatives for 24 h, tissues were paraffin-embedded and stained for LC3B using biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:100) and Vectastain ABC. For formalin-fixed samples, heat-mediated antigen retrieval was performed either in citrate buffer (pH 6.0) or in EDTA buffer (pH 8.0). Scale bar, 20 μm.

    Article Snippet: Next, the slides were incubated for 30 min with biotinylated anti-rabbit or anti-mouse immunoglobulins (Vectastain ABC Kit, Vector Laboratories), 1:200 diluted in staining buffer.

    Techniques: Immunohistochemistry, Isolation, Mouse Assay, Staining

    Figure 4. Liver from autophagy-deficient Atg7 F/F Alb-Cre + mice but not from autophagy-competent Atg7 +/+ Alb-Cre + shows immunohistochemical staining for LC3A and LC3B. Liver samples were isolated from fed mice (control) or from mice that underwent starvation for 48 h. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for LC3A ( A ) and LC3B ( B ) using rabbit polyclonal anti-LC3A (Abgent, 1:100) and biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:100) in combination with Vectastain ABC. Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar, 20 μm. The LC3A and LC3B positive area was quantified. ***p

    Journal: Autophagy

    Article Title: Immunohistochemical analysis of macroautophagy

    doi: 10.4161/auto.22968

    Figure Lengend Snippet: Figure 4. Liver from autophagy-deficient Atg7 F/F Alb-Cre + mice but not from autophagy-competent Atg7 +/+ Alb-Cre + shows immunohistochemical staining for LC3A and LC3B. Liver samples were isolated from fed mice (control) or from mice that underwent starvation for 48 h. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for LC3A ( A ) and LC3B ( B ) using rabbit polyclonal anti-LC3A (Abgent, 1:100) and biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:100) in combination with Vectastain ABC. Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar, 20 μm. The LC3A and LC3B positive area was quantified. ***p

    Article Snippet: Next, the slides were incubated for 30 min with biotinylated anti-rabbit or anti-mouse immunoglobulins (Vectastain ABC Kit, Vector Laboratories), 1:200 diluted in staining buffer.

    Techniques: Mouse Assay, Immunohistochemistry, Staining, Isolation

    Figure 9. LC3B dots are detectable in frozen liver sections using a staining procedure with alkaline phosphatase and are localized on the surface of lipid droplets. Liver samples were isolated from Atg7 +/+ Alb-Cre + ( A ) or Atg7 F/F Alb-Cre + mice ( B ) that were fed normal chow (control) or underwent starvation for 48 h. After fixation in 4% paraformaldehyde, frozen sections were stained for LC3B using biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:1,000) and Vectastain ABC-AP kit, containing biotinylated alkaline phosphatase instead of biotinylated horseradish peroxidase. LC3B-positive dots (arrows) were detected on the surface of lipid droplets. These structures could be stained using oil red O. Scale bar, 20 μm.

    Journal: Autophagy

    Article Title: Immunohistochemical analysis of macroautophagy

    doi: 10.4161/auto.22968

    Figure Lengend Snippet: Figure 9. LC3B dots are detectable in frozen liver sections using a staining procedure with alkaline phosphatase and are localized on the surface of lipid droplets. Liver samples were isolated from Atg7 +/+ Alb-Cre + ( A ) or Atg7 F/F Alb-Cre + mice ( B ) that were fed normal chow (control) or underwent starvation for 48 h. After fixation in 4% paraformaldehyde, frozen sections were stained for LC3B using biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:1,000) and Vectastain ABC-AP kit, containing biotinylated alkaline phosphatase instead of biotinylated horseradish peroxidase. LC3B-positive dots (arrows) were detected on the surface of lipid droplets. These structures could be stained using oil red O. Scale bar, 20 μm.

    Article Snippet: Next, the slides were incubated for 30 min with biotinylated anti-rabbit or anti-mouse immunoglobulins (Vectastain ABC Kit, Vector Laboratories), 1:200 diluted in staining buffer.

    Techniques: Staining, Isolation, Mouse Assay

    Figure 8. Immunohistochemical staining of LC3B is enhanced in frozen liver sections. Liver samples were isolated from Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice ( A ) or from transgenic Gfp-Lc3 tg/+ or Gfp-Lc3 tg/tg mice ( B ). Some animals were fed normal chow (control), others underwent starvation for 48 h. After fixation in acetone, frozen sections were stained for LC3B using biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:1,000 [ Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + samples] or 1:30,000 [ Gfp-Lc3 tg/+ and Gfp-Lc3 tg/tg samples]) and Vectastain ABC. Scale bar, 40 μm.

    Journal: Autophagy

    Article Title: Immunohistochemical analysis of macroautophagy

    doi: 10.4161/auto.22968

    Figure Lengend Snippet: Figure 8. Immunohistochemical staining of LC3B is enhanced in frozen liver sections. Liver samples were isolated from Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice ( A ) or from transgenic Gfp-Lc3 tg/+ or Gfp-Lc3 tg/tg mice ( B ). Some animals were fed normal chow (control), others underwent starvation for 48 h. After fixation in acetone, frozen sections were stained for LC3B using biotinylated mouse monoclonal anti-LC3B (clone 5F10, Nanotools, 1:1,000 [ Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + samples] or 1:30,000 [ Gfp-Lc3 tg/+ and Gfp-Lc3 tg/tg samples]) and Vectastain ABC. Scale bar, 40 μm.

    Article Snippet: Next, the slides were incubated for 30 min with biotinylated anti-rabbit or anti-mouse immunoglobulins (Vectastain ABC Kit, Vector Laboratories), 1:200 diluted in staining buffer.

    Techniques: Immunohistochemistry, Staining, Isolation, Mouse Assay, Transgenic Assay

    Figure 14. BECN1 is upregulated in mouse tissue after starvation. ( A ) Liver samples were isolated from fed Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice (control) or from mice that underwent starvation for 48 h. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for BECN1 using rabbit polyclonal anti-BECN1 (Lifespan, 1:1,000) and Vectastain ABC. Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar 20 μm. The BECN1 positive area was quantified. ***p

    Journal: Autophagy

    Article Title: Immunohistochemical analysis of macroautophagy

    doi: 10.4161/auto.22968

    Figure Lengend Snippet: Figure 14. BECN1 is upregulated in mouse tissue after starvation. ( A ) Liver samples were isolated from fed Atg7 +/+ Alb-Cre + and Atg7 F/F Alb-Cre + mice (control) or from mice that underwent starvation for 48 h. After fixation in neutral buffered formalin for 24 h, tissues were paraffin-embedded and stained for BECN1 using rabbit polyclonal anti-BECN1 (Lifespan, 1:1,000) and Vectastain ABC. Heat-mediated antigen retrieval was performed in citrate buffer (pH 6.0). Scale bar 20 μm. The BECN1 positive area was quantified. ***p

    Article Snippet: Next, the slides were incubated for 30 min with biotinylated anti-rabbit or anti-mouse immunoglobulins (Vectastain ABC Kit, Vector Laboratories), 1:200 diluted in staining buffer.

    Techniques: Isolation, Mouse Assay, Staining

    The PrP Sc amyloid load is substantially decreased in PLTP −/− mice fed a standard chow diet. A, B: PET blot analyses of frontal tissue sections of WT or PLTP −/− mice not inoculated with prions and euthanized at 233 days (dpi) while they were healthy. C–E: PET blot analyses of frontal tissue sections of WT or PLTP −/− mice inoculated with 22L prion strain, fed a standard chow diet or Western-type cholesterol-rich diet, and all euthanized on the same day (196 dpi), when they were in an asymptomatic stage (C, D) or while they were at the terminal stage of the disease (E). The SAF84 antibody was used to detect PrP SC proteins and the Vectastain ABC-AmP kit (Vector Laboratories) was used to reveal antibody binding.

    Journal: Journal of Lipid Research

    Article Title: Plasma cholesterol level determines in vivo prion propagation [S]

    doi: 10.1194/jlr.M073718

    Figure Lengend Snippet: The PrP Sc amyloid load is substantially decreased in PLTP −/− mice fed a standard chow diet. A, B: PET blot analyses of frontal tissue sections of WT or PLTP −/− mice not inoculated with prions and euthanized at 233 days (dpi) while they were healthy. C–E: PET blot analyses of frontal tissue sections of WT or PLTP −/− mice inoculated with 22L prion strain, fed a standard chow diet or Western-type cholesterol-rich diet, and all euthanized on the same day (196 dpi), when they were in an asymptomatic stage (C, D) or while they were at the terminal stage of the disease (E). The SAF84 antibody was used to detect PrP SC proteins and the Vectastain ABC-AmP kit (Vector Laboratories) was used to reveal antibody binding.

    Article Snippet: The SAF84 antibody was used to label PrPSc and the Vectastain ABC-AmP kit (Vector Laboratories) was used to reveal antibody binding.

    Techniques: Mouse Assay, Positron Emission Tomography, Western Blot, Plasmid Preparation, Binding Assay

    Focused ultrasound delivery of BAM-10 to the brain reduces Aβ plaque pathology in TgCRND8 mice. Coronal sections were stained with streptavadin-Cy2 for ( A ) biotinylated BAM-10 and ( B ) anti-Aβ antibody 6F3D for plaques to demonstrate that 6F3D binds to plaques which are strongly (arrows) and weakly (arrowheads) positive for BAM-10 ( C ). Once it was confirmed that BAM-10 does not interfere with 6F3D plaque detection, sections for mice in each treatment group were stained with 6F3D and the stereology software was used to draw contours outlining the FUS-targeted region (determined from MRI post-treatment scans) on the right side of the brain and an equivalent region on the contralateral side ( D ). Plaques were counted and measured at high magnification ( E ), using stereological methods. In 4 days, the mean ( F ) count, ( G ) size and ( H ) surface area of Aβ plaques on the right, MRIgFUS-targeted side of the brain was consistently reduced in comparison to the left side of the brain only for the BAM-10/FUS-treated mice ( F–H ; n = 6, paired t-tests, p = 0.008, p = 0.048 and p = 0.003, respectively). This difference is unique to BAM-10/FUS treatment as there was no significant difference between right and left side of the brain in mice from other treatment groups ( F′–H′ : BAM-10 treated group, n = 6, paired t-tests, p = 0.294, p = 0.941 and p = 0.402; F″–H″ : Untreated group, n = 6, paired t-tests, p = 0.502, p = 0.690, p = 0.610). Scale bars: A–C = 100 µm (inset = 20 µm); D = 1 mm; E = 50 µm.

    Journal: PLoS ONE

    Article Title: Antibodies Targeted to the Brain with Image-Guided Focused Ultrasound Reduces Amyloid-? Plaque Load in the TgCRND8 Mouse Model of Alzheimer's Disease

    doi: 10.1371/journal.pone.0010549

    Figure Lengend Snippet: Focused ultrasound delivery of BAM-10 to the brain reduces Aβ plaque pathology in TgCRND8 mice. Coronal sections were stained with streptavadin-Cy2 for ( A ) biotinylated BAM-10 and ( B ) anti-Aβ antibody 6F3D for plaques to demonstrate that 6F3D binds to plaques which are strongly (arrows) and weakly (arrowheads) positive for BAM-10 ( C ). Once it was confirmed that BAM-10 does not interfere with 6F3D plaque detection, sections for mice in each treatment group were stained with 6F3D and the stereology software was used to draw contours outlining the FUS-targeted region (determined from MRI post-treatment scans) on the right side of the brain and an equivalent region on the contralateral side ( D ). Plaques were counted and measured at high magnification ( E ), using stereological methods. In 4 days, the mean ( F ) count, ( G ) size and ( H ) surface area of Aβ plaques on the right, MRIgFUS-targeted side of the brain was consistently reduced in comparison to the left side of the brain only for the BAM-10/FUS-treated mice ( F–H ; n = 6, paired t-tests, p = 0.008, p = 0.048 and p = 0.003, respectively). This difference is unique to BAM-10/FUS treatment as there was no significant difference between right and left side of the brain in mice from other treatment groups ( F′–H′ : BAM-10 treated group, n = 6, paired t-tests, p = 0.294, p = 0.941 and p = 0.402; F″–H″ : Untreated group, n = 6, paired t-tests, p = 0.502, p = 0.690, p = 0.610). Scale bars: A–C = 100 µm (inset = 20 µm); D = 1 mm; E = 50 µm.

    Article Snippet: 6F3D-biotin complexes were revealed with streptavadin-horseradish peroxidase (Vectastain Elite, Vector Laboratories, Burlingame, CA, USA) and DAB.

    Techniques: Mouse Assay, Staining, Software, Magnetic Resonance Imaging