immpress peroxidase micropolymer conjugated horse anti mouse  (Vector Laboratories)


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    ImmPRESS Horse Anti Mouse IgG PLUS Polymer Kit Peroxidase
    Description:
    The ImmPRESS PLUS Polymer Kit is a polymerized reporter enzyme staining system that uses novel conjugation and micropolymer chemistries to create a highly sensitive ready to use one step detection system This unique micropolymer of highly active enzyme peroxidase or alkaline phosphatase is attached to our affinity purified secondary antibodies producing reagents with outstanding sensitivity and low background The peroxidase micropolymers of the ImmPRESS HRP polymer reagent limit steric interference and provide enhanced accessibility to the target avoiding the disadvantages of other polymer systems that use large dextrans or other macromolecules as backbones The result is crisp strong staining of antibody targets especially nuclear and membrane antigens such as Ki67 estrogen receptor bcl 2 CD3 CD8 and CD10 and greater sensitivity than other polymer systems
    Catalog Number:
    mp-7802
    Price:
    None
    Size:
    1 Kit
    Category:
    Protein chromogenic detection reagents or kits or substrates
    Buy from Supplier


    Structured Review

    Vector Laboratories immpress peroxidase micropolymer conjugated horse anti mouse
    ImmPRESS Horse Anti Mouse IgG PLUS Polymer Kit Peroxidase
    The ImmPRESS PLUS Polymer Kit is a polymerized reporter enzyme staining system that uses novel conjugation and micropolymer chemistries to create a highly sensitive ready to use one step detection system This unique micropolymer of highly active enzyme peroxidase or alkaline phosphatase is attached to our affinity purified secondary antibodies producing reagents with outstanding sensitivity and low background The peroxidase micropolymers of the ImmPRESS HRP polymer reagent limit steric interference and provide enhanced accessibility to the target avoiding the disadvantages of other polymer systems that use large dextrans or other macromolecules as backbones The result is crisp strong staining of antibody targets especially nuclear and membrane antigens such as Ki67 estrogen receptor bcl 2 CD3 CD8 and CD10 and greater sensitivity than other polymer systems
    https://www.bioz.com/result/immpress peroxidase micropolymer conjugated horse anti mouse/product/Vector Laboratories
    Average 90 stars, based on 1782 article reviews
    Price from $9.99 to $1999.99
    immpress peroxidase micropolymer conjugated horse anti mouse - by Bioz Stars, 2021-02
    90/100 stars

    Images

    1) Product Images from "A novel animal model for neuroinflammation and white matter degeneration"

    Article Title: A novel animal model for neuroinflammation and white matter degeneration

    Journal: PeerJ

    doi: 10.7717/peerj.3905

    Massive mouse IgG proteins in brain tissue transduced with scAAV8-D1shRNAs. (A) Three shRNAs targeting different sites of mouse Drd1 mRNA were designed to suppress Drd1 expression. All three Drd1 shRNAs were under the control of mouse U6 promoter in scAAV-hrGFP vector. (B) The coordinates for the stereotaxic injection of scAAV8 virus into mouse striatum. Transduction efficiency of scAAV8 was examined by hrGFP expression in mouse striatum 4 weeks after surgery. Immunohistochemical staining of striatal Drd1 protein was conducted using mouse anti-Drd1 antibody in control mice (C) injected with scAAV8-hrGFP virus or mice injected with scAAV8-D1shRNA1 virus (D) 7 weeks post-injection. The virus was injected only into the left hemisphere as indicated by the arrows. ImmPRESS peroxidase-micropolymer conjugated horse anti-mouse IgG secondary antibody (without primary antibodies) was used for immunostaining of the control brain section (E) from mice injected with scAAV8-hrGFP virus and the brain section (F) from mice injected with scAAV8-D1shRNA1 virus. (G) Western blot analysis of Drd1 expression in the striatum of wildtype and Drd1 knockout mice as well as mice injected with scAAV8-D1shRNA1 virus. A putative Drd1 band migrating between 100 to 200 kD (red arrowhead) may be oligomers of Drd1 proteins that are absent in the knockout mice and reduced in the mice injected with scAAV8-D1shRNA1 virus. Mouse IgG heavy (50 kD) and light chains (25 kD) were detected only in mice injected with scAAV8-D1shRNA1 virus. The same set of samples was analyzed in Western blot with the anti-mouse IgG secondary antibody only (H). Massive mouse IgG was confirmed in mouse brain tissue transduced with scAAV8-D1shRNA1 virus.
    Figure Legend Snippet: Massive mouse IgG proteins in brain tissue transduced with scAAV8-D1shRNAs. (A) Three shRNAs targeting different sites of mouse Drd1 mRNA were designed to suppress Drd1 expression. All three Drd1 shRNAs were under the control of mouse U6 promoter in scAAV-hrGFP vector. (B) The coordinates for the stereotaxic injection of scAAV8 virus into mouse striatum. Transduction efficiency of scAAV8 was examined by hrGFP expression in mouse striatum 4 weeks after surgery. Immunohistochemical staining of striatal Drd1 protein was conducted using mouse anti-Drd1 antibody in control mice (C) injected with scAAV8-hrGFP virus or mice injected with scAAV8-D1shRNA1 virus (D) 7 weeks post-injection. The virus was injected only into the left hemisphere as indicated by the arrows. ImmPRESS peroxidase-micropolymer conjugated horse anti-mouse IgG secondary antibody (without primary antibodies) was used for immunostaining of the control brain section (E) from mice injected with scAAV8-hrGFP virus and the brain section (F) from mice injected with scAAV8-D1shRNA1 virus. (G) Western blot analysis of Drd1 expression in the striatum of wildtype and Drd1 knockout mice as well as mice injected with scAAV8-D1shRNA1 virus. A putative Drd1 band migrating between 100 to 200 kD (red arrowhead) may be oligomers of Drd1 proteins that are absent in the knockout mice and reduced in the mice injected with scAAV8-D1shRNA1 virus. Mouse IgG heavy (50 kD) and light chains (25 kD) were detected only in mice injected with scAAV8-D1shRNA1 virus. The same set of samples was analyzed in Western blot with the anti-mouse IgG secondary antibody only (H). Massive mouse IgG was confirmed in mouse brain tissue transduced with scAAV8-D1shRNA1 virus.

    Techniques Used: Transduction, Expressing, Plasmid Preparation, Injection, Immunohistochemistry, Staining, Mouse Assay, Immunostaining, Western Blot, Knock-Out

    2) Product Images from "Minimal contribution of ERK1/2-MAPK signalling towards the maintenance of oncogenic GNAQQ209P-driven uveal melanomas in zebrafish"

    Article Title: Minimal contribution of ERK1/2-MAPK signalling towards the maintenance of oncogenic GNAQQ209P-driven uveal melanomas in zebrafish

    Journal: Oncotarget

    doi: 10.18632/oncotarget.9207

    Validation of GNAQ Q209P -driven choroidal hyperplasia and uveal tumours in Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish Transverse sections of formalin-fixed and paraffin-embedded zebrafish eye tissues were stained by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (blue). Representative H E and IHC images are shown for the pre-malignant A.I-A.V. and malignant B.I-B.V. stages. (A.I) H E staining illustrating a 5x magnification of a benign choroidal hyperplasia in a 2-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (A.II-A.V) Magnifications of the region depicted in the black dashed box in A.I. (A.II) H E staining of a melanin-bleached section, revealing multiple cellular layers of the thickened choroid. (A.III) Negative control section for IHC staining. (A.IV) GNAQ expression in thickened choroid. (A.V) Hyperplastic choroidal melanocytes are positive for PCNA (dark brown nuclei), especially at the junctional zone between RPE and choroid. (B.I) H E staining illustrating a 5x magnification of a uveal tumour developing in a 5-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (B.II) Negative control section for IHC staining. (B.III) Malignant cells expressing GNAQ. (B.IV) Positive immunoreactivity to the melanocytic differentiation marker tyrosinase, indicating the melanocytic origin of malignant cells. (B.V) Numerous PCNA-positive nuclei (dark brown) in malignant melanocytes. Abbreviations: Tyr, tyrosinase; PCNA, proliferating cell nuclear antigen. Scale bars, 100 μm (A.I, B.I); 20 μm (A.II-A.V; B.II-B.V).
    Figure Legend Snippet: Validation of GNAQ Q209P -driven choroidal hyperplasia and uveal tumours in Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish Transverse sections of formalin-fixed and paraffin-embedded zebrafish eye tissues were stained by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (blue). Representative H E and IHC images are shown for the pre-malignant A.I-A.V. and malignant B.I-B.V. stages. (A.I) H E staining illustrating a 5x magnification of a benign choroidal hyperplasia in a 2-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (A.II-A.V) Magnifications of the region depicted in the black dashed box in A.I. (A.II) H E staining of a melanin-bleached section, revealing multiple cellular layers of the thickened choroid. (A.III) Negative control section for IHC staining. (A.IV) GNAQ expression in thickened choroid. (A.V) Hyperplastic choroidal melanocytes are positive for PCNA (dark brown nuclei), especially at the junctional zone between RPE and choroid. (B.I) H E staining illustrating a 5x magnification of a uveal tumour developing in a 5-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (B.II) Negative control section for IHC staining. (B.III) Malignant cells expressing GNAQ. (B.IV) Positive immunoreactivity to the melanocytic differentiation marker tyrosinase, indicating the melanocytic origin of malignant cells. (B.V) Numerous PCNA-positive nuclei (dark brown) in malignant melanocytes. Abbreviations: Tyr, tyrosinase; PCNA, proliferating cell nuclear antigen. Scale bars, 100 μm (A.I, B.I); 20 μm (A.II-A.V; B.II-B.V).

    Techniques Used: Staining, Immunohistochemistry, Negative Control, Expressing, Marker

    Oncogenic GNAQ Q209P -mediated activation of ERK and YAP signalling in choroidal melanocytes at the junction between RPE and choroid A-D. Transverse sections of formalin-fixed and paraffin-embedded eye tissues of F 1 generation, 5-month-old Tg ( mitfa :GNAQ Q209P ) zebrafish. (A) H E staining demonstrating choroidal hyperplasia (black arrowheads). White dashed box indicates the region of the choroid magnified in B-D. (B-D) Transverse sections of formalin-fixed and paraffin-embedded eye tissues were stained by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (blue). (B) Negative control: section incubated with 1x PBS instead of primary antibody. (C) Immunoreactivity to pERK1/2 (read-out of ERK activation; black arrowheads) in melanocytes at the interface between the RPE and choroid. (D) YAP-positive nuclei (read-out of YAP activation; black arrowheads) in the same cells. Scale bars, 20 μm.
    Figure Legend Snippet: Oncogenic GNAQ Q209P -mediated activation of ERK and YAP signalling in choroidal melanocytes at the junction between RPE and choroid A-D. Transverse sections of formalin-fixed and paraffin-embedded eye tissues of F 1 generation, 5-month-old Tg ( mitfa :GNAQ Q209P ) zebrafish. (A) H E staining demonstrating choroidal hyperplasia (black arrowheads). White dashed box indicates the region of the choroid magnified in B-D. (B-D) Transverse sections of formalin-fixed and paraffin-embedded eye tissues were stained by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (blue). (B) Negative control: section incubated with 1x PBS instead of primary antibody. (C) Immunoreactivity to pERK1/2 (read-out of ERK activation; black arrowheads) in melanocytes at the interface between the RPE and choroid. (D) YAP-positive nuclei (read-out of YAP activation; black arrowheads) in the same cells. Scale bars, 20 μm.

    Techniques Used: Activation Assay, Staining, Immunohistochemistry, Negative Control, Incubation

    Sporadic ERK activation contrasted with ubiquitous nuclear YAP in malignancies in Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish Sections of formalin-fixed and paraffin-embedded zebrafish eye specimens were stained for GNAQ, pERK1/2, and YAP by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (purple nuclei). A.I-A.III. Representative IHC images of benign hyperproliferative choroid. (A.I) Uniform expression of GNAQ in hyperplastic choroidal melanocytes. (A.II) Only a few cells (black arrowheads) are immunoreactive to pERK1/2. (A.III) Comparatively, more cells displayed nuclear YAP (brown nuclei), although again, this was more noticeable for cells residing at the interface. B.I-B.III; C.I-C.III. Representative IHC images of malignant choroidal melanocytes in two independent GNAQ Q209P -driven uveal tumours. (B.I, C.I) Transformed melanocytes expressing GNAQ. (B.II, C.II) Malignant cells showing only sporadic immunoreactivity to pERK1/2 (cells showing positive immunoreactivity are indicated with black arrowheads). (B.III, C.III) Ubiquitous YAP nuclear localization (brown nuclei) in transformed uveal melanocytes. Images are representative of sections from three animals. Abbreviations: RPE, retinal pigmented epithelium; RBC, red blood cells. Scale bars, 20 μm.
    Figure Legend Snippet: Sporadic ERK activation contrasted with ubiquitous nuclear YAP in malignancies in Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish Sections of formalin-fixed and paraffin-embedded zebrafish eye specimens were stained for GNAQ, pERK1/2, and YAP by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (purple nuclei). A.I-A.III. Representative IHC images of benign hyperproliferative choroid. (A.I) Uniform expression of GNAQ in hyperplastic choroidal melanocytes. (A.II) Only a few cells (black arrowheads) are immunoreactive to pERK1/2. (A.III) Comparatively, more cells displayed nuclear YAP (brown nuclei), although again, this was more noticeable for cells residing at the interface. B.I-B.III; C.I-C.III. Representative IHC images of malignant choroidal melanocytes in two independent GNAQ Q209P -driven uveal tumours. (B.I, C.I) Transformed melanocytes expressing GNAQ. (B.II, C.II) Malignant cells showing only sporadic immunoreactivity to pERK1/2 (cells showing positive immunoreactivity are indicated with black arrowheads). (B.III, C.III) Ubiquitous YAP nuclear localization (brown nuclei) in transformed uveal melanocytes. Images are representative of sections from three animals. Abbreviations: RPE, retinal pigmented epithelium; RBC, red blood cells. Scale bars, 20 μm.

    Techniques Used: Activation Assay, Staining, Immunohistochemistry, Expressing, Transformation Assay

    3) Product Images from "A novel animal model for neuroinflammation and white matter degeneration"

    Article Title: A novel animal model for neuroinflammation and white matter degeneration

    Journal: PeerJ

    doi: 10.7717/peerj.3905

    Massive mouse IgG proteins in brain tissue transduced with scAAV8-D1shRNAs. (A) Three shRNAs targeting different sites of mouse Drd1 mRNA were designed to suppress Drd1 expression. All three Drd1 shRNAs were under the control of mouse U6 promoter in scAAV-hrGFP vector. (B) The coordinates for the stereotaxic injection of scAAV8 virus into mouse striatum. Transduction efficiency of scAAV8 was examined by hrGFP expression in mouse striatum 4 weeks after surgery. Immunohistochemical staining of striatal Drd1 protein was conducted using mouse anti-Drd1 antibody in control mice (C) injected with scAAV8-hrGFP virus or mice injected with scAAV8-D1shRNA1 virus (D) 7 weeks post-injection. The virus was injected only into the left hemisphere as indicated by the arrows. ImmPRESS peroxidase-micropolymer conjugated horse anti-mouse IgG secondary antibody (without primary antibodies) was used for immunostaining of the control brain section (E) from mice injected with scAAV8-hrGFP virus and the brain section (F) from mice injected with scAAV8-D1shRNA1 virus. (G) Western blot analysis of Drd1 expression in the striatum of wildtype and Drd1 knockout mice as well as mice injected with scAAV8-D1shRNA1 virus. A putative Drd1 band migrating between 100 to 200 kD (red arrowhead) may be oligomers of Drd1 proteins that are absent in the knockout mice and reduced in the mice injected with scAAV8-D1shRNA1 virus. Mouse IgG heavy (50 kD) and light chains (25 kD) were detected only in mice injected with scAAV8-D1shRNA1 virus. The same set of samples was analyzed in Western blot with the anti-mouse IgG secondary antibody only (H). Massive mouse IgG was confirmed in mouse brain tissue transduced with scAAV8-D1shRNA1 virus.
    Figure Legend Snippet: Massive mouse IgG proteins in brain tissue transduced with scAAV8-D1shRNAs. (A) Three shRNAs targeting different sites of mouse Drd1 mRNA were designed to suppress Drd1 expression. All three Drd1 shRNAs were under the control of mouse U6 promoter in scAAV-hrGFP vector. (B) The coordinates for the stereotaxic injection of scAAV8 virus into mouse striatum. Transduction efficiency of scAAV8 was examined by hrGFP expression in mouse striatum 4 weeks after surgery. Immunohistochemical staining of striatal Drd1 protein was conducted using mouse anti-Drd1 antibody in control mice (C) injected with scAAV8-hrGFP virus or mice injected with scAAV8-D1shRNA1 virus (D) 7 weeks post-injection. The virus was injected only into the left hemisphere as indicated by the arrows. ImmPRESS peroxidase-micropolymer conjugated horse anti-mouse IgG secondary antibody (without primary antibodies) was used for immunostaining of the control brain section (E) from mice injected with scAAV8-hrGFP virus and the brain section (F) from mice injected with scAAV8-D1shRNA1 virus. (G) Western blot analysis of Drd1 expression in the striatum of wildtype and Drd1 knockout mice as well as mice injected with scAAV8-D1shRNA1 virus. A putative Drd1 band migrating between 100 to 200 kD (red arrowhead) may be oligomers of Drd1 proteins that are absent in the knockout mice and reduced in the mice injected with scAAV8-D1shRNA1 virus. Mouse IgG heavy (50 kD) and light chains (25 kD) were detected only in mice injected with scAAV8-D1shRNA1 virus. The same set of samples was analyzed in Western blot with the anti-mouse IgG secondary antibody only (H). Massive mouse IgG was confirmed in mouse brain tissue transduced with scAAV8-D1shRNA1 virus.

    Techniques Used: Transduction, Expressing, Plasmid Preparation, Injection, Immunohistochemistry, Staining, Mouse Assay, Immunostaining, Western Blot, Knock-Out

    4) Product Images from "Injection site vaccinology of a recombinant vaccinia-based vector reveals diverse innate immune signatures"

    Article Title: Injection site vaccinology of a recombinant vaccinia-based vector reveals diverse innate immune signatures

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1009215

    Vaccine genome read alignments, histology and immunohistochemistry. (A) RNA-Seq reads from each of the five groups aligned to the three viral genomes (the vector, SCV, and the two recombinant immunogen inserts from ZIKV and CHIKV); MQ—quadriceps muscles from mock vaccinated, SCV12hQ mice quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice 12 hours post vaccination, SCVd7Q -quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice taken 7 days post vaccination, MF–feet from mock vaccinated mice 7 days post vaccination, and SCVd7F - feet from SCV-ZIKA/CHIK vaccinated mice 7 days post vaccination. The number of viral reads is expressed as a percentage of the number of reads mapping to the mouse genome, with 3 biological replicates providing the SD ( S1 Fig ). The bars plotting to ≈0% had values ranging from 0 to 3.5x10 -5 %. (B) RNA-Seq reads from each of the five groups aligned to the house-keeping gene, RPL13A, also expressed as a percentage of the number of reads mapping to the mouse genome. (C) IGV visualization of reads aligned to the recombinant structural polyprotein immunogens of ZIKV (prME) and CHIKV (C-E3-E2-6K-E1), which are encoded in the SCV-ZIKA/CHIK vaccine. All reads from all replicates are shown (for details see ( S1A Table ). As expected, no reads mapped to the non-structural genes of ZIKV or CHIKV (NS1-5 and nsP1-4, respectively), as these are not encoded in SCV-ZIKA/CHIK. (Vertical purple lines for ZIKV indicate base call errors after a string of Gs). (Reads mapping to the SCV genome are shown in S1B Table ). (D) H E staining of injection site 12 hours post vaccination. Dotted ovals indicate muscle cells in early stages of necrosis (pink staining). (E) Top; IHC with anti-Ly6G staining for neutrophils (parallel section to D focusing on area of infiltrates). Bottom: Apoptag staining of the same area, illustrating apoptosis within areas of infiltrating cells. (F) Top: IHC for CHIK capsid protein. Bottom: parallel section showing IHC for ZIKA E protein. (G) Cell death annotation from Cytoscape analysis of up-regulated DEGs at 12 hours post vaccination (MQ vs SCV12hQ) ( S2C Table ) divided into non-apoptotic signatures (left) and apoptotic signatures (right). KEGG Pathways (purple), Go process (black), Reactome Pathways (brown), UniProt Keywords (green).
    Figure Legend Snippet: Vaccine genome read alignments, histology and immunohistochemistry. (A) RNA-Seq reads from each of the five groups aligned to the three viral genomes (the vector, SCV, and the two recombinant immunogen inserts from ZIKV and CHIKV); MQ—quadriceps muscles from mock vaccinated, SCV12hQ mice quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice 12 hours post vaccination, SCVd7Q -quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice taken 7 days post vaccination, MF–feet from mock vaccinated mice 7 days post vaccination, and SCVd7F - feet from SCV-ZIKA/CHIK vaccinated mice 7 days post vaccination. The number of viral reads is expressed as a percentage of the number of reads mapping to the mouse genome, with 3 biological replicates providing the SD ( S1 Fig ). The bars plotting to ≈0% had values ranging from 0 to 3.5x10 -5 %. (B) RNA-Seq reads from each of the five groups aligned to the house-keeping gene, RPL13A, also expressed as a percentage of the number of reads mapping to the mouse genome. (C) IGV visualization of reads aligned to the recombinant structural polyprotein immunogens of ZIKV (prME) and CHIKV (C-E3-E2-6K-E1), which are encoded in the SCV-ZIKA/CHIK vaccine. All reads from all replicates are shown (for details see ( S1A Table ). As expected, no reads mapped to the non-structural genes of ZIKV or CHIKV (NS1-5 and nsP1-4, respectively), as these are not encoded in SCV-ZIKA/CHIK. (Vertical purple lines for ZIKV indicate base call errors after a string of Gs). (Reads mapping to the SCV genome are shown in S1B Table ). (D) H E staining of injection site 12 hours post vaccination. Dotted ovals indicate muscle cells in early stages of necrosis (pink staining). (E) Top; IHC with anti-Ly6G staining for neutrophils (parallel section to D focusing on area of infiltrates). Bottom: Apoptag staining of the same area, illustrating apoptosis within areas of infiltrating cells. (F) Top: IHC for CHIK capsid protein. Bottom: parallel section showing IHC for ZIKA E protein. (G) Cell death annotation from Cytoscape analysis of up-regulated DEGs at 12 hours post vaccination (MQ vs SCV12hQ) ( S2C Table ) divided into non-apoptotic signatures (left) and apoptotic signatures (right). KEGG Pathways (purple), Go process (black), Reactome Pathways (brown), UniProt Keywords (green).

    Techniques Used: Immunohistochemistry, RNA Sequencing Assay, Plasmid Preparation, Recombinant, Mouse Assay, Staining, Injection

    5) Product Images from "Immunohistochemical Characterization and Sensitivity to Human Adenovirus Serotypes 3, 5, and 11p of New Cell Lines Derived from Human Diffuse Grade II to IV Gliomas"

    Article Title: Immunohistochemical Characterization and Sensitivity to Human Adenovirus Serotypes 3, 5, and 11p of New Cell Lines Derived from Human Diffuse Grade II to IV Gliomas

    Journal: Translational Oncology

    doi: 10.1016/j.tranon.2017.07.002

    Western blot analysis of adenovirus receptor expression in established and commercially available glioma cell lines (A172, U251, and U87-MG). “+” indicates positive control cells; A431 for DSG-2, 025 (original tumor lysate) for CAR, and U251 for CD46.
    Figure Legend Snippet: Western blot analysis of adenovirus receptor expression in established and commercially available glioma cell lines (A172, U251, and U87-MG). “+” indicates positive control cells; A431 for DSG-2, 025 (original tumor lysate) for CAR, and U251 for CD46.

    Techniques Used: Western Blot, Expressing, Positive Control

    Expression of adenovirus receptors (CAR, CD46, and DSG-2) in glioma tissues and cultured glioma cells. (A) Adenovirus receptor expression (brown color) was variable in paraffin sections of patient gliomas for which hematoxylin (blue color) was used as counterstain. Anaplastic oligodendroglioma 014-AOD was used as positive control in CAR and CD46 staining and rat kidney in DSG-2 staining. Among gliomas especially, 014-AOD had abundant expression of adenovirus receptors. Scale bar 100 μm. (B) Fluorescence images of immunocytochemically (red color) stained cultured cells. Nuclei were stained with DAPI (blue color). Especially CD46, receptor for serotype 11p, was well preserved among patient glioma-derived cells. A549 and 293 cells were used as positive controls. Scale bar 20 μm.
    Figure Legend Snippet: Expression of adenovirus receptors (CAR, CD46, and DSG-2) in glioma tissues and cultured glioma cells. (A) Adenovirus receptor expression (brown color) was variable in paraffin sections of patient gliomas for which hematoxylin (blue color) was used as counterstain. Anaplastic oligodendroglioma 014-AOD was used as positive control in CAR and CD46 staining and rat kidney in DSG-2 staining. Among gliomas especially, 014-AOD had abundant expression of adenovirus receptors. Scale bar 100 μm. (B) Fluorescence images of immunocytochemically (red color) stained cultured cells. Nuclei were stained with DAPI (blue color). Especially CD46, receptor for serotype 11p, was well preserved among patient glioma-derived cells. A549 and 293 cells were used as positive controls. Scale bar 20 μm.

    Techniques Used: Expressing, Cell Culture, Positive Control, Staining, Fluorescence, Derivative Assay

    6) Product Images from "Injection site vaccinology of a recombinant vaccinia-based vector reveals diverse innate immune signatures"

    Article Title: Injection site vaccinology of a recombinant vaccinia-based vector reveals diverse innate immune signatures

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1009215

    Vaccine genome read alignments, histology and immunohistochemistry. (A) RNA-Seq reads from each of the five groups aligned to the three viral genomes (the vector, SCV, and the two recombinant immunogen inserts from ZIKV and CHIKV); MQ—quadriceps muscles from mock vaccinated, SCV12hQ mice quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice 12 hours post vaccination, SCVd7Q -quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice taken 7 days post vaccination, MF–feet from mock vaccinated mice 7 days post vaccination, and SCVd7F - feet from SCV-ZIKA/CHIK vaccinated mice 7 days post vaccination. The number of viral reads is expressed as a percentage of the number of reads mapping to the mouse genome, with 3 biological replicates providing the SD ( S1 Fig ). The bars plotting to ≈0% had values ranging from 0 to 3.5x10 -5 %. (B) RNA-Seq reads from each of the five groups aligned to the house-keeping gene, RPL13A, also expressed as a percentage of the number of reads mapping to the mouse genome. (C) IGV visualization of reads aligned to the recombinant structural polyprotein immunogens of ZIKV (prME) and CHIKV (C-E3-E2-6K-E1), which are encoded in the SCV-ZIKA/CHIK vaccine. All reads from all replicates are shown (for details see ( S1A Table ). As expected, no reads mapped to the non-structural genes of ZIKV or CHIKV (NS1-5 and nsP1-4, respectively), as these are not encoded in SCV-ZIKA/CHIK. (Vertical purple lines for ZIKV indicate base call errors after a string of Gs). (Reads mapping to the SCV genome are shown in S1B Table ). (D) H E staining of injection site 12 hours post vaccination. Dotted ovals indicate muscle cells in early stages of necrosis (pink staining). (E) Top; IHC with anti-Ly6G staining for neutrophils (parallel section to D focusing on area of infiltrates). Bottom: Apoptag staining of the same area, illustrating apoptosis within areas of infiltrating cells. (F) Top: IHC for CHIK capsid protein. Bottom: parallel section showing IHC for ZIKA E protein. (G) Cell death annotation from Cytoscape analysis of up-regulated DEGs at 12 hours post vaccination (MQ vs SCV12hQ) ( S2C Table ) divided into non-apoptotic signatures (left) and apoptotic signatures (right). KEGG Pathways (purple), Go process (black), Reactome Pathways (brown), UniProt Keywords (green).
    Figure Legend Snippet: Vaccine genome read alignments, histology and immunohistochemistry. (A) RNA-Seq reads from each of the five groups aligned to the three viral genomes (the vector, SCV, and the two recombinant immunogen inserts from ZIKV and CHIKV); MQ—quadriceps muscles from mock vaccinated, SCV12hQ mice quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice 12 hours post vaccination, SCVd7Q -quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice taken 7 days post vaccination, MF–feet from mock vaccinated mice 7 days post vaccination, and SCVd7F - feet from SCV-ZIKA/CHIK vaccinated mice 7 days post vaccination. The number of viral reads is expressed as a percentage of the number of reads mapping to the mouse genome, with 3 biological replicates providing the SD ( S1 Fig ). The bars plotting to ≈0% had values ranging from 0 to 3.5x10 -5 %. (B) RNA-Seq reads from each of the five groups aligned to the house-keeping gene, RPL13A, also expressed as a percentage of the number of reads mapping to the mouse genome. (C) IGV visualization of reads aligned to the recombinant structural polyprotein immunogens of ZIKV (prME) and CHIKV (C-E3-E2-6K-E1), which are encoded in the SCV-ZIKA/CHIK vaccine. All reads from all replicates are shown (for details see ( S1A Table ). As expected, no reads mapped to the non-structural genes of ZIKV or CHIKV (NS1-5 and nsP1-4, respectively), as these are not encoded in SCV-ZIKA/CHIK. (Vertical purple lines for ZIKV indicate base call errors after a string of Gs). (Reads mapping to the SCV genome are shown in S1B Table ). (D) H E staining of injection site 12 hours post vaccination. Dotted ovals indicate muscle cells in early stages of necrosis (pink staining). (E) Top; IHC with anti-Ly6G staining for neutrophils (parallel section to D focusing on area of infiltrates). Bottom: Apoptag staining of the same area, illustrating apoptosis within areas of infiltrating cells. (F) Top: IHC for CHIK capsid protein. Bottom: parallel section showing IHC for ZIKA E protein. (G) Cell death annotation from Cytoscape analysis of up-regulated DEGs at 12 hours post vaccination (MQ vs SCV12hQ) ( S2C Table ) divided into non-apoptotic signatures (left) and apoptotic signatures (right). KEGG Pathways (purple), Go process (black), Reactome Pathways (brown), UniProt Keywords (green).

    Techniques Used: Immunohistochemistry, RNA Sequencing Assay, Plasmid Preparation, Recombinant, Mouse Assay, Staining, Injection

    7) Product Images from "Systems vaccinology analysis of a recombinant vaccinia-based vector reveals diverse innate immune signatures at the injection site"

    Article Title: Systems vaccinology analysis of a recombinant vaccinia-based vector reveals diverse innate immune signatures at the injection site

    Journal: bioRxiv

    doi: 10.1101/2020.08.17.254938

    Bowtie2 alignments to viral genomes. Raw FASTQ files were assessed for quality using FastQC and MultiQC tools. Sequencing adapters were trimmed using Trimmomatic (0.36.6) 1 where reads with an average quality score over a 4 base sliding window of less than 20 were removed. Trimmed reads were aligned using Bowtie2 (v2.3.4.1) 2 to a combined reference that included the GRCm38 primary assembly and the GENCODE M23 gene model. Vaccinia virus Copenhagen (M35027.1), Zika virus strain ZikaSPH2015 (KU321639.1), and chikungunya virus (AM258992.1). Primary proper pair reads aligned to viral features, including CDS and mature peptide features, were counted using SAMtools (v1.9). A Bar graph of raw data shown in B . 1. Bolger AM, Lohse M, Usadel B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. 2014:30(15)2114-20. 2. Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nature Methods. 2012:9(4)357-9.
    Figure Legend Snippet: Bowtie2 alignments to viral genomes. Raw FASTQ files were assessed for quality using FastQC and MultiQC tools. Sequencing adapters were trimmed using Trimmomatic (0.36.6) 1 where reads with an average quality score over a 4 base sliding window of less than 20 were removed. Trimmed reads were aligned using Bowtie2 (v2.3.4.1) 2 to a combined reference that included the GRCm38 primary assembly and the GENCODE M23 gene model. Vaccinia virus Copenhagen (M35027.1), Zika virus strain ZikaSPH2015 (KU321639.1), and chikungunya virus (AM258992.1). Primary proper pair reads aligned to viral features, including CDS and mature peptide features, were counted using SAMtools (v1.9). A Bar graph of raw data shown in B . 1. Bolger AM, Lohse M, Usadel B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. 2014:30(15)2114-20. 2. Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nature Methods. 2012:9(4)357-9.

    Techniques Used: Sequencing

    Read alignments to the viral genomes. (A) RNA-Seq reads from each of the five groups aligned to the three viral genomes (the vector, SCV, and the two recombinant immunogen inserts from ZIKV and CHIKV); MQ - quadriceps muscles from mock vaccinated, SCV12hQ mice quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice 12 hours post vaccination, SCVd7Q -quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice taken 7 days post vaccination, MF – feet from mock vaccinated mice 7 days post vaccination, and SCVd7F - feet from SCV-ZIKA/CHIK vaccinated mice 7 days post vaccination. The number of viral reads is expressed as a percentage of the number of reads mapping to the mouse genome, with 3 biological replicates providing the SD ( Supplementary Fig. 1 ). The bars plotting to ≈0% had values ranging from 0 to 3.5×10 −5 %. (B) RNA-Seq reads from each of the five groups aligned to the house-keeping gene, RPL13A, also expressed as a percentage of the number of reads mapping to the mouse genome. (C) IGV visualization of reads aligned to the recombinant structural polyprotein immunogens of ZIKV (prME) and CHIKV (C-E3-E2-6K-E1), which are encoded in the SCV-ZIKA/CHIK vaccine. All reads from all replicates are shown (for details see (S1A Table). As expected, no reads mapped to the non-structural genes of ZIKV or CHIKV (NS1-5 and nsP1-4, respectively), as these are not encoded in SCV-ZIKA/CHIK. (Vertical purple lines for ZIKV indicate base call errors after a string of Gs). (Reads mapping to the SCV genome are shown in S1B Table). (D) H E staining of injection site 12 hours post vaccination. Dotted ovals indicate muscle cells in early stages of necrosis (pink staining). (E) Top; IHC with anti-Ly6G staining for neutrophils (parallel section to D focusing on area of infiltrates). Bottom: Apoptag staining of the same area, illustrating apoptosis within areas of infiltrating cells. (F) Top: IHC for CHIK capsid protein. Bottom: parallel section showing IHC for ZIKA E protein. (G) Cell death annotation from Cytoscape analysis of up-regulated DEGs at 12 hours post vaccination (MQ vs SCV12Q) (S2C Table) divided into non-apoptotic signatures (left) and apoptotic signatures (right). KEGG Pathways (purple), Go process (black), Reactome Pathways (brown), UniProt Keywords (green).
    Figure Legend Snippet: Read alignments to the viral genomes. (A) RNA-Seq reads from each of the five groups aligned to the three viral genomes (the vector, SCV, and the two recombinant immunogen inserts from ZIKV and CHIKV); MQ - quadriceps muscles from mock vaccinated, SCV12hQ mice quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice 12 hours post vaccination, SCVd7Q -quadriceps muscles from SCV-ZIKA/CHIK vaccinated mice taken 7 days post vaccination, MF – feet from mock vaccinated mice 7 days post vaccination, and SCVd7F - feet from SCV-ZIKA/CHIK vaccinated mice 7 days post vaccination. The number of viral reads is expressed as a percentage of the number of reads mapping to the mouse genome, with 3 biological replicates providing the SD ( Supplementary Fig. 1 ). The bars plotting to ≈0% had values ranging from 0 to 3.5×10 −5 %. (B) RNA-Seq reads from each of the five groups aligned to the house-keeping gene, RPL13A, also expressed as a percentage of the number of reads mapping to the mouse genome. (C) IGV visualization of reads aligned to the recombinant structural polyprotein immunogens of ZIKV (prME) and CHIKV (C-E3-E2-6K-E1), which are encoded in the SCV-ZIKA/CHIK vaccine. All reads from all replicates are shown (for details see (S1A Table). As expected, no reads mapped to the non-structural genes of ZIKV or CHIKV (NS1-5 and nsP1-4, respectively), as these are not encoded in SCV-ZIKA/CHIK. (Vertical purple lines for ZIKV indicate base call errors after a string of Gs). (Reads mapping to the SCV genome are shown in S1B Table). (D) H E staining of injection site 12 hours post vaccination. Dotted ovals indicate muscle cells in early stages of necrosis (pink staining). (E) Top; IHC with anti-Ly6G staining for neutrophils (parallel section to D focusing on area of infiltrates). Bottom: Apoptag staining of the same area, illustrating apoptosis within areas of infiltrating cells. (F) Top: IHC for CHIK capsid protein. Bottom: parallel section showing IHC for ZIKA E protein. (G) Cell death annotation from Cytoscape analysis of up-regulated DEGs at 12 hours post vaccination (MQ vs SCV12Q) (S2C Table) divided into non-apoptotic signatures (left) and apoptotic signatures (right). KEGG Pathways (purple), Go process (black), Reactome Pathways (brown), UniProt Keywords (green).

    Techniques Used: RNA Sequencing Assay, Plasmid Preparation, Recombinant, Mouse Assay, Staining, Injection, Immunohistochemistry

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    Article Snippet: .. After incubation with the primary antibodies, ImmPRESS peroxidase-micropolymer conjugated horse anti-mouse, anti-rabbit, anti-goat IgG (Vector Labs) antibodies were used as the secondary antibody. .. Chromogenic reaction was conducted with ImmPACT NovaRED Peroxidase Substrate (Vector Labs, SK-4805) for 5 min with rotational shaking.

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    Vector Laboratories immpact novared peroxidase hrp substrate
    Validation of GNAQ Q209P -driven choroidal hyperplasia and uveal tumours in Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish Transverse sections of formalin-fixed and paraffin-embedded zebrafish eye tissues were stained by IHC, visualized by <t>ImmPact</t> <t>NovaRed</t> peroxidase <t>(HRP)</t> substrate then counterstained with hematoxylin (blue). Representative H E and IHC images are shown for the pre-malignant A.I-A.V. and malignant B.I-B.V. stages. (A.I) H E staining illustrating a 5x magnification of a benign choroidal hyperplasia in a 2-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (A.II-A.V) Magnifications of the region depicted in the black dashed box in A.I. (A.II) H E staining of a melanin-bleached section, revealing multiple cellular layers of the thickened choroid. (A.III) Negative control section for IHC staining. (A.IV) GNAQ expression in thickened choroid. (A.V) Hyperplastic choroidal melanocytes are positive for PCNA (dark brown nuclei), especially at the junctional zone between RPE and choroid. (B.I) H E staining illustrating a 5x magnification of a uveal tumour developing in a 5-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (B.II) Negative control section for IHC staining. (B.III) Malignant cells expressing GNAQ. (B.IV) Positive immunoreactivity to the melanocytic differentiation marker tyrosinase, indicating the melanocytic origin of malignant cells. (B.V) Numerous PCNA-positive nuclei (dark brown) in malignant melanocytes. Abbreviations: Tyr, tyrosinase; PCNA, proliferating cell nuclear antigen. Scale bars, 100 μm (A.I, B.I); 20 μm (A.II-A.V; B.II-B.V).
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    Validation of GNAQ Q209P -driven choroidal hyperplasia and uveal tumours in Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish Transverse sections of formalin-fixed and paraffin-embedded zebrafish eye tissues were stained by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (blue). Representative H E and IHC images are shown for the pre-malignant A.I-A.V. and malignant B.I-B.V. stages. (A.I) H E staining illustrating a 5x magnification of a benign choroidal hyperplasia in a 2-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (A.II-A.V) Magnifications of the region depicted in the black dashed box in A.I. (A.II) H E staining of a melanin-bleached section, revealing multiple cellular layers of the thickened choroid. (A.III) Negative control section for IHC staining. (A.IV) GNAQ expression in thickened choroid. (A.V) Hyperplastic choroidal melanocytes are positive for PCNA (dark brown nuclei), especially at the junctional zone between RPE and choroid. (B.I) H E staining illustrating a 5x magnification of a uveal tumour developing in a 5-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (B.II) Negative control section for IHC staining. (B.III) Malignant cells expressing GNAQ. (B.IV) Positive immunoreactivity to the melanocytic differentiation marker tyrosinase, indicating the melanocytic origin of malignant cells. (B.V) Numerous PCNA-positive nuclei (dark brown) in malignant melanocytes. Abbreviations: Tyr, tyrosinase; PCNA, proliferating cell nuclear antigen. Scale bars, 100 μm (A.I, B.I); 20 μm (A.II-A.V; B.II-B.V).

    Journal: Oncotarget

    Article Title: Minimal contribution of ERK1/2-MAPK signalling towards the maintenance of oncogenic GNAQQ209P-driven uveal melanomas in zebrafish

    doi: 10.18632/oncotarget.9207

    Figure Lengend Snippet: Validation of GNAQ Q209P -driven choroidal hyperplasia and uveal tumours in Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish Transverse sections of formalin-fixed and paraffin-embedded zebrafish eye tissues were stained by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (blue). Representative H E and IHC images are shown for the pre-malignant A.I-A.V. and malignant B.I-B.V. stages. (A.I) H E staining illustrating a 5x magnification of a benign choroidal hyperplasia in a 2-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (A.II-A.V) Magnifications of the region depicted in the black dashed box in A.I. (A.II) H E staining of a melanin-bleached section, revealing multiple cellular layers of the thickened choroid. (A.III) Negative control section for IHC staining. (A.IV) GNAQ expression in thickened choroid. (A.V) Hyperplastic choroidal melanocytes are positive for PCNA (dark brown nuclei), especially at the junctional zone between RPE and choroid. (B.I) H E staining illustrating a 5x magnification of a uveal tumour developing in a 5-month-old Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish. (B.II) Negative control section for IHC staining. (B.III) Malignant cells expressing GNAQ. (B.IV) Positive immunoreactivity to the melanocytic differentiation marker tyrosinase, indicating the melanocytic origin of malignant cells. (B.V) Numerous PCNA-positive nuclei (dark brown) in malignant melanocytes. Abbreviations: Tyr, tyrosinase; PCNA, proliferating cell nuclear antigen. Scale bars, 100 μm (A.I, B.I); 20 μm (A.II-A.V; B.II-B.V).

    Article Snippet: After washing, sections were incubated with avidin biotin complex (ABC) reagent (Vector labs) for 30 minutes at room temperature then developed with ImmPACT NovaRED Peroxidase (HRP) substrate (Vector labs) for 2 minutes at room temperature, gently washed with tap water, then counterstained with hematoxylin, dehydrated, and finally mounted with Pertex mounting medium (Cell Path) and left to dry overnight at room temperature.

    Techniques: Staining, Immunohistochemistry, Negative Control, Expressing, Marker

    Oncogenic GNAQ Q209P -mediated activation of ERK and YAP signalling in choroidal melanocytes at the junction between RPE and choroid A-D. Transverse sections of formalin-fixed and paraffin-embedded eye tissues of F 1 generation, 5-month-old Tg ( mitfa :GNAQ Q209P ) zebrafish. (A) H E staining demonstrating choroidal hyperplasia (black arrowheads). White dashed box indicates the region of the choroid magnified in B-D. (B-D) Transverse sections of formalin-fixed and paraffin-embedded eye tissues were stained by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (blue). (B) Negative control: section incubated with 1x PBS instead of primary antibody. (C) Immunoreactivity to pERK1/2 (read-out of ERK activation; black arrowheads) in melanocytes at the interface between the RPE and choroid. (D) YAP-positive nuclei (read-out of YAP activation; black arrowheads) in the same cells. Scale bars, 20 μm.

    Journal: Oncotarget

    Article Title: Minimal contribution of ERK1/2-MAPK signalling towards the maintenance of oncogenic GNAQQ209P-driven uveal melanomas in zebrafish

    doi: 10.18632/oncotarget.9207

    Figure Lengend Snippet: Oncogenic GNAQ Q209P -mediated activation of ERK and YAP signalling in choroidal melanocytes at the junction between RPE and choroid A-D. Transverse sections of formalin-fixed and paraffin-embedded eye tissues of F 1 generation, 5-month-old Tg ( mitfa :GNAQ Q209P ) zebrafish. (A) H E staining demonstrating choroidal hyperplasia (black arrowheads). White dashed box indicates the region of the choroid magnified in B-D. (B-D) Transverse sections of formalin-fixed and paraffin-embedded eye tissues were stained by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (blue). (B) Negative control: section incubated with 1x PBS instead of primary antibody. (C) Immunoreactivity to pERK1/2 (read-out of ERK activation; black arrowheads) in melanocytes at the interface between the RPE and choroid. (D) YAP-positive nuclei (read-out of YAP activation; black arrowheads) in the same cells. Scale bars, 20 μm.

    Article Snippet: After washing, sections were incubated with avidin biotin complex (ABC) reagent (Vector labs) for 30 minutes at room temperature then developed with ImmPACT NovaRED Peroxidase (HRP) substrate (Vector labs) for 2 minutes at room temperature, gently washed with tap water, then counterstained with hematoxylin, dehydrated, and finally mounted with Pertex mounting medium (Cell Path) and left to dry overnight at room temperature.

    Techniques: Activation Assay, Staining, Immunohistochemistry, Negative Control, Incubation

    Sporadic ERK activation contrasted with ubiquitous nuclear YAP in malignancies in Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish Sections of formalin-fixed and paraffin-embedded zebrafish eye specimens were stained for GNAQ, pERK1/2, and YAP by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (purple nuclei). A.I-A.III. Representative IHC images of benign hyperproliferative choroid. (A.I) Uniform expression of GNAQ in hyperplastic choroidal melanocytes. (A.II) Only a few cells (black arrowheads) are immunoreactive to pERK1/2. (A.III) Comparatively, more cells displayed nuclear YAP (brown nuclei), although again, this was more noticeable for cells residing at the interface. B.I-B.III; C.I-C.III. Representative IHC images of malignant choroidal melanocytes in two independent GNAQ Q209P -driven uveal tumours. (B.I, C.I) Transformed melanocytes expressing GNAQ. (B.II, C.II) Malignant cells showing only sporadic immunoreactivity to pERK1/2 (cells showing positive immunoreactivity are indicated with black arrowheads). (B.III, C.III) Ubiquitous YAP nuclear localization (brown nuclei) in transformed uveal melanocytes. Images are representative of sections from three animals. Abbreviations: RPE, retinal pigmented epithelium; RBC, red blood cells. Scale bars, 20 μm.

    Journal: Oncotarget

    Article Title: Minimal contribution of ERK1/2-MAPK signalling towards the maintenance of oncogenic GNAQQ209P-driven uveal melanomas in zebrafish

    doi: 10.18632/oncotarget.9207

    Figure Lengend Snippet: Sporadic ERK activation contrasted with ubiquitous nuclear YAP in malignancies in Tg ( mitfa :GNAQ Q209P ;p53 M214K/M214K ) zebrafish Sections of formalin-fixed and paraffin-embedded zebrafish eye specimens were stained for GNAQ, pERK1/2, and YAP by IHC, visualized by ImmPact NovaRed peroxidase (HRP) substrate then counterstained with hematoxylin (purple nuclei). A.I-A.III. Representative IHC images of benign hyperproliferative choroid. (A.I) Uniform expression of GNAQ in hyperplastic choroidal melanocytes. (A.II) Only a few cells (black arrowheads) are immunoreactive to pERK1/2. (A.III) Comparatively, more cells displayed nuclear YAP (brown nuclei), although again, this was more noticeable for cells residing at the interface. B.I-B.III; C.I-C.III. Representative IHC images of malignant choroidal melanocytes in two independent GNAQ Q209P -driven uveal tumours. (B.I, C.I) Transformed melanocytes expressing GNAQ. (B.II, C.II) Malignant cells showing only sporadic immunoreactivity to pERK1/2 (cells showing positive immunoreactivity are indicated with black arrowheads). (B.III, C.III) Ubiquitous YAP nuclear localization (brown nuclei) in transformed uveal melanocytes. Images are representative of sections from three animals. Abbreviations: RPE, retinal pigmented epithelium; RBC, red blood cells. Scale bars, 20 μm.

    Article Snippet: After washing, sections were incubated with avidin biotin complex (ABC) reagent (Vector labs) for 30 minutes at room temperature then developed with ImmPACT NovaRED Peroxidase (HRP) substrate (Vector labs) for 2 minutes at room temperature, gently washed with tap water, then counterstained with hematoxylin, dehydrated, and finally mounted with Pertex mounting medium (Cell Path) and left to dry overnight at room temperature.

    Techniques: Activation Assay, Staining, Immunohistochemistry, Expressing, Transformation Assay