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  • 99
    Thermo Fisher cell fluorescence
    Cell Fluorescence, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 143 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher cell disruption buffer
    Cell Disruption Buffer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 108 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher cell lysis
    Cell Lysis, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 3738 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Applications Inc hbec cells
    Hbec Cells, supplied by Cell Applications Inc, used in various techniques. Bioz Stars score: 88/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher raji cells
    Raji Cells, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 547 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Applications Inc hdlmvec cells
    AR of ccRCC cells influences angiogenesis and lymphoangiogenesis. a Outline of co-culture system. Endothelial cells (HUVEC and <t>HDLMVEC)</t> were co-cultured with ccRCC cells with indicated treatments (with/without overexpression/knockdown of AR) for 72 h, and then the endothelial cells were harvested. After harvest, the endothelial cells were maintained in mixed conditioned medium (CM from co-culture system) with fresh media at the ratio of 1:1. b , d Wound-healing assays of HUVEC and HDLMVEC cells co-cultured with A498 b and SW839 d with altered AR expression at indicated time points. Scale bar in b , d 10 μm. c , e Quantitation of endothelial cells migration in wound-healing assay after co-culture with indicated ccRCC cells as described in b , d . f , h Transwell invasion assay of HUVEC ( upper ) and HDLMVEC ( lower ) after co-culture with A498 cells overexpressing AR ( f ) and SW839 cells with AR knocked down ( h ). After 24 h, the invaded cells were stained and five random fields per well were analyzed. Scale bar in f , h , 5 μm. g , i Quantitation of the transwell invasion described in f , h . j A representative image of tube formation assays of HUVEC and HDLMVEC after co-culture with A498 overexpressing AR. The HUVEC cells during coculture with A498 cells were also blocked with VEGF-A neutralizing antibody (R D Systems) at the concentration of 0.08 µg/ml. The endothelial cells were pre-stained with Calcein AM at the concentration of 2 µM and monitored with fluorescence microscope. l Tube formation assay of HUVEC and HDLMVEC after co-culture with SW839 cells with AR knockdown. The SW839 cells were infected with lentivirus for AR and sh-VEGF-C were used for co-culture with HDLMVEC cells for the rescue assay. Scale bar in j , l , 10 μm. k , m Quantitation of the tube number described in j , l . The functional assays were all replicated four times. * p
    Hdlmvec Cells, supplied by Cell Applications Inc, used in various techniques. Bioz Stars score: 91/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Applications Inc bce cells
    AR of ccRCC cells influences angiogenesis and lymphoangiogenesis. a Outline of co-culture system. Endothelial cells (HUVEC and <t>HDLMVEC)</t> were co-cultured with ccRCC cells with indicated treatments (with/without overexpression/knockdown of AR) for 72 h, and then the endothelial cells were harvested. After harvest, the endothelial cells were maintained in mixed conditioned medium (CM from co-culture system) with fresh media at the ratio of 1:1. b , d Wound-healing assays of HUVEC and HDLMVEC cells co-cultured with A498 b and SW839 d with altered AR expression at indicated time points. Scale bar in b , d 10 μm. c , e Quantitation of endothelial cells migration in wound-healing assay after co-culture with indicated ccRCC cells as described in b , d . f , h Transwell invasion assay of HUVEC ( upper ) and HDLMVEC ( lower ) after co-culture with A498 cells overexpressing AR ( f ) and SW839 cells with AR knocked down ( h ). After 24 h, the invaded cells were stained and five random fields per well were analyzed. Scale bar in f , h , 5 μm. g , i Quantitation of the transwell invasion described in f , h . j A representative image of tube formation assays of HUVEC and HDLMVEC after co-culture with A498 overexpressing AR. The HUVEC cells during coculture with A498 cells were also blocked with VEGF-A neutralizing antibody (R D Systems) at the concentration of 0.08 µg/ml. The endothelial cells were pre-stained with Calcein AM at the concentration of 2 µM and monitored with fluorescence microscope. l Tube formation assay of HUVEC and HDLMVEC after co-culture with SW839 cells with AR knockdown. The SW839 cells were infected with lentivirus for AR and sh-VEGF-C were used for co-culture with HDLMVEC cells for the rescue assay. Scale bar in j , l , 10 μm. k , m Quantitation of the tube number described in j , l . The functional assays were all replicated four times. * p
    Bce Cells, supplied by Cell Applications Inc, used in various techniques. Bioz Stars score: 88/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher top10 competent cells
    AR of ccRCC cells influences angiogenesis and lymphoangiogenesis. a Outline of co-culture system. Endothelial cells (HUVEC and <t>HDLMVEC)</t> were co-cultured with ccRCC cells with indicated treatments (with/without overexpression/knockdown of AR) for 72 h, and then the endothelial cells were harvested. After harvest, the endothelial cells were maintained in mixed conditioned medium (CM from co-culture system) with fresh media at the ratio of 1:1. b , d Wound-healing assays of HUVEC and HDLMVEC cells co-cultured with A498 b and SW839 d with altered AR expression at indicated time points. Scale bar in b , d 10 μm. c , e Quantitation of endothelial cells migration in wound-healing assay after co-culture with indicated ccRCC cells as described in b , d . f , h Transwell invasion assay of HUVEC ( upper ) and HDLMVEC ( lower ) after co-culture with A498 cells overexpressing AR ( f ) and SW839 cells with AR knocked down ( h ). After 24 h, the invaded cells were stained and five random fields per well were analyzed. Scale bar in f , h , 5 μm. g , i Quantitation of the transwell invasion described in f , h . j A representative image of tube formation assays of HUVEC and HDLMVEC after co-culture with A498 overexpressing AR. The HUVEC cells during coculture with A498 cells were also blocked with VEGF-A neutralizing antibody (R D Systems) at the concentration of 0.08 µg/ml. The endothelial cells were pre-stained with Calcein AM at the concentration of 2 µM and monitored with fluorescence microscope. l Tube formation assay of HUVEC and HDLMVEC after co-culture with SW839 cells with AR knockdown. The SW839 cells were infected with lentivirus for AR and sh-VEGF-C were used for co-culture with HDLMVEC cells for the rescue assay. Scale bar in j , l , 10 μm. k , m Quantitation of the tube number described in j , l . The functional assays were all replicated four times. * p
    Top10 Competent Cells, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1605 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher cell suspension buffer
    AR of ccRCC cells influences angiogenesis and lymphoangiogenesis. a Outline of co-culture system. Endothelial cells (HUVEC and <t>HDLMVEC)</t> were co-cultured with ccRCC cells with indicated treatments (with/without overexpression/knockdown of AR) for 72 h, and then the endothelial cells were harvested. After harvest, the endothelial cells were maintained in mixed conditioned medium (CM from co-culture system) with fresh media at the ratio of 1:1. b , d Wound-healing assays of HUVEC and HDLMVEC cells co-cultured with A498 b and SW839 d with altered AR expression at indicated time points. Scale bar in b , d 10 μm. c , e Quantitation of endothelial cells migration in wound-healing assay after co-culture with indicated ccRCC cells as described in b , d . f , h Transwell invasion assay of HUVEC ( upper ) and HDLMVEC ( lower ) after co-culture with A498 cells overexpressing AR ( f ) and SW839 cells with AR knocked down ( h ). After 24 h, the invaded cells were stained and five random fields per well were analyzed. Scale bar in f , h , 5 μm. g , i Quantitation of the transwell invasion described in f , h . j A representative image of tube formation assays of HUVEC and HDLMVEC after co-culture with A498 overexpressing AR. The HUVEC cells during coculture with A498 cells were also blocked with VEGF-A neutralizing antibody (R D Systems) at the concentration of 0.08 µg/ml. The endothelial cells were pre-stained with Calcein AM at the concentration of 2 µM and monitored with fluorescence microscope. l Tube formation assay of HUVEC and HDLMVEC after co-culture with SW839 cells with AR knockdown. The SW839 cells were infected with lentivirus for AR and sh-VEGF-C were used for co-culture with HDLMVEC cells for the rescue assay. Scale bar in j , l , 10 μm. k , m Quantitation of the tube number described in j , l . The functional assays were all replicated four times. * p
    Cell Suspension Buffer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 27 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher hek293 cells
    Immunostaining of <t>HEK293</t> cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.
    Hek293 Cells, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 29257 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Graph Pad Software Inc cnt cells
    Immunostaining of <t>HEK293</t> cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.
    Cnt Cells, supplied by Graph Pad Software Inc, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher cell permeant
    Immunostaining of <t>HEK293</t> cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.
    Cell Permeant, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 162 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Applications Inc cell growth media
    Immunostaining of <t>HEK293</t> cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.
    Cell Growth Media, supplied by Cell Applications Inc, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher cells to ct kit
    Immunostaining of <t>HEK293</t> cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.
    Cells To Ct Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 507 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher mammalian cells
    Immunostaining of <t>HEK293</t> cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.
    Mammalian Cells, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 271 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher taqman cells
    Immunostaining of <t>HEK293</t> cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.
    Taqman Cells, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher cells kit
    Immunostaining of <t>HEK293</t> cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.
    Cells Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 70 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Sonics & Materials vibra cell vcx130
    Immunostaining of <t>HEK293</t> cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.
    Vibra Cell Vcx130, supplied by Sonics & Materials, used in various techniques. Bioz Stars score: 93/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    AR of ccRCC cells influences angiogenesis and lymphoangiogenesis. a Outline of co-culture system. Endothelial cells (HUVEC and HDLMVEC) were co-cultured with ccRCC cells with indicated treatments (with/without overexpression/knockdown of AR) for 72 h, and then the endothelial cells were harvested. After harvest, the endothelial cells were maintained in mixed conditioned medium (CM from co-culture system) with fresh media at the ratio of 1:1. b , d Wound-healing assays of HUVEC and HDLMVEC cells co-cultured with A498 b and SW839 d with altered AR expression at indicated time points. Scale bar in b , d 10 μm. c , e Quantitation of endothelial cells migration in wound-healing assay after co-culture with indicated ccRCC cells as described in b , d . f , h Transwell invasion assay of HUVEC ( upper ) and HDLMVEC ( lower ) after co-culture with A498 cells overexpressing AR ( f ) and SW839 cells with AR knocked down ( h ). After 24 h, the invaded cells were stained and five random fields per well were analyzed. Scale bar in f , h , 5 μm. g , i Quantitation of the transwell invasion described in f , h . j A representative image of tube formation assays of HUVEC and HDLMVEC after co-culture with A498 overexpressing AR. The HUVEC cells during coculture with A498 cells were also blocked with VEGF-A neutralizing antibody (R D Systems) at the concentration of 0.08 µg/ml. The endothelial cells were pre-stained with Calcein AM at the concentration of 2 µM and monitored with fluorescence microscope. l Tube formation assay of HUVEC and HDLMVEC after co-culture with SW839 cells with AR knockdown. The SW839 cells were infected with lentivirus for AR and sh-VEGF-C were used for co-culture with HDLMVEC cells for the rescue assay. Scale bar in j , l , 10 μm. k , m Quantitation of the tube number described in j , l . The functional assays were all replicated four times. * p

    Journal: Nature Communications

    Article Title: Androgen receptor increases hematogenous metastasis yet decreases lymphatic metastasis of renal cell carcinoma

    doi: 10.1038/s41467-017-00701-6

    Figure Lengend Snippet: AR of ccRCC cells influences angiogenesis and lymphoangiogenesis. a Outline of co-culture system. Endothelial cells (HUVEC and HDLMVEC) were co-cultured with ccRCC cells with indicated treatments (with/without overexpression/knockdown of AR) for 72 h, and then the endothelial cells were harvested. After harvest, the endothelial cells were maintained in mixed conditioned medium (CM from co-culture system) with fresh media at the ratio of 1:1. b , d Wound-healing assays of HUVEC and HDLMVEC cells co-cultured with A498 b and SW839 d with altered AR expression at indicated time points. Scale bar in b , d 10 μm. c , e Quantitation of endothelial cells migration in wound-healing assay after co-culture with indicated ccRCC cells as described in b , d . f , h Transwell invasion assay of HUVEC ( upper ) and HDLMVEC ( lower ) after co-culture with A498 cells overexpressing AR ( f ) and SW839 cells with AR knocked down ( h ). After 24 h, the invaded cells were stained and five random fields per well were analyzed. Scale bar in f , h , 5 μm. g , i Quantitation of the transwell invasion described in f , h . j A representative image of tube formation assays of HUVEC and HDLMVEC after co-culture with A498 overexpressing AR. The HUVEC cells during coculture with A498 cells were also blocked with VEGF-A neutralizing antibody (R D Systems) at the concentration of 0.08 µg/ml. The endothelial cells were pre-stained with Calcein AM at the concentration of 2 µM and monitored with fluorescence microscope. l Tube formation assay of HUVEC and HDLMVEC after co-culture with SW839 cells with AR knockdown. The SW839 cells were infected with lentivirus for AR and sh-VEGF-C were used for co-culture with HDLMVEC cells for the rescue assay. Scale bar in j , l , 10 μm. k , m Quantitation of the tube number described in j , l . The functional assays were all replicated four times. * p

    Article Snippet: HDLMVEC cells were purchased from Cell Applications, Inc. (San Diego, CA) and maintained with endothelial cell growth medium in attachment factor solution pre-coated flasks.

    Techniques: Co-Culture Assay, Cell Culture, Over Expression, Expressing, Quantitation Assay, Migration, Wound Healing Assay, Transwell Invasion Assay, Staining, Concentration Assay, Fluorescence, Microscopy, Tube Formation Assay, Infection, Rescue Assay, Functional Assay

    Immunostaining of HEK293 cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.

    Journal: The Journal of Neuroscience

    Article Title: Surface Targeting of the Dopamine Transporter Involves Discrete Epitopes in the Distal C Terminus But Does Not Require Canonical PDZ Domain Interactions

    doi: 10.1523/JNEUROSCI.1863-04.2004

    Figure Lengend Snippet: Immunostaining of HEK293 cells stably expressing EGFP-PICK1 alone or coexpressing WT, +Ala, 3xAla_618-620, or 3xAla_615-617 DAT and EGFP-PICK1. The cells were fixed and incubated with primary MAB369 anti-DAT antibody (1:1000), followed by secondary goat anti-rat Alexa Fluor 568 antibody and visualization by confocal fluorescence microscopy, as described in Materials and Methods. The anti-DAT (red) is shown on the left, the EGFP-PICK1 (green) is in the middle, and the overlay is on the right. Scale bar, 10 μm. All pictures have the same magnification.

    Article Snippet: Membrane protein (10 μg), prepared from HEK293 cells as described above, was assayed in a total volume of 250 μl using a sodium phosphate buffer (50 m m Na2 HPO4 -NaH2 PO4 , pH 7.4) containing ∼0.25 n m [125 I]RTI-55 (PE Applied Biosystems, Foster City, CA) and increasing concentrations of competing nonlabeled RTI-55.

    Techniques: Immunostaining, Stable Transfection, Expressing, Incubation, Fluorescence, Microscopy

    Surface expression of WT and mutants transporters assessed by biotinylation. A , Surface biotinylation of HEK293 cells expressing WT, ΔLKV, β 2 -23, β 2 -3, or +Ala (left). Comparison of nontransfected cells with WT transfected cells (right). Surface biotinylated protein was purified using streptavidin beads and analyzed by SDS-PAGE, followed by immunoblotting with the M2 anti-FLAG antibody, as described in Materials and Methods. The surface-expressed hDAT migrated as a single glycosylated band corresponding to a molecular weight of ∼90 kDa. B , SDS-PAGE and immunoblotting with the M2 anti-FLAG antibody of total cellular protein (15 μg) from cells expressing WT, ΔLKV, β 2 -23, β 2 ). The gel shown is representative of at least three independent experiments.

    Journal: The Journal of Neuroscience

    Article Title: Surface Targeting of the Dopamine Transporter Involves Discrete Epitopes in the Distal C Terminus But Does Not Require Canonical PDZ Domain Interactions

    doi: 10.1523/JNEUROSCI.1863-04.2004

    Figure Lengend Snippet: Surface expression of WT and mutants transporters assessed by biotinylation. A , Surface biotinylation of HEK293 cells expressing WT, ΔLKV, β 2 -23, β 2 -3, or +Ala (left). Comparison of nontransfected cells with WT transfected cells (right). Surface biotinylated protein was purified using streptavidin beads and analyzed by SDS-PAGE, followed by immunoblotting with the M2 anti-FLAG antibody, as described in Materials and Methods. The surface-expressed hDAT migrated as a single glycosylated band corresponding to a molecular weight of ∼90 kDa. B , SDS-PAGE and immunoblotting with the M2 anti-FLAG antibody of total cellular protein (15 μg) from cells expressing WT, ΔLKV, β 2 -23, β 2 ). The gel shown is representative of at least three independent experiments.

    Article Snippet: Membrane protein (10 μg), prepared from HEK293 cells as described above, was assayed in a total volume of 250 μl using a sodium phosphate buffer (50 m m Na2 HPO4 -NaH2 PO4 , pH 7.4) containing ∼0.25 n m [125 I]RTI-55 (PE Applied Biosystems, Foster City, CA) and increasing concentrations of competing nonlabeled RTI-55.

    Techniques: Expressing, Transfection, Purification, SDS Page, Molecular Weight

    Effect of proteasome 26S inhibition on degradation of the DAT and ΔLKV in HEK293 cells. A , The ΔLKV and the WT ( B ) after incubation with MG132 for 0, 3, 6, or 9 hr, followed by incubation with PNGaseF or EndoH. Samples were analyzed by SDS-PAGE, followed by immunoblotting with the MAB369 anti-DAT antibody, as described in Materials and Methods. The fully mature N-glycosylated hDAT, which is sensitive to PNGaseF, eluted corresponding to a molecular weight of 90 kDa. The immature lyglycosylated form, which is sensitive to EndoH, eluted at ∼60 kDa. Complete deglycosylation by PNGaseF or EndoH results in elution of the DAT at ∼50 kDa. Protein from untransfected cells is included in the right lane as control.

    Journal: The Journal of Neuroscience

    Article Title: Surface Targeting of the Dopamine Transporter Involves Discrete Epitopes in the Distal C Terminus But Does Not Require Canonical PDZ Domain Interactions

    doi: 10.1523/JNEUROSCI.1863-04.2004

    Figure Lengend Snippet: Effect of proteasome 26S inhibition on degradation of the DAT and ΔLKV in HEK293 cells. A , The ΔLKV and the WT ( B ) after incubation with MG132 for 0, 3, 6, or 9 hr, followed by incubation with PNGaseF or EndoH. Samples were analyzed by SDS-PAGE, followed by immunoblotting with the MAB369 anti-DAT antibody, as described in Materials and Methods. The fully mature N-glycosylated hDAT, which is sensitive to PNGaseF, eluted corresponding to a molecular weight of 90 kDa. The immature lyglycosylated form, which is sensitive to EndoH, eluted at ∼60 kDa. Complete deglycosylation by PNGaseF or EndoH results in elution of the DAT at ∼50 kDa. Protein from untransfected cells is included in the right lane as control.

    Article Snippet: Membrane protein (10 μg), prepared from HEK293 cells as described above, was assayed in a total volume of 250 μl using a sodium phosphate buffer (50 m m Na2 HPO4 -NaH2 PO4 , pH 7.4) containing ∼0.25 n m [125 I]RTI-55 (PE Applied Biosystems, Foster City, CA) and increasing concentrations of competing nonlabeled RTI-55.

    Techniques: Inhibition, Incubation, SDS Page, Molecular Weight