lipid transfection  (Qiagen)

 
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    Name:
    Effectene Transfection Reagent
    Description:
    For DNA transfection of primary cells and sensitive cell lines Kit contents Qiagen Effectene Transfection Reagent 1mL For 160 Transfections in 60mm Dishes or 640 Transfections in 12 well Plates Eukaryotic Cell Type For DNA Transfection of Primary Cells and Sensitive Cell Lines Non liposomal Lipid Formulation in Conjonction with a DNA Condensing Enhancer High Efficiency in the Presence of Serum Suitable for High throughput Screening Includes Enhancer Buffer Benefits High efficiency in the presence of serum Efficient transfection with low DNA amounts Far lower cytotoxicity and gentler than many alternatives Suitable for high throughpu
    Catalog Number:
    301425
    Price:
    339
    Category:
    Effectene Transfection Reagent
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    Structured Review

    Qiagen lipid transfection
    Effectene Transfection Reagent
    For DNA transfection of primary cells and sensitive cell lines Kit contents Qiagen Effectene Transfection Reagent 1mL For 160 Transfections in 60mm Dishes or 640 Transfections in 12 well Plates Eukaryotic Cell Type For DNA Transfection of Primary Cells and Sensitive Cell Lines Non liposomal Lipid Formulation in Conjonction with a DNA Condensing Enhancer High Efficiency in the Presence of Serum Suitable for High throughput Screening Includes Enhancer Buffer Benefits High efficiency in the presence of serum Efficient transfection with low DNA amounts Far lower cytotoxicity and gentler than many alternatives Suitable for high throughpu
    https://www.bioz.com/result/lipid transfection/product/Qiagen
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    lipid transfection - by Bioz Stars, 2020-07
    99/100 stars

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    Related Articles

    Transfection:

    Article Title: Combined effects of 17-DMAG and TNF on cells through a mechanism related to the NF-kappaB pathway
    Article Snippet: .. The control pGL3 vector DNA (0.2 μg) (purchased from Clontech Co.) or similar amounts of Luciferase-NF-κB reporter vectors (Clontech Co., USA), was cotransfected into cells with the pRL-SV40 vector using the effectene transfection reagent (Qiagen, USA) for 20 h. Cells were then treated with DMSO (0.016%, v/v), TNF (10 ng/ml), 17-DMAG (0.05 μM), or TNF (10 ng/ml) plus 17-DMAG (0.05 μM) for 24 h. Luciferase activity was measured using the Dual-Luciferase Reporter Assay System (Promega, USA) according to the manufacturer's protocol. .. The transcriptional activity of NF-κB in the cells transfected with the Luciferase-NF-κB reporter vectors were calculated as folds of those in the relative cells transfected with the empty vector.

    Article Title: Effect of APOL1 disease risk variants on APOL1 gene product
    Article Snippet: .. Equal copy number/amount of plasmid was transfected using Effectene Transfection Regent Kit (Qiagen cat # 301425). ..

    Article Title: Glyceraldehyde-3-Phosphate Dehydrogenase in Retinal Microvasculature: Implications for the Development and Progression of Diabetic Retinopathy
    Article Snippet: .. Transfection complexes were formed with transfection reagent (Effectene; Qiagen, Valencia, CA) and were incubated with BRECs for 8 hours before incubation in 5 mM or 20 mM glucose media for 4 days. .. Parallel incubations were carried out by incubating the cells in the transfection reagent alone for 8 hours followed by incubation in 5 mM or 20 mM glucose media. (Our previous studies have shown that control plasmid vectors [pGL3] or scrambled siRNA have no effect on retinal endothelial cells , ).

    Article Title: Role of Bruton’s Tyrosine Kinase inhibitors in HIV-1 infected cells
    Article Snippet: .. HIV-1 proviral DNA construct pNL4.3 (5 µg) was also transfected in U937 and Jurkat cells using Effectene (Qiagen) according to the manufacturer's instructions ( ). .. Both uninfected U937 and infected U1 monocytic cells were washed with PBS without Mg2+ and Ca2+ and resuspended in lysis buffer (50 mM Tris–HCl pH 7.5, 120 mM NaCl, 5 mM EDTA, 0.5% NP-40, 50 mM NaF, 0.2 mM Na3 VO4 , 1 mM DTT, and one complete protease cocktail tablet per 50 ml) and incubated on ice for 20 minutes, with gentle vortexing.

    Article Title: Fluorescent labeling of tetracysteine-tagged proteins in intact cells
    Article Snippet: .. Routinely, we use HEK293 cells transiently transfected with any standard transfection method using either calcium phosphate or transfection reagents such as Effectene (Qiagen), Lipofectamine (Invitrogen) or GeneJuice (Novagen). ..

    Article Title: EBV noncoding RNA EBER2 interacts with host RNA-binding proteins to regulate viral gene expression
    Article Snippet: .. HEK 293T cells were transfected with EBER2 expression plasmid (pBS-5×EBER2) together with each of the FLAG expression plasmids using Effectene (Qiagen). .. UV irradiation followed by immunoprecipitation was carried out as described ( ).

    Article Title: Human Airway Epithelial Cells Sense Pseudomonas aeruginosa Infection via Recognition of Flagellin by Toll-Like Receptor 5
    Article Snippet: .. Efficient transfection of the primary HAECs was obtained by using the Effectene reagent. .. Firefly and Rinilla luciferase reporter genes were introduced into the cells as reporters for the dual-luciferase assay.

    Article Title: ELMO1 and Dock180, a Bipartite Rac1 Guanine Nucleotide Exchange Factor, Promote Human Glioma Cell Invasion
    Article Snippet: .. A pCG plasmid encoding c-Myc– and His-tagged full-length human ELMO1 (from Dr. J. Skowronski, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; ref. ) and/or a pCXN2 plasmid encoding Flag-tagged full-length human Dock180 (from Dr. M. Matsuda, Kyoto University, Kyoto, Japan) was transfected into U87MG and U251MG cells using Effectene following the manufacturer’s instructions (Qiagen). .. Forty-eight hours posttransfection, the cells were used in in vitro cell migration, invasion, and ex vivo brain slice assays.

    Luciferase:

    Article Title: Combined effects of 17-DMAG and TNF on cells through a mechanism related to the NF-kappaB pathway
    Article Snippet: .. The control pGL3 vector DNA (0.2 μg) (purchased from Clontech Co.) or similar amounts of Luciferase-NF-κB reporter vectors (Clontech Co., USA), was cotransfected into cells with the pRL-SV40 vector using the effectene transfection reagent (Qiagen, USA) for 20 h. Cells were then treated with DMSO (0.016%, v/v), TNF (10 ng/ml), 17-DMAG (0.05 μM), or TNF (10 ng/ml) plus 17-DMAG (0.05 μM) for 24 h. Luciferase activity was measured using the Dual-Luciferase Reporter Assay System (Promega, USA) according to the manufacturer's protocol. .. The transcriptional activity of NF-κB in the cells transfected with the Luciferase-NF-κB reporter vectors were calculated as folds of those in the relative cells transfected with the empty vector.

    Reporter Assay:

    Article Title: Combined effects of 17-DMAG and TNF on cells through a mechanism related to the NF-kappaB pathway
    Article Snippet: .. The control pGL3 vector DNA (0.2 μg) (purchased from Clontech Co.) or similar amounts of Luciferase-NF-κB reporter vectors (Clontech Co., USA), was cotransfected into cells with the pRL-SV40 vector using the effectene transfection reagent (Qiagen, USA) for 20 h. Cells were then treated with DMSO (0.016%, v/v), TNF (10 ng/ml), 17-DMAG (0.05 μM), or TNF (10 ng/ml) plus 17-DMAG (0.05 μM) for 24 h. Luciferase activity was measured using the Dual-Luciferase Reporter Assay System (Promega, USA) according to the manufacturer's protocol. .. The transcriptional activity of NF-κB in the cells transfected with the Luciferase-NF-κB reporter vectors were calculated as folds of those in the relative cells transfected with the empty vector.

    Construct:

    Article Title: Role of Bruton’s Tyrosine Kinase inhibitors in HIV-1 infected cells
    Article Snippet: .. HIV-1 proviral DNA construct pNL4.3 (5 µg) was also transfected in U937 and Jurkat cells using Effectene (Qiagen) according to the manufacturer's instructions ( ). .. Both uninfected U937 and infected U1 monocytic cells were washed with PBS without Mg2+ and Ca2+ and resuspended in lysis buffer (50 mM Tris–HCl pH 7.5, 120 mM NaCl, 5 mM EDTA, 0.5% NP-40, 50 mM NaF, 0.2 mM Na3 VO4 , 1 mM DTT, and one complete protease cocktail tablet per 50 ml) and incubated on ice for 20 minutes, with gentle vortexing.

    Incubation:

    Article Title: Glyceraldehyde-3-Phosphate Dehydrogenase in Retinal Microvasculature: Implications for the Development and Progression of Diabetic Retinopathy
    Article Snippet: .. Transfection complexes were formed with transfection reagent (Effectene; Qiagen, Valencia, CA) and were incubated with BRECs for 8 hours before incubation in 5 mM or 20 mM glucose media for 4 days. .. Parallel incubations were carried out by incubating the cells in the transfection reagent alone for 8 hours followed by incubation in 5 mM or 20 mM glucose media. (Our previous studies have shown that control plasmid vectors [pGL3] or scrambled siRNA have no effect on retinal endothelial cells , ).

    Activity Assay:

    Article Title: Combined effects of 17-DMAG and TNF on cells through a mechanism related to the NF-kappaB pathway
    Article Snippet: .. The control pGL3 vector DNA (0.2 μg) (purchased from Clontech Co.) or similar amounts of Luciferase-NF-κB reporter vectors (Clontech Co., USA), was cotransfected into cells with the pRL-SV40 vector using the effectene transfection reagent (Qiagen, USA) for 20 h. Cells were then treated with DMSO (0.016%, v/v), TNF (10 ng/ml), 17-DMAG (0.05 μM), or TNF (10 ng/ml) plus 17-DMAG (0.05 μM) for 24 h. Luciferase activity was measured using the Dual-Luciferase Reporter Assay System (Promega, USA) according to the manufacturer's protocol. .. The transcriptional activity of NF-κB in the cells transfected with the Luciferase-NF-κB reporter vectors were calculated as folds of those in the relative cells transfected with the empty vector.

    Expressing:

    Article Title: EBV noncoding RNA EBER2 interacts with host RNA-binding proteins to regulate viral gene expression
    Article Snippet: .. HEK 293T cells were transfected with EBER2 expression plasmid (pBS-5×EBER2) together with each of the FLAG expression plasmids using Effectene (Qiagen). .. UV irradiation followed by immunoprecipitation was carried out as described ( ).

    Plasmid Preparation:

    Article Title: Combined effects of 17-DMAG and TNF on cells through a mechanism related to the NF-kappaB pathway
    Article Snippet: .. The control pGL3 vector DNA (0.2 μg) (purchased from Clontech Co.) or similar amounts of Luciferase-NF-κB reporter vectors (Clontech Co., USA), was cotransfected into cells with the pRL-SV40 vector using the effectene transfection reagent (Qiagen, USA) for 20 h. Cells were then treated with DMSO (0.016%, v/v), TNF (10 ng/ml), 17-DMAG (0.05 μM), or TNF (10 ng/ml) plus 17-DMAG (0.05 μM) for 24 h. Luciferase activity was measured using the Dual-Luciferase Reporter Assay System (Promega, USA) according to the manufacturer's protocol. .. The transcriptional activity of NF-κB in the cells transfected with the Luciferase-NF-κB reporter vectors were calculated as folds of those in the relative cells transfected with the empty vector.

    Article Title: Effect of APOL1 disease risk variants on APOL1 gene product
    Article Snippet: .. Equal copy number/amount of plasmid was transfected using Effectene Transfection Regent Kit (Qiagen cat # 301425). ..

    Article Title: EBV noncoding RNA EBER2 interacts with host RNA-binding proteins to regulate viral gene expression
    Article Snippet: .. HEK 293T cells were transfected with EBER2 expression plasmid (pBS-5×EBER2) together with each of the FLAG expression plasmids using Effectene (Qiagen). .. UV irradiation followed by immunoprecipitation was carried out as described ( ).

    Article Title: ELMO1 and Dock180, a Bipartite Rac1 Guanine Nucleotide Exchange Factor, Promote Human Glioma Cell Invasion
    Article Snippet: .. A pCG plasmid encoding c-Myc– and His-tagged full-length human ELMO1 (from Dr. J. Skowronski, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; ref. ) and/or a pCXN2 plasmid encoding Flag-tagged full-length human Dock180 (from Dr. M. Matsuda, Kyoto University, Kyoto, Japan) was transfected into U87MG and U251MG cells using Effectene following the manufacturer’s instructions (Qiagen). .. Forty-eight hours posttransfection, the cells were used in in vitro cell migration, invasion, and ex vivo brain slice assays.

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  • 92
    Qiagen transfection complexes
    Exosome concentration affects electroporation efficiency. To determine the optimal exosome concentration for <t>transfection,</t> exosomes at four different concentrations (0.25–4 µg/µl) were mixed with an Alexa Fluor 488-tagged siRNA (2 nmol/ml) and electroporated. Samples containing exosomes and siRNA, without an electric pulse, were used as a negative control. To eliminate the excess siRNA, the exosomes were purified using latex beads and the presence of siRNA in the exosomes was determined by FACS analysis.
    Transfection Complexes, supplied by Qiagen, used in various techniques. Bioz Stars score: 92/100, based on 32 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/transfection complexes/product/Qiagen
    Average 92 stars, based on 32 article reviews
    Price from $9.99 to $1999.99
    transfection complexes - by Bioz Stars, 2020-07
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    85
    Qiagen standard lipid based transfection
    <t>Transfection</t> and Transduction Efficiency
    Standard Lipid Based Transfection, supplied by Qiagen, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/standard lipid based transfection/product/Qiagen
    Average 85 stars, based on 1 article reviews
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    85/100 stars
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    99
    Qiagen superfect
    Comparative transfection efficiency of FuGENE HD (FuGENE), X-tremeGENE (X-treme), <t>SuperFect</t> (Super), Lipofectamine 2000 (L-2000), Lipofectamine RNAiMAX (L-iMAX), INT, and mTat/PEI/INT using siRNA targeting β - actin in HSC-3 cells. β - actin mRNA was measured by QRT-PCR and then % remaining β - actin mRNA expression was calculated based on control as 100% ( a ). Cell viability of each of the reagent groups in HaCaT cells were evaluated by MTT assay under the same siRNA and transfection reagent volume ( b ). * p
    Superfect, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 1366 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/superfect/product/Qiagen
    Average 99 stars, based on 1366 article reviews
    Price from $9.99 to $1999.99
    superfect - by Bioz Stars, 2020-07
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    99
    Qiagen cationic lipid transfection reagent
    Mutational disruption of NK1R-mediated sequestration of β-arrestin prevents inhibition of F-MOR endocytosis in N2A cells (A) Confocal micrographs of N2A cells co-expressing either full-length HA-NK1R (left panels, red) or truncated HA-NK1R (right panels, red) with β-arrestin2-EGFP (green). Incubation with 10μM SP in cells expressing the truncated HA-NK1R (355x) resulted in less co-localization with receptors in endosomal clusters and higher cytoplasmic β-arrestin2-EGFP distribution (left column), compared to β-arrestin2-EGFP distribution in the wild type HA-NK1R under the same conditions (right column). Scale bar, 10μm. (B) In N2A cells co-expressing F-MOR (red) and HA-NK1 355x receptors (green), both morphine and DAMGO were able to drive internalization of F-MORs even in the presence of substance P. Scale bar, 10μm. (C) Quantification using ratiometric staining of F-MORs in N2A cells co-expressing HA-NK1 355x revealed that co-incubation with SP produced comparable levels of F-MOR internalization compared to treatment with either MS or DG alone. Bar graphs represent mean internalization determined from ~50 cell bodies selected at random in the N2A cultures and averaged over four independent <t>transfections.</t> (D) Biochemical assay of F-MOR internalization by surface biotinylation confirmed that F-MOR internalization induced by either MS or DG (lanes 3 and 5) was not significantly inhibited by co-application of SP (lanes 4 and 6; n= 3 independent experiments).
    Cationic Lipid Transfection Reagent, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cationic lipid transfection reagent/product/Qiagen
    Average 99 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    cationic lipid transfection reagent - by Bioz Stars, 2020-07
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    Image Search Results


    Exosome concentration affects electroporation efficiency. To determine the optimal exosome concentration for transfection, exosomes at four different concentrations (0.25–4 µg/µl) were mixed with an Alexa Fluor 488-tagged siRNA (2 nmol/ml) and electroporated. Samples containing exosomes and siRNA, without an electric pulse, were used as a negative control. To eliminate the excess siRNA, the exosomes were purified using latex beads and the presence of siRNA in the exosomes was determined by FACS analysis.

    Journal: Nucleic Acids Research

    Article Title: Plasma exosomes can deliver exogenous short interfering RNA to monocytes and lymphocytes

    doi: 10.1093/nar/gks463

    Figure Lengend Snippet: Exosome concentration affects electroporation efficiency. To determine the optimal exosome concentration for transfection, exosomes at four different concentrations (0.25–4 µg/µl) were mixed with an Alexa Fluor 488-tagged siRNA (2 nmol/ml) and electroporated. Samples containing exosomes and siRNA, without an electric pulse, were used as a negative control. To eliminate the excess siRNA, the exosomes were purified using latex beads and the presence of siRNA in the exosomes was determined by FACS analysis.

    Article Snippet: To examine the efficiency of chemical transfection, a siRNA against MAPK1 was mixed with transfection reagent to allow for the formation of transfection complexes (siRNA embedded in lipid micelles), according to the manufacturer’s instructions (RNAi Human/Mouse Starter Kit, Quiagen).

    Techniques: Concentration Assay, Electroporation, Transfection, Negative Control, Purification, FACS

    Flow cytometry and northern slot blot of exosomes following chemical transfection. siRNA was embedded into lipid micelles according to the manufacturer’s instructions. The embedded siRNA was mixed with exosomes and incubated overnight. To eliminate the excess siRNA embedded in transfection complexes, the exosomes were purified using latex beads. The beads were washed twice with PBS to eliminate the excess of un-specific bound siRNA molecules. The presence of fluorescently labeled siRNA was determined by flow cytometry and northern blotting (slot blot). Panel A–D shows the results of flow cytometry for the detection of Alexa Fluor-488 labeled siRNA, beads only ( A ), beads plus siRNA ( B ), beads plus siRNA embedded in lipid micelles (negative control; C ) and chemically transfected exosomes ( D ). Panels E and F show the results of northern blotting using a digoxigenin-labeled probe against the administered MAPK1 siRNA. Panel E shows the presence of siRNA in exosomes and micelles. The chemically transfected exosomes were mixed with their parental cells HTB-177 and the delivery of siRNA from the exosomes to cells was determined and compared with direct transfection of cells ( F ). Non-transfected exosomes and cells were used as negative control.

    Journal: Nucleic Acids Research

    Article Title: Plasma exosomes can deliver exogenous short interfering RNA to monocytes and lymphocytes

    doi: 10.1093/nar/gks463

    Figure Lengend Snippet: Flow cytometry and northern slot blot of exosomes following chemical transfection. siRNA was embedded into lipid micelles according to the manufacturer’s instructions. The embedded siRNA was mixed with exosomes and incubated overnight. To eliminate the excess siRNA embedded in transfection complexes, the exosomes were purified using latex beads. The beads were washed twice with PBS to eliminate the excess of un-specific bound siRNA molecules. The presence of fluorescently labeled siRNA was determined by flow cytometry and northern blotting (slot blot). Panel A–D shows the results of flow cytometry for the detection of Alexa Fluor-488 labeled siRNA, beads only ( A ), beads plus siRNA ( B ), beads plus siRNA embedded in lipid micelles (negative control; C ) and chemically transfected exosomes ( D ). Panels E and F show the results of northern blotting using a digoxigenin-labeled probe against the administered MAPK1 siRNA. Panel E shows the presence of siRNA in exosomes and micelles. The chemically transfected exosomes were mixed with their parental cells HTB-177 and the delivery of siRNA from the exosomes to cells was determined and compared with direct transfection of cells ( F ). Non-transfected exosomes and cells were used as negative control.

    Article Snippet: To examine the efficiency of chemical transfection, a siRNA against MAPK1 was mixed with transfection reagent to allow for the formation of transfection complexes (siRNA embedded in lipid micelles), according to the manufacturer’s instructions (RNAi Human/Mouse Starter Kit, Quiagen).

    Techniques: Flow Cytometry, Cytometry, Northern Blot, Dot Blot, Transfection, Incubation, Purification, Labeling, Negative Control

    Transfection and Transduction Efficiency

    Journal: Laboratory investigation; a journal of technical methods and pathology

    Article Title: Immortalized Liver Endothelial Cells: A Cell Culture Model for Studies of Motility and Angiogenesis

    doi: 10.1038/labinvest.2010.132

    Figure Lengend Snippet: Transfection and Transduction Efficiency

    Article Snippet: Standard lipid-based transfection of plasmid DNA was performed using Effectene reagent (Qiagen) per the manufacturer's specifications.

    Techniques: Transfection, Transduction

    Comparative transfection efficiency of FuGENE HD (FuGENE), X-tremeGENE (X-treme), SuperFect (Super), Lipofectamine 2000 (L-2000), Lipofectamine RNAiMAX (L-iMAX), INT, and mTat/PEI/INT using siRNA targeting β - actin in HSC-3 cells. β - actin mRNA was measured by QRT-PCR and then % remaining β - actin mRNA expression was calculated based on control as 100% ( a ). Cell viability of each of the reagent groups in HaCaT cells were evaluated by MTT assay under the same siRNA and transfection reagent volume ( b ). * p

    Journal: Journal of Nanobiotechnology

    Article Title: Efficient siRNA delivery and gene silencing using a lipopolypeptide hybrid vector mediated by a caveolae-mediated and temperature-dependent endocytic pathway

    doi: 10.1186/s12951-019-0444-8

    Figure Lengend Snippet: Comparative transfection efficiency of FuGENE HD (FuGENE), X-tremeGENE (X-treme), SuperFect (Super), Lipofectamine 2000 (L-2000), Lipofectamine RNAiMAX (L-iMAX), INT, and mTat/PEI/INT using siRNA targeting β - actin in HSC-3 cells. β - actin mRNA was measured by QRT-PCR and then % remaining β - actin mRNA expression was calculated based on control as 100% ( a ). Cell viability of each of the reagent groups in HaCaT cells were evaluated by MTT assay under the same siRNA and transfection reagent volume ( b ). * p

    Article Snippet: Non-viral vectors To compare transfection efficiency of siRNA targeting β -actin (Santa Cruz Biotechnology, Inc. Dallas, TX) with several commercial reagents, Lipofectamine 2000 (Invitrogen, Carlsbad, CA), which is a cationic lipid-based transfection reagent, FuGENE HD (Promega Corp., Fitchburg, WI), which is a blend of lipids and other components, SuperFect (Qiagen, Valencia, CA), which is an activated dendrimer-based reagent, Lipofectamine RNAiMAX (Invitrogen), which is a cationic lipid-based transfection reagent, X-tremeGENE (Roche, Switzerland), which is a blend of lipids and other components, and INT (Polyplus-transfection, France) were used.

    Techniques: Transfection, Quantitative RT-PCR, Expressing, MTT Assay

    Mutational disruption of NK1R-mediated sequestration of β-arrestin prevents inhibition of F-MOR endocytosis in N2A cells (A) Confocal micrographs of N2A cells co-expressing either full-length HA-NK1R (left panels, red) or truncated HA-NK1R (right panels, red) with β-arrestin2-EGFP (green). Incubation with 10μM SP in cells expressing the truncated HA-NK1R (355x) resulted in less co-localization with receptors in endosomal clusters and higher cytoplasmic β-arrestin2-EGFP distribution (left column), compared to β-arrestin2-EGFP distribution in the wild type HA-NK1R under the same conditions (right column). Scale bar, 10μm. (B) In N2A cells co-expressing F-MOR (red) and HA-NK1 355x receptors (green), both morphine and DAMGO were able to drive internalization of F-MORs even in the presence of substance P. Scale bar, 10μm. (C) Quantification using ratiometric staining of F-MORs in N2A cells co-expressing HA-NK1 355x revealed that co-incubation with SP produced comparable levels of F-MOR internalization compared to treatment with either MS or DG alone. Bar graphs represent mean internalization determined from ~50 cell bodies selected at random in the N2A cultures and averaged over four independent transfections. (D) Biochemical assay of F-MOR internalization by surface biotinylation confirmed that F-MOR internalization induced by either MS or DG (lanes 3 and 5) was not significantly inhibited by co-application of SP (lanes 4 and 6; n= 3 independent experiments).

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    Article Title: Neurokinin1 receptors regulate morphine-induced endocytosis and desensitization of mu opioid receptors in CNS neurons

    doi: 10.1523/JNEUROSCI.4315-08.2009

    Figure Lengend Snippet: Mutational disruption of NK1R-mediated sequestration of β-arrestin prevents inhibition of F-MOR endocytosis in N2A cells (A) Confocal micrographs of N2A cells co-expressing either full-length HA-NK1R (left panels, red) or truncated HA-NK1R (right panels, red) with β-arrestin2-EGFP (green). Incubation with 10μM SP in cells expressing the truncated HA-NK1R (355x) resulted in less co-localization with receptors in endosomal clusters and higher cytoplasmic β-arrestin2-EGFP distribution (left column), compared to β-arrestin2-EGFP distribution in the wild type HA-NK1R under the same conditions (right column). Scale bar, 10μm. (B) In N2A cells co-expressing F-MOR (red) and HA-NK1 355x receptors (green), both morphine and DAMGO were able to drive internalization of F-MORs even in the presence of substance P. Scale bar, 10μm. (C) Quantification using ratiometric staining of F-MORs in N2A cells co-expressing HA-NK1 355x revealed that co-incubation with SP produced comparable levels of F-MOR internalization compared to treatment with either MS or DG alone. Bar graphs represent mean internalization determined from ~50 cell bodies selected at random in the N2A cultures and averaged over four independent transfections. (D) Biochemical assay of F-MOR internalization by surface biotinylation confirmed that F-MOR internalization induced by either MS or DG (lanes 3 and 5) was not significantly inhibited by co-application of SP (lanes 4 and 6; n= 3 independent experiments).

    Article Snippet: Transfections were performed using a cationic lipid transfection reagent (Effectene; Qiagen, Hilden, Germany).

    Techniques: Inhibition, Expressing, Incubation, Staining, Produced, Mass Spectrometry, Transfection

    Heterologous regulation of F-MOR endocytosis in mouse neuroblastoma (N2A) cells (A) Dual-labeled confocal fluorescence micrographs show F-MOR (red) and HA-NK1R (green) transiently expressed in N2A cells. In untreated cells, both F-MOR and HA-NK1R showed a plasma membrane distribution, with minimal labeled receptors present internally in the cell (top row). Incubation with either 10μM MS or 10μM SP for 30 minutes resulted in a selective increase in either labeled F-MORs (second row, first panel) or HA-NK1Rs (third row, middle panel), respectively, present internally in the cell. Simultaneous incubation with MS and SP produced visibly less F-MOR internalization compared to incubation with MS alone (bottom row, first panel). Scale bar, 10μm. (B) Quantification of ratiometric staining in N2A cells show inhibition of F-MOR internalization in response to both MS and DG when SP is also present. Bar graphs represent mean internalization determined from ~50 cell bodies selected at random in N2A cultures and averaged over five independent transfections (**p

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    Article Title: Neurokinin1 receptors regulate morphine-induced endocytosis and desensitization of mu opioid receptors in CNS neurons

    doi: 10.1523/JNEUROSCI.4315-08.2009

    Figure Lengend Snippet: Heterologous regulation of F-MOR endocytosis in mouse neuroblastoma (N2A) cells (A) Dual-labeled confocal fluorescence micrographs show F-MOR (red) and HA-NK1R (green) transiently expressed in N2A cells. In untreated cells, both F-MOR and HA-NK1R showed a plasma membrane distribution, with minimal labeled receptors present internally in the cell (top row). Incubation with either 10μM MS or 10μM SP for 30 minutes resulted in a selective increase in either labeled F-MORs (second row, first panel) or HA-NK1Rs (third row, middle panel), respectively, present internally in the cell. Simultaneous incubation with MS and SP produced visibly less F-MOR internalization compared to incubation with MS alone (bottom row, first panel). Scale bar, 10μm. (B) Quantification of ratiometric staining in N2A cells show inhibition of F-MOR internalization in response to both MS and DG when SP is also present. Bar graphs represent mean internalization determined from ~50 cell bodies selected at random in N2A cultures and averaged over five independent transfections (**p

    Article Snippet: Transfections were performed using a cationic lipid transfection reagent (Effectene; Qiagen, Hilden, Germany).

    Techniques: Labeling, Fluorescence, Incubation, Mass Spectrometry, Produced, Staining, Inhibition, Transfection