hek 293 a human embryonic kidney cell line  (ATCC)


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    ATCC hek 293 a human embryonic kidney cell line
    Phosphorylation-independent MAb production and epitope mapping. (A) Reactivity of MAbs with TDP-43-Flag recombinant protein of about 46 kDa (lane 1) overexpressed in <t>HEK</t> <t>293</t> cells by Western blot. Anti-Flag MAb (Flag) was used as the positive control antibody. Leucine-rich repeat-containing protein 15 (LRRC15)-His fusion protein (lane 2) served as the negative control, which was revealed by anti-His MAb (HIS). (B) Schematic representation of the TDP-43 fragments employed for epitope mapping. (C) Mapping of TDP-43 domains with epitopes recognized by the 10 generated MAbs. This mapping was performed across the entire length of TDP-43 using immunoblot and indirect peptide ELISA. RRM, RNA-recognition motif; NLS, bipartite nuclear localization signal; NES, nuclear export signal.
    Hek 293 A Human Embryonic Kidney Cell Line, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Production and characterization of novel monoclonal antibodies against pathological human TDP-43 proteins"

    Article Title: Production and characterization of novel monoclonal antibodies against pathological human TDP-43 proteins

    Journal: Journal of Neuropathology and Experimental Neurology

    doi: 10.1093/jnen/nlae042

    Phosphorylation-independent MAb production and epitope mapping. (A) Reactivity of MAbs with TDP-43-Flag recombinant protein of about 46 kDa (lane 1) overexpressed in HEK 293 cells by Western blot. Anti-Flag MAb (Flag) was used as the positive control antibody. Leucine-rich repeat-containing protein 15 (LRRC15)-His fusion protein (lane 2) served as the negative control, which was revealed by anti-His MAb (HIS). (B) Schematic representation of the TDP-43 fragments employed for epitope mapping. (C) Mapping of TDP-43 domains with epitopes recognized by the 10 generated MAbs. This mapping was performed across the entire length of TDP-43 using immunoblot and indirect peptide ELISA. RRM, RNA-recognition motif; NLS, bipartite nuclear localization signal; NES, nuclear export signal.
    Figure Legend Snippet: Phosphorylation-independent MAb production and epitope mapping. (A) Reactivity of MAbs with TDP-43-Flag recombinant protein of about 46 kDa (lane 1) overexpressed in HEK 293 cells by Western blot. Anti-Flag MAb (Flag) was used as the positive control antibody. Leucine-rich repeat-containing protein 15 (LRRC15)-His fusion protein (lane 2) served as the negative control, which was revealed by anti-His MAb (HIS). (B) Schematic representation of the TDP-43 fragments employed for epitope mapping. (C) Mapping of TDP-43 domains with epitopes recognized by the 10 generated MAbs. This mapping was performed across the entire length of TDP-43 using immunoblot and indirect peptide ELISA. RRM, RNA-recognition motif; NLS, bipartite nuclear localization signal; NES, nuclear export signal.

    Techniques Used: Recombinant, Western Blot, Positive Control, Negative Control, Generated, Peptide ELISA

    flp in t rex human embryonic kidney 293 hek293 cell line  (Thermo Fisher)


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    Thermo Fisher flp in t rex human embryonic kidney 293 hek293 cell line
    A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from <t>HEK293,</t> GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.
    Flp In T Rex Human Embryonic Kidney 293 Hek293 Cell Line, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "GTPBP8 plays a role in mitoribosome formation in human mitochondria"

    Article Title: GTPBP8 plays a role in mitoribosome formation in human mitochondria

    Journal: Nature Communications

    doi: 10.1038/s41467-024-50011-x

    A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.
    Figure Legend Snippet: A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.

    Techniques Used: Western Blot, Knock-Out, SDS Page, Control, Cell Culture, Standard Deviation, Two Tailed Test, Mass Spectrometry, Staining

    A In vivo translation assay of GTPBP8 KO1 and GTPBP8 KO2 cell lines compared to HEK293. Cells were labeled with a [ 35 S]-methionine and cysteine mix upon cytosolic translation inhibition. Whole cell lysates were resolved by SDS-PAGE and the signal was later detected via autoradiography. Loading was determined using Coomassie staining ( n = 3 independent experiments). B Mitoribosome profiling analysis of GTPBP8 KO1 compared to HEK293. The ratio of the mitoribosome-protected fragments (RPFs) in GTPBP8 KO1 vs control for each mitochondrial transcript is represented on the y -axis. The data were determined via MitoRiboSeq and refer to a single experiment. C Quantitative real-time PCR of mt-mRNAs and mt-rRNAs steady-state levels in HEK293 cells depleted of GTPBP8 compared to control. Total RNA isolated from HEK293, GTPBP8 KO1 and GTPBP8 KO2 was used ( n = 3 biological replicates, data presented as mean values ± 1 SD). Student’s two-tailed t -test was performed. P -values and source data are provided in a Source Data file. The significance cut-off was set at p < 0.05, with * p < 0.05, ** p < 0.01, and *** p < 0.001. Source data are provided as a Source Data file.
    Figure Legend Snippet: A In vivo translation assay of GTPBP8 KO1 and GTPBP8 KO2 cell lines compared to HEK293. Cells were labeled with a [ 35 S]-methionine and cysteine mix upon cytosolic translation inhibition. Whole cell lysates were resolved by SDS-PAGE and the signal was later detected via autoradiography. Loading was determined using Coomassie staining ( n = 3 independent experiments). B Mitoribosome profiling analysis of GTPBP8 KO1 compared to HEK293. The ratio of the mitoribosome-protected fragments (RPFs) in GTPBP8 KO1 vs control for each mitochondrial transcript is represented on the y -axis. The data were determined via MitoRiboSeq and refer to a single experiment. C Quantitative real-time PCR of mt-mRNAs and mt-rRNAs steady-state levels in HEK293 cells depleted of GTPBP8 compared to control. Total RNA isolated from HEK293, GTPBP8 KO1 and GTPBP8 KO2 was used ( n = 3 biological replicates, data presented as mean values ± 1 SD). Student’s two-tailed t -test was performed. P -values and source data are provided in a Source Data file. The significance cut-off was set at p < 0.05, with * p < 0.05, ** p < 0.01, and *** p < 0.001. Source data are provided as a Source Data file.

    Techniques Used: In Vivo, Labeling, Inhibition, SDS Page, Autoradiography, Staining, Control, Real-time Polymerase Chain Reaction, Isolation, Two Tailed Test

    A Western blotting analyses of the mt-LSU and mt-SSU MRPs and selected assembly factors steady-state levels in GTPBP8-depleted cells. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE and membranes were blotted with antibodies against GTPBP8, mt-LSU MRPs (mL37, uL3m, bL28m, mL49, uL12m, uL15m), mt-SSU MRPs (uS15m, uS16m, uS17m, mS22, mS37) and others (RBFA, GTPBP10, MRM3, TRMT10C, MTG1). SDHA was used as a loading control. B Quantification of MRPs steady-state levels in GTPBP8 knock-out cells compared to control HEK293. Quantification from western blotting ( n = 3 biological replicates) was performed using the Image Lab software and protein signals were normalized to SDHA. Student’s two-tailed t -test was performed. Source data and P -values are provided in a Source Data file. C LFQ mass spectrometry analyses of MitoCarta3.0 mito-pathways in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis. Each dot refers to a protein belonging to mt-LSU MRPs, mt-LSU assembly factors, mt-SSU MRPs, mt-SSU assembly factors, mt-RNA processing proteins, mt-transcription and mt-translation pathways. A summary of boxplots is provided in a Supplementary Data File. The complete list of log 2 (FC) values is presented in Supplementary Data . D Sucrose gradient centrifugation analysis to assess mitoribosome sedimentation patterns in GTPBP8 KO1 and GTPBP8 KO2 compared to HEK293 control cells. Mitochondrial lysates were loaded onto 10–30 % isokinetic sucrose gradients and obtained fractions were analyzed via western blotting. Membranes were probed for mt-LSU MRP mL37 and mt-SSU MRP uS15m. Below, quantification using Image Lab software of mL37 and uS15m in the monosome fractions from independent replicates is represented ( n = 3 independent experiments). The samples derived from the same experiment were processed in parallel. The y -axis represents the signal detected in the monosome fraction normalized for the total signal of each sample’s gradient. Data are presented as mean values +/− SD. Student’s two-tailed t -test was performed. P -values and source data are provided as a Source Data file.
    Figure Legend Snippet: A Western blotting analyses of the mt-LSU and mt-SSU MRPs and selected assembly factors steady-state levels in GTPBP8-depleted cells. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE and membranes were blotted with antibodies against GTPBP8, mt-LSU MRPs (mL37, uL3m, bL28m, mL49, uL12m, uL15m), mt-SSU MRPs (uS15m, uS16m, uS17m, mS22, mS37) and others (RBFA, GTPBP10, MRM3, TRMT10C, MTG1). SDHA was used as a loading control. B Quantification of MRPs steady-state levels in GTPBP8 knock-out cells compared to control HEK293. Quantification from western blotting ( n = 3 biological replicates) was performed using the Image Lab software and protein signals were normalized to SDHA. Student’s two-tailed t -test was performed. Source data and P -values are provided in a Source Data file. C LFQ mass spectrometry analyses of MitoCarta3.0 mito-pathways in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis. Each dot refers to a protein belonging to mt-LSU MRPs, mt-LSU assembly factors, mt-SSU MRPs, mt-SSU assembly factors, mt-RNA processing proteins, mt-transcription and mt-translation pathways. A summary of boxplots is provided in a Supplementary Data File. The complete list of log 2 (FC) values is presented in Supplementary Data . D Sucrose gradient centrifugation analysis to assess mitoribosome sedimentation patterns in GTPBP8 KO1 and GTPBP8 KO2 compared to HEK293 control cells. Mitochondrial lysates were loaded onto 10–30 % isokinetic sucrose gradients and obtained fractions were analyzed via western blotting. Membranes were probed for mt-LSU MRP mL37 and mt-SSU MRP uS15m. Below, quantification using Image Lab software of mL37 and uS15m in the monosome fractions from independent replicates is represented ( n = 3 independent experiments). The samples derived from the same experiment were processed in parallel. The y -axis represents the signal detected in the monosome fraction normalized for the total signal of each sample’s gradient. Data are presented as mean values +/− SD. Student’s two-tailed t -test was performed. P -values and source data are provided as a Source Data file.

    Techniques Used: Western Blot, SDS Page, Control, Knock-Out, Software, Two Tailed Test, Mass Spectrometry, Gradient Centrifugation, Sedimentation, Derivative Assay

    A Western blotting analysis of control HEK293, GTPBP8 KO1 , GTPBP8 KO2 , GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A samples to confirm GTPBP8 knock-out compared to the rescue cell lines. Anti-GTPBP8 antibody was used to compare GTPBP8 endogenous expression with the overexpression in rescue cell lines. SDHA was used as a loading control ( n = 1 independent experiments). B Western blotting analysis to test OxPhos steady-state levels in GTPBP8 KO1 and GTPBP8 KO2 compared to wild-type HEK293 and rescue cell lines GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Overexpression was induced prior to the experiment using 1 ng/ml and 50 ng/ml of doxycycline. Whole-cell lysates were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. SDHA and SDHB were used as loading controls ( n = 1). C Sucrose gradient centrifugation analysis to assess mitoribosome sedimentation patterns in GTPBP8 KO1 and GTPBP8 KO2 compared to HEK293, GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Mitochondrial lysates were loaded onto 10-30 % isokinetic sucrose gradients and obtained fractions were analyzed via western blotting. Membranes were probed for mt-LSU MRP, mL37, and mt-SSU MRP, uS15m ( n = 3 independent experiments). D In vivo translation assay of GTPBP8 KO1 and GTPBP8 KO2 cell lines compared to HEK293, GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Cells were labeled with a [ 35 S]-methionine and cysteine mix upon cytosolic translation inhibition. Whole cell lysates were resolved by SDS-PAGE and the signal was later detected via autoradiography. The loading was determined using Coomassie staining ( n = 1 independent experiment). Source data are provided as a Source Data file.
    Figure Legend Snippet: A Western blotting analysis of control HEK293, GTPBP8 KO1 , GTPBP8 KO2 , GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A samples to confirm GTPBP8 knock-out compared to the rescue cell lines. Anti-GTPBP8 antibody was used to compare GTPBP8 endogenous expression with the overexpression in rescue cell lines. SDHA was used as a loading control ( n = 1 independent experiments). B Western blotting analysis to test OxPhos steady-state levels in GTPBP8 KO1 and GTPBP8 KO2 compared to wild-type HEK293 and rescue cell lines GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Overexpression was induced prior to the experiment using 1 ng/ml and 50 ng/ml of doxycycline. Whole-cell lysates were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. SDHA and SDHB were used as loading controls ( n = 1). C Sucrose gradient centrifugation analysis to assess mitoribosome sedimentation patterns in GTPBP8 KO1 and GTPBP8 KO2 compared to HEK293, GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Mitochondrial lysates were loaded onto 10-30 % isokinetic sucrose gradients and obtained fractions were analyzed via western blotting. Membranes were probed for mt-LSU MRP, mL37, and mt-SSU MRP, uS15m ( n = 3 independent experiments). D In vivo translation assay of GTPBP8 KO1 and GTPBP8 KO2 cell lines compared to HEK293, GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Cells were labeled with a [ 35 S]-methionine and cysteine mix upon cytosolic translation inhibition. Whole cell lysates were resolved by SDS-PAGE and the signal was later detected via autoradiography. The loading was determined using Coomassie staining ( n = 1 independent experiment). Source data are provided as a Source Data file.

    Techniques Used: Western Blot, Control, Knock-Out, Expressing, Over Expression, SDS Page, Gradient Centrifugation, Sedimentation, In Vivo, Labeling, Inhibition, Autoradiography, Staining

    human embryonic kidney cell line hek 293  (ATCC)


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    ATCC human embryonic kidney cell line hek 293
    Human Embryonic Kidney Cell Line Hek 293, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    human embryonic kidney cell line hek 293  (ATCC)


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    ATCC human embryonic kidney cell line hek 293
    Human Embryonic Kidney Cell Line Hek 293, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    hek293 cells human embryonic kidney 293 cell line  (Millipore)

     
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    Millipore hek293 cells human embryonic kidney 293 cell line
    Hek293 Cells Human Embryonic Kidney 293 Cell Line, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    human embryonic kidney cell line hek 293  (ATCC)


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    ATCC human embryonic kidney cell line hek 293
    The effect of statins on viability and growth of ( a ) stem ADMSC and non-cancerous <t>HEK</t> <t>293</t> cells and ( b ) cancer MiaPaCa-2 cells. ( a ) ADMSC—human adipose-derived mesenchymal stem cells, HEK 293—human embryonic kidney cells, exposure to statins—24 h, concentrations 0—100 µM, control—methanol, ( b ) previously published data , MiaPaCa-2—pancreatic cancer cells, exposure to statins—24 h, concentrations 0—40 µM, control—methanol.
    Human Embryonic Kidney Cell Line Hek 293, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Highly variable biological effects of statins on cancer, non-cancer, and stem cells in vitro"

    Article Title: Highly variable biological effects of statins on cancer, non-cancer, and stem cells in vitro

    Journal: Scientific Reports

    doi: 10.1038/s41598-024-62615-w

    The effect of statins on viability and growth of ( a ) stem ADMSC and non-cancerous HEK 293 cells and ( b ) cancer MiaPaCa-2 cells. ( a ) ADMSC—human adipose-derived mesenchymal stem cells, HEK 293—human embryonic kidney cells, exposure to statins—24 h, concentrations 0—100 µM, control—methanol, ( b ) previously published data , MiaPaCa-2—pancreatic cancer cells, exposure to statins—24 h, concentrations 0—40 µM, control—methanol.
    Figure Legend Snippet: The effect of statins on viability and growth of ( a ) stem ADMSC and non-cancerous HEK 293 cells and ( b ) cancer MiaPaCa-2 cells. ( a ) ADMSC—human adipose-derived mesenchymal stem cells, HEK 293—human embryonic kidney cells, exposure to statins—24 h, concentrations 0—100 µM, control—methanol, ( b ) previously published data , MiaPaCa-2—pancreatic cancer cells, exposure to statins—24 h, concentrations 0—40 µM, control—methanol.

    Techniques Used: Derivative Assay

    Comparison of the effect of statins on the growth and viability of pancreatic cancer MiaPaCa-2 cells, non-cancerous HEK 293 cells, and ADMSC stem cells. Concentration of statins—20 µM, Time—exposure to statins—24, 48, and 72 h, control—methanol.
    Figure Legend Snippet: Comparison of the effect of statins on the growth and viability of pancreatic cancer MiaPaCa-2 cells, non-cancerous HEK 293 cells, and ADMSC stem cells. Concentration of statins—20 µM, Time—exposure to statins—24, 48, and 72 h, control—methanol.

    Techniques Used: Comparison, Concentration Assay

    cell lines human embryonic kidney hek 293 atcc cat crl 1573  (ATCC)


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    ATCC cell lines human embryonic kidney hek 293 atcc cat crl 1573
    Cell Lines Human Embryonic Kidney Hek 293 Atcc Cat Crl 1573, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    human embryonic kidney 293 hek293 cell lines  (ATCC)


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    ATCC human embryonic kidney 293 hek293 cell lines
    Human Embryonic Kidney 293 Hek293 Cell Lines, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    human embryonic kidney epithelial ‒ hek 293 cell lines  (ATCC)


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    ATCC human embryonic kidney epithelial ‒ hek 293 cell lines
    Relative ACE2 ( A ) and TMPRSS2( B ) mRNA levels in <t>HEK-293-hACE2</t> cells at different time points after transfection with LNP-si ACE2 (40 nM si ACE2 ) and LNP-si TMPRSS2 (40 nM si TMPRSS2 ), respectively. The control cells in ( A ) and ( B ) received LNP-siSCR. Relative ACE2 mRNA level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( C ). Relative ACE2 protein level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( D ). The uncropped blots are shown in . Values are expressed as the mean ± standard error of the mean (n = 3). Statistical significance between the control and treatment groups were analyzed using t -test (**p < 0.01, ***p<0.001, ****p<0.0001; p < 0.05 was considered statistically significant).
    Human Embryonic Kidney Epithelial ‒ Hek 293 Cell Lines, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Lipid Nanoparticle-Based Inhibitors for SARS-CoV-2 Host Cell Infection"

    Article Title: Lipid Nanoparticle-Based Inhibitors for SARS-CoV-2 Host Cell Infection

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S448005

    Relative ACE2 ( A ) and TMPRSS2( B ) mRNA levels in HEK-293-hACE2 cells at different time points after transfection with LNP-si ACE2 (40 nM si ACE2 ) and LNP-si TMPRSS2 (40 nM si TMPRSS2 ), respectively. The control cells in ( A ) and ( B ) received LNP-siSCR. Relative ACE2 mRNA level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( C ). Relative ACE2 protein level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( D ). The uncropped blots are shown in . Values are expressed as the mean ± standard error of the mean (n = 3). Statistical significance between the control and treatment groups were analyzed using t -test (**p < 0.01, ***p<0.001, ****p<0.0001; p < 0.05 was considered statistically significant).
    Figure Legend Snippet: Relative ACE2 ( A ) and TMPRSS2( B ) mRNA levels in HEK-293-hACE2 cells at different time points after transfection with LNP-si ACE2 (40 nM si ACE2 ) and LNP-si TMPRSS2 (40 nM si TMPRSS2 ), respectively. The control cells in ( A ) and ( B ) received LNP-siSCR. Relative ACE2 mRNA level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( C ). Relative ACE2 protein level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( D ). The uncropped blots are shown in . Values are expressed as the mean ± standard error of the mean (n = 3). Statistical significance between the control and treatment groups were analyzed using t -test (**p < 0.01, ***p<0.001, ****p<0.0001; p < 0.05 was considered statistically significant).

    Techniques Used: Transfection

    Transduction efficiency (luciferase expression) of SARS-CoV-2 S pseudotyped luciferase lentivirus at the given MOIs in HEK-293 and HEK-293-hACE2 cell lines ( A ). Percentage inhibition of infection in ACE2 KD and TMPRSS2 KD HEK-293-hACE2 cells ( B ). Percentage inhibition of infection in ACE2 KD ( C ) and TMPRSS2 KD ( D ) in airway epithelial Calu-3 cell line at different siRNA concentrations (LNP-siRNA: 1, 2, 10, 20 and 40 nM; presented in the graph as log 10 values). The nonlinear regression curve fit analysis of siRNA concentration vs inhibition is presented in the graphs. Values are expressed as the mean ± standard error of the mean (n =3). Statistical analysis was performed using one-way ANOVA, followed by Tukey’s test (*p < 0.05, **p<0.01; p < 0.05 was considered statistically significant).
    Figure Legend Snippet: Transduction efficiency (luciferase expression) of SARS-CoV-2 S pseudotyped luciferase lentivirus at the given MOIs in HEK-293 and HEK-293-hACE2 cell lines ( A ). Percentage inhibition of infection in ACE2 KD and TMPRSS2 KD HEK-293-hACE2 cells ( B ). Percentage inhibition of infection in ACE2 KD ( C ) and TMPRSS2 KD ( D ) in airway epithelial Calu-3 cell line at different siRNA concentrations (LNP-siRNA: 1, 2, 10, 20 and 40 nM; presented in the graph as log 10 values). The nonlinear regression curve fit analysis of siRNA concentration vs inhibition is presented in the graphs. Values are expressed as the mean ± standard error of the mean (n =3). Statistical analysis was performed using one-way ANOVA, followed by Tukey’s test (*p < 0.05, **p<0.01; p < 0.05 was considered statistically significant).

    Techniques Used: Transduction, Luciferase, Expressing, Inhibition, Infection, Concentration Assay


    Structured Review

    iCell Gene Therapeutics human embryonic kidney cell line hek 293 t
    Human Embryonic Kidney Cell Line Hek 293 T, supplied by iCell Gene Therapeutics, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC hek 293 a human embryonic kidney cell line
    Phosphorylation-independent MAb production and epitope mapping. (A) Reactivity of MAbs with TDP-43-Flag recombinant protein of about 46 kDa (lane 1) overexpressed in <t>HEK</t> <t>293</t> cells by Western blot. Anti-Flag MAb (Flag) was used as the positive control antibody. Leucine-rich repeat-containing protein 15 (LRRC15)-His fusion protein (lane 2) served as the negative control, which was revealed by anti-His MAb (HIS). (B) Schematic representation of the TDP-43 fragments employed for epitope mapping. (C) Mapping of TDP-43 domains with epitopes recognized by the 10 generated MAbs. This mapping was performed across the entire length of TDP-43 using immunoblot and indirect peptide ELISA. RRM, RNA-recognition motif; NLS, bipartite nuclear localization signal; NES, nuclear export signal.
    Hek 293 A Human Embryonic Kidney Cell Line, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher flp in t rex human embryonic kidney 293 hek293 cell line
    A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from <t>HEK293,</t> GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.
    Flp In T Rex Human Embryonic Kidney 293 Hek293 Cell Line, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC human embryonic kidney cell line hek 293
    A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from <t>HEK293,</t> GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.
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    Millipore hek293 cells human embryonic kidney 293 cell line
    A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from <t>HEK293,</t> GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.
    Hek293 Cells Human Embryonic Kidney 293 Cell Line, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC cell lines human embryonic kidney hek 293 atcc cat crl 1573
    A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from <t>HEK293,</t> GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.
    Cell Lines Human Embryonic Kidney Hek 293 Atcc Cat Crl 1573, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC human embryonic kidney 293 hek293 cell lines
    A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from <t>HEK293,</t> GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.
    Human Embryonic Kidney 293 Hek293 Cell Lines, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC human embryonic kidney epithelial ‒ hek 293 cell lines
    Relative ACE2 ( A ) and TMPRSS2( B ) mRNA levels in <t>HEK-293-hACE2</t> cells at different time points after transfection with LNP-si ACE2 (40 nM si ACE2 ) and LNP-si TMPRSS2 (40 nM si TMPRSS2 ), respectively. The control cells in ( A ) and ( B ) received LNP-siSCR. Relative ACE2 mRNA level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( C ). Relative ACE2 protein level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( D ). The uncropped blots are shown in . Values are expressed as the mean ± standard error of the mean (n = 3). Statistical significance between the control and treatment groups were analyzed using t -test (**p < 0.01, ***p<0.001, ****p<0.0001; p < 0.05 was considered statistically significant).
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    iCell Gene Therapeutics human embryonic kidney cell line hek 293 t
    Relative ACE2 ( A ) and TMPRSS2( B ) mRNA levels in <t>HEK-293-hACE2</t> cells at different time points after transfection with LNP-si ACE2 (40 nM si ACE2 ) and LNP-si TMPRSS2 (40 nM si TMPRSS2 ), respectively. The control cells in ( A ) and ( B ) received LNP-siSCR. Relative ACE2 mRNA level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( C ). Relative ACE2 protein level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( D ). The uncropped blots are shown in . Values are expressed as the mean ± standard error of the mean (n = 3). Statistical significance between the control and treatment groups were analyzed using t -test (**p < 0.01, ***p<0.001, ****p<0.0001; p < 0.05 was considered statistically significant).
    Human Embryonic Kidney Cell Line Hek 293 T, supplied by iCell Gene Therapeutics, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Phosphorylation-independent MAb production and epitope mapping. (A) Reactivity of MAbs with TDP-43-Flag recombinant protein of about 46 kDa (lane 1) overexpressed in HEK 293 cells by Western blot. Anti-Flag MAb (Flag) was used as the positive control antibody. Leucine-rich repeat-containing protein 15 (LRRC15)-His fusion protein (lane 2) served as the negative control, which was revealed by anti-His MAb (HIS). (B) Schematic representation of the TDP-43 fragments employed for epitope mapping. (C) Mapping of TDP-43 domains with epitopes recognized by the 10 generated MAbs. This mapping was performed across the entire length of TDP-43 using immunoblot and indirect peptide ELISA. RRM, RNA-recognition motif; NLS, bipartite nuclear localization signal; NES, nuclear export signal.

    Journal: Journal of Neuropathology and Experimental Neurology

    Article Title: Production and characterization of novel monoclonal antibodies against pathological human TDP-43 proteins

    doi: 10.1093/jnen/nlae042

    Figure Lengend Snippet: Phosphorylation-independent MAb production and epitope mapping. (A) Reactivity of MAbs with TDP-43-Flag recombinant protein of about 46 kDa (lane 1) overexpressed in HEK 293 cells by Western blot. Anti-Flag MAb (Flag) was used as the positive control antibody. Leucine-rich repeat-containing protein 15 (LRRC15)-His fusion protein (lane 2) served as the negative control, which was revealed by anti-His MAb (HIS). (B) Schematic representation of the TDP-43 fragments employed for epitope mapping. (C) Mapping of TDP-43 domains with epitopes recognized by the 10 generated MAbs. This mapping was performed across the entire length of TDP-43 using immunoblot and indirect peptide ELISA. RRM, RNA-recognition motif; NLS, bipartite nuclear localization signal; NES, nuclear export signal.

    Article Snippet: Two cell lines, SH-SY5Y (a human neuroblastoma cell line) and HEK 293 (a human embryonic kidney cell line), were sourced from the American Type Culture Collection (ATCC, Manassas, VA).

    Techniques: Recombinant, Western Blot, Positive Control, Negative Control, Generated, Peptide ELISA

    A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.

    Journal: Nature Communications

    Article Title: GTPBP8 plays a role in mitoribosome formation in human mitochondria

    doi: 10.1038/s41467-024-50011-x

    Figure Lengend Snippet: A Western blotting to confirm GTPBP8 knock-out. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were used for SDS-PAGE analysis. GTPBP8 antibody was used to detect GTPBP8 endogenous levels; SDHA antibody was used as a loading control. B Growth rates of GTPBP8 knock-out cells in galactose medium. HEK293 (black), GTPBP8 KO1 (red), and GTPBP8 KO2 (yellow) were cultured in DMEM containing 0.9 g/l galactose ( n = 3 independent experiments). The mean average cell number per each time point is indicated, and error bars = ± 1 standard deviation (SD). Statistical analyses were performed using the Student’s two-tailed t -test. P -values and source data are provided in the Source Data file. C Western blotting to assess OxPhos complexes steady-state levels in GTPBP8 knock-out cell lines. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. GAPDH was used as a loading control. D LFQ mass spectrometry analyses of OxPhos complexes in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis for each OxPhos complex. Each dot refers to a protein, which constitutes the specific complex. A summary of boxplots is provided in a Source Data file. All the log 2 (FC) values are listed in Supplementary Data . E Assessment of OxPhos complex I and complex IV activities in GTPBP8 knock-out cell lines compared to control. Mitochondrial lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 cell lines were resolved by BN-PAGE and stained using complex I (CI) and complex IV (CIV) -specific substrates ( n = 1 independent experiment). Coomassie staining was used as a loading control. SC supercomplexes. Source data are provided as a Source Data file.

    Article Snippet: Stable mammalian cell lines that enable doxycycline-inducible expression of C-terminally FLAG-tagged GTPBP8 (GTPBP8::FLAG) were generated using the Flp-In T-Rex human embryonic kidney 293 (HEK293) cell line (Invitrogen).

    Techniques: Western Blot, Knock-Out, SDS Page, Control, Cell Culture, Standard Deviation, Two Tailed Test, Mass Spectrometry, Staining

    A In vivo translation assay of GTPBP8 KO1 and GTPBP8 KO2 cell lines compared to HEK293. Cells were labeled with a [ 35 S]-methionine and cysteine mix upon cytosolic translation inhibition. Whole cell lysates were resolved by SDS-PAGE and the signal was later detected via autoradiography. Loading was determined using Coomassie staining ( n = 3 independent experiments). B Mitoribosome profiling analysis of GTPBP8 KO1 compared to HEK293. The ratio of the mitoribosome-protected fragments (RPFs) in GTPBP8 KO1 vs control for each mitochondrial transcript is represented on the y -axis. The data were determined via MitoRiboSeq and refer to a single experiment. C Quantitative real-time PCR of mt-mRNAs and mt-rRNAs steady-state levels in HEK293 cells depleted of GTPBP8 compared to control. Total RNA isolated from HEK293, GTPBP8 KO1 and GTPBP8 KO2 was used ( n = 3 biological replicates, data presented as mean values ± 1 SD). Student’s two-tailed t -test was performed. P -values and source data are provided in a Source Data file. The significance cut-off was set at p < 0.05, with * p < 0.05, ** p < 0.01, and *** p < 0.001. Source data are provided as a Source Data file.

    Journal: Nature Communications

    Article Title: GTPBP8 plays a role in mitoribosome formation in human mitochondria

    doi: 10.1038/s41467-024-50011-x

    Figure Lengend Snippet: A In vivo translation assay of GTPBP8 KO1 and GTPBP8 KO2 cell lines compared to HEK293. Cells were labeled with a [ 35 S]-methionine and cysteine mix upon cytosolic translation inhibition. Whole cell lysates were resolved by SDS-PAGE and the signal was later detected via autoradiography. Loading was determined using Coomassie staining ( n = 3 independent experiments). B Mitoribosome profiling analysis of GTPBP8 KO1 compared to HEK293. The ratio of the mitoribosome-protected fragments (RPFs) in GTPBP8 KO1 vs control for each mitochondrial transcript is represented on the y -axis. The data were determined via MitoRiboSeq and refer to a single experiment. C Quantitative real-time PCR of mt-mRNAs and mt-rRNAs steady-state levels in HEK293 cells depleted of GTPBP8 compared to control. Total RNA isolated from HEK293, GTPBP8 KO1 and GTPBP8 KO2 was used ( n = 3 biological replicates, data presented as mean values ± 1 SD). Student’s two-tailed t -test was performed. P -values and source data are provided in a Source Data file. The significance cut-off was set at p < 0.05, with * p < 0.05, ** p < 0.01, and *** p < 0.001. Source data are provided as a Source Data file.

    Article Snippet: Stable mammalian cell lines that enable doxycycline-inducible expression of C-terminally FLAG-tagged GTPBP8 (GTPBP8::FLAG) were generated using the Flp-In T-Rex human embryonic kidney 293 (HEK293) cell line (Invitrogen).

    Techniques: In Vivo, Labeling, Inhibition, SDS Page, Autoradiography, Staining, Control, Real-time Polymerase Chain Reaction, Isolation, Two Tailed Test

    A Western blotting analyses of the mt-LSU and mt-SSU MRPs and selected assembly factors steady-state levels in GTPBP8-depleted cells. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE and membranes were blotted with antibodies against GTPBP8, mt-LSU MRPs (mL37, uL3m, bL28m, mL49, uL12m, uL15m), mt-SSU MRPs (uS15m, uS16m, uS17m, mS22, mS37) and others (RBFA, GTPBP10, MRM3, TRMT10C, MTG1). SDHA was used as a loading control. B Quantification of MRPs steady-state levels in GTPBP8 knock-out cells compared to control HEK293. Quantification from western blotting ( n = 3 biological replicates) was performed using the Image Lab software and protein signals were normalized to SDHA. Student’s two-tailed t -test was performed. Source data and P -values are provided in a Source Data file. C LFQ mass spectrometry analyses of MitoCarta3.0 mito-pathways in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis. Each dot refers to a protein belonging to mt-LSU MRPs, mt-LSU assembly factors, mt-SSU MRPs, mt-SSU assembly factors, mt-RNA processing proteins, mt-transcription and mt-translation pathways. A summary of boxplots is provided in a Supplementary Data File. The complete list of log 2 (FC) values is presented in Supplementary Data . D Sucrose gradient centrifugation analysis to assess mitoribosome sedimentation patterns in GTPBP8 KO1 and GTPBP8 KO2 compared to HEK293 control cells. Mitochondrial lysates were loaded onto 10–30 % isokinetic sucrose gradients and obtained fractions were analyzed via western blotting. Membranes were probed for mt-LSU MRP mL37 and mt-SSU MRP uS15m. Below, quantification using Image Lab software of mL37 and uS15m in the monosome fractions from independent replicates is represented ( n = 3 independent experiments). The samples derived from the same experiment were processed in parallel. The y -axis represents the signal detected in the monosome fraction normalized for the total signal of each sample’s gradient. Data are presented as mean values +/− SD. Student’s two-tailed t -test was performed. P -values and source data are provided as a Source Data file.

    Journal: Nature Communications

    Article Title: GTPBP8 plays a role in mitoribosome formation in human mitochondria

    doi: 10.1038/s41467-024-50011-x

    Figure Lengend Snippet: A Western blotting analyses of the mt-LSU and mt-SSU MRPs and selected assembly factors steady-state levels in GTPBP8-depleted cells. Whole-cell lysates from HEK293, GTPBP8 KO1 and GTPBP8 KO2 were resolved via SDS-PAGE and membranes were blotted with antibodies against GTPBP8, mt-LSU MRPs (mL37, uL3m, bL28m, mL49, uL12m, uL15m), mt-SSU MRPs (uS15m, uS16m, uS17m, mS22, mS37) and others (RBFA, GTPBP10, MRM3, TRMT10C, MTG1). SDHA was used as a loading control. B Quantification of MRPs steady-state levels in GTPBP8 knock-out cells compared to control HEK293. Quantification from western blotting ( n = 3 biological replicates) was performed using the Image Lab software and protein signals were normalized to SDHA. Student’s two-tailed t -test was performed. Source data and P -values are provided in a Source Data file. C LFQ mass spectrometry analyses of MitoCarta3.0 mito-pathways in GTPBP8 knock-out cell lines compared to HEK293 ( n = 3 independent experiments). Log 2 (FC) of GTPBP8 KO1 vs HEK293 (red) and GTPBP8 KO2 vs HEK293 (yellow) is represented on the y -axis. Each dot refers to a protein belonging to mt-LSU MRPs, mt-LSU assembly factors, mt-SSU MRPs, mt-SSU assembly factors, mt-RNA processing proteins, mt-transcription and mt-translation pathways. A summary of boxplots is provided in a Supplementary Data File. The complete list of log 2 (FC) values is presented in Supplementary Data . D Sucrose gradient centrifugation analysis to assess mitoribosome sedimentation patterns in GTPBP8 KO1 and GTPBP8 KO2 compared to HEK293 control cells. Mitochondrial lysates were loaded onto 10–30 % isokinetic sucrose gradients and obtained fractions were analyzed via western blotting. Membranes were probed for mt-LSU MRP mL37 and mt-SSU MRP uS15m. Below, quantification using Image Lab software of mL37 and uS15m in the monosome fractions from independent replicates is represented ( n = 3 independent experiments). The samples derived from the same experiment were processed in parallel. The y -axis represents the signal detected in the monosome fraction normalized for the total signal of each sample’s gradient. Data are presented as mean values +/− SD. Student’s two-tailed t -test was performed. P -values and source data are provided as a Source Data file.

    Article Snippet: Stable mammalian cell lines that enable doxycycline-inducible expression of C-terminally FLAG-tagged GTPBP8 (GTPBP8::FLAG) were generated using the Flp-In T-Rex human embryonic kidney 293 (HEK293) cell line (Invitrogen).

    Techniques: Western Blot, SDS Page, Control, Knock-Out, Software, Two Tailed Test, Mass Spectrometry, Gradient Centrifugation, Sedimentation, Derivative Assay

    A Western blotting analysis of control HEK293, GTPBP8 KO1 , GTPBP8 KO2 , GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A samples to confirm GTPBP8 knock-out compared to the rescue cell lines. Anti-GTPBP8 antibody was used to compare GTPBP8 endogenous expression with the overexpression in rescue cell lines. SDHA was used as a loading control ( n = 1 independent experiments). B Western blotting analysis to test OxPhos steady-state levels in GTPBP8 KO1 and GTPBP8 KO2 compared to wild-type HEK293 and rescue cell lines GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Overexpression was induced prior to the experiment using 1 ng/ml and 50 ng/ml of doxycycline. Whole-cell lysates were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. SDHA and SDHB were used as loading controls ( n = 1). C Sucrose gradient centrifugation analysis to assess mitoribosome sedimentation patterns in GTPBP8 KO1 and GTPBP8 KO2 compared to HEK293, GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Mitochondrial lysates were loaded onto 10-30 % isokinetic sucrose gradients and obtained fractions were analyzed via western blotting. Membranes were probed for mt-LSU MRP, mL37, and mt-SSU MRP, uS15m ( n = 3 independent experiments). D In vivo translation assay of GTPBP8 KO1 and GTPBP8 KO2 cell lines compared to HEK293, GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Cells were labeled with a [ 35 S]-methionine and cysteine mix upon cytosolic translation inhibition. Whole cell lysates were resolved by SDS-PAGE and the signal was later detected via autoradiography. The loading was determined using Coomassie staining ( n = 1 independent experiment). Source data are provided as a Source Data file.

    Journal: Nature Communications

    Article Title: GTPBP8 plays a role in mitoribosome formation in human mitochondria

    doi: 10.1038/s41467-024-50011-x

    Figure Lengend Snippet: A Western blotting analysis of control HEK293, GTPBP8 KO1 , GTPBP8 KO2 , GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A samples to confirm GTPBP8 knock-out compared to the rescue cell lines. Anti-GTPBP8 antibody was used to compare GTPBP8 endogenous expression with the overexpression in rescue cell lines. SDHA was used as a loading control ( n = 1 independent experiments). B Western blotting analysis to test OxPhos steady-state levels in GTPBP8 KO1 and GTPBP8 KO2 compared to wild-type HEK293 and rescue cell lines GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Overexpression was induced prior to the experiment using 1 ng/ml and 50 ng/ml of doxycycline. Whole-cell lysates were resolved via SDS-PAGE. Membranes were probed using the OxPhos antibody cocktail against nuclear-encoded proteins of complexes V (ATP5A) and I (NDUFB8) and mtDNA-encoded complex IV protein MTCO2. SDHA and SDHB were used as loading controls ( n = 1). C Sucrose gradient centrifugation analysis to assess mitoribosome sedimentation patterns in GTPBP8 KO1 and GTPBP8 KO2 compared to HEK293, GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Mitochondrial lysates were loaded onto 10-30 % isokinetic sucrose gradients and obtained fractions were analyzed via western blotting. Membranes were probed for mt-LSU MRP, mL37, and mt-SSU MRP, uS15m ( n = 3 independent experiments). D In vivo translation assay of GTPBP8 KO1 and GTPBP8 KO2 cell lines compared to HEK293, GTPBP8 RESCUE_wt and GTPBP8 RESCUE_S124A . Cells were labeled with a [ 35 S]-methionine and cysteine mix upon cytosolic translation inhibition. Whole cell lysates were resolved by SDS-PAGE and the signal was later detected via autoradiography. The loading was determined using Coomassie staining ( n = 1 independent experiment). Source data are provided as a Source Data file.

    Article Snippet: Stable mammalian cell lines that enable doxycycline-inducible expression of C-terminally FLAG-tagged GTPBP8 (GTPBP8::FLAG) were generated using the Flp-In T-Rex human embryonic kidney 293 (HEK293) cell line (Invitrogen).

    Techniques: Western Blot, Control, Knock-Out, Expressing, Over Expression, SDS Page, Gradient Centrifugation, Sedimentation, In Vivo, Labeling, Inhibition, Autoradiography, Staining

    Relative ACE2 ( A ) and TMPRSS2( B ) mRNA levels in HEK-293-hACE2 cells at different time points after transfection with LNP-si ACE2 (40 nM si ACE2 ) and LNP-si TMPRSS2 (40 nM si TMPRSS2 ), respectively. The control cells in ( A ) and ( B ) received LNP-siSCR. Relative ACE2 mRNA level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( C ). Relative ACE2 protein level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( D ). The uncropped blots are shown in . Values are expressed as the mean ± standard error of the mean (n = 3). Statistical significance between the control and treatment groups were analyzed using t -test (**p < 0.01, ***p<0.001, ****p<0.0001; p < 0.05 was considered statistically significant).

    Journal: International Journal of Nanomedicine

    Article Title: Lipid Nanoparticle-Based Inhibitors for SARS-CoV-2 Host Cell Infection

    doi: 10.2147/IJN.S448005

    Figure Lengend Snippet: Relative ACE2 ( A ) and TMPRSS2( B ) mRNA levels in HEK-293-hACE2 cells at different time points after transfection with LNP-si ACE2 (40 nM si ACE2 ) and LNP-si TMPRSS2 (40 nM si TMPRSS2 ), respectively. The control cells in ( A ) and ( B ) received LNP-siSCR. Relative ACE2 mRNA level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( C ). Relative ACE2 protein level in Calu-3 cells 72 hours after receiving LNP-si ACE2 (40 nM si ACE2 ) against control cells that received LNP-siSCR ( D ). The uncropped blots are shown in . Values are expressed as the mean ± standard error of the mean (n = 3). Statistical significance between the control and treatment groups were analyzed using t -test (**p < 0.01, ***p<0.001, ****p<0.0001; p < 0.05 was considered statistically significant).

    Article Snippet: Human lung epithelial ‒ Calu-3 (CVCL_0609) and human embryonic kidney epithelial ‒ HEK-293 cell lines were obtained from ATCC (Manassas, VA).

    Techniques: Transfection

    Transduction efficiency (luciferase expression) of SARS-CoV-2 S pseudotyped luciferase lentivirus at the given MOIs in HEK-293 and HEK-293-hACE2 cell lines ( A ). Percentage inhibition of infection in ACE2 KD and TMPRSS2 KD HEK-293-hACE2 cells ( B ). Percentage inhibition of infection in ACE2 KD ( C ) and TMPRSS2 KD ( D ) in airway epithelial Calu-3 cell line at different siRNA concentrations (LNP-siRNA: 1, 2, 10, 20 and 40 nM; presented in the graph as log 10 values). The nonlinear regression curve fit analysis of siRNA concentration vs inhibition is presented in the graphs. Values are expressed as the mean ± standard error of the mean (n =3). Statistical analysis was performed using one-way ANOVA, followed by Tukey’s test (*p < 0.05, **p<0.01; p < 0.05 was considered statistically significant).

    Journal: International Journal of Nanomedicine

    Article Title: Lipid Nanoparticle-Based Inhibitors for SARS-CoV-2 Host Cell Infection

    doi: 10.2147/IJN.S448005

    Figure Lengend Snippet: Transduction efficiency (luciferase expression) of SARS-CoV-2 S pseudotyped luciferase lentivirus at the given MOIs in HEK-293 and HEK-293-hACE2 cell lines ( A ). Percentage inhibition of infection in ACE2 KD and TMPRSS2 KD HEK-293-hACE2 cells ( B ). Percentage inhibition of infection in ACE2 KD ( C ) and TMPRSS2 KD ( D ) in airway epithelial Calu-3 cell line at different siRNA concentrations (LNP-siRNA: 1, 2, 10, 20 and 40 nM; presented in the graph as log 10 values). The nonlinear regression curve fit analysis of siRNA concentration vs inhibition is presented in the graphs. Values are expressed as the mean ± standard error of the mean (n =3). Statistical analysis was performed using one-way ANOVA, followed by Tukey’s test (*p < 0.05, **p<0.01; p < 0.05 was considered statistically significant).

    Article Snippet: Human lung epithelial ‒ Calu-3 (CVCL_0609) and human embryonic kidney epithelial ‒ HEK-293 cell lines were obtained from ATCC (Manassas, VA).

    Techniques: Transduction, Luciferase, Expressing, Inhibition, Infection, Concentration Assay