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anti mouse secondary antibodies  (Jackson Immuno)


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    Structured Review

    Jackson Immuno anti mouse secondary antibodies
    Anti Mouse Secondary Antibodies, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 93/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/fitc+anti+mouse+secondary+antibody/Fluorescein+(FITC)+AffiniPure+F(ab')%E2%82%82+Fragment+Goat+Anti-Human+IgG/pm41912133-112-8-11
    Average 93 stars, based on 19 article reviews
    anti mouse secondary antibodies - by Bioz Stars, 2026-09
    93/100 stars

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    Related Articles

    Incubation:

    Article Title: Treadmill exercise ameliorates social isolation-induced depression through neuronal generation in rat pups.
    Article Snippet: The sections were then incubated overnight with rabbit polyclonal anti-5-HT antibody (1:500; Immuno Star, Hudson, WI, USA) and mouse monoclonal anti-TPH antibody (1:500; Oncogene Research Products, Cambridge, UK). .. The sections were next incubated for 90 min with CY3 anti-rabbit secondary antibody (Vector Laboratories, Burlingame, CA, USA) and FITC anti-mouse secondary antibody (Jackson ImmunoResearch Laboratories, West Grove, PA, USA). .. The sections were then mounted on gelatin-coated glass slides, http://www.e-jer.org 629https://doi.org/10.12965/jer.1735180.590 and the coverslips were mounted using fluorescent mounting medium (DakoCytomation, Carpinteria, CA, USA).

    Article Title: Endogenous interferon-gamma is required for efficient skeletal muscle regeneration.
    Article Snippet: For myoblast accumulation, sections were fixed in cold acetone, washed, blocked, and incubated with a MyoD antibody overnight (1:20, Santa Cruz Biotechnologies). .. Afterward, samples were washed with PBS and subsequently incubated with FITC anti-mouse secondary antibody (Jackson ImmunoResearch). ..

    Article Title: A Dual Protein-mRNA Localization Screen Reveals Compartmentalized Translation and Widespread Co-translational RNA Targeting.
    Article Snippet: For P-body labelling, coverslips were incubated overnight at 4 Cwith a monoclonal mouse antibody (Santa Cruz sc-8418; see Figure S2 of Stoecklin et al., 2006), diluted 1/250 in PBS containing 0.1% BSA. .. Coverslips were washed three times with PBS, 10 minutes each time, and incubated with an FITC anti-mouse secondary antibody (Jackson ImmunoResearch 115-096-006) diluted 1/200 in PBS with 0.1% BSA. ..

    Article Title: NeuN/Rbfox3 nuclear and cytoplasmic isoforms differentially regulate alternative splicing and nonsense-mediated decay of Rbfox2.
    Article Snippet: .. Cells were incubated with anti-NeuN 1:150 in block for 1 hour, washed 3 times with PBST, then incubated with FITC anti-mouse secondary antibody (Jackson Immunolabs) 1:200 in block. .. Cells were washed 3 times with PBST, with 0.5 mg/ml DAPI included in the second wash and mounted with ProLong Gold (Invitrogen).

    Article Title: A localization screen reveals translation factories and widespread co-translational RNA targeting
    Article Snippet: For P-body labelling, coverslips were incubated overnight at 4°C with a monoclonal mouse antibody (Santa Cruz sc-8418), diluted 1/250 in PBS containing 0.1% BSA. .. Coverslips were washed three times with PBS, 10 minutes each time, and incubated with an FITC anti-mouse secondary antibody (Jackson ImmunoResearch 115096-006) diluted 1/200 in PBS with 0.1% BSA. ..

    Article Title: Treadmill exercise ameliorates social isolation-induced depression through neuronal generation in rat pups
    Article Snippet: The sections were then incubated overnight with rabbit polyclonal anti-5-HT antibody (1:500; Immuno Star, Hudson, WI, USA) and mouse monoclonal anti-TPH antibody (1:500; Oncogene Research Products, Cambridge, UK). .. The sections were next incubated for 90 min with CY3 anti-rabbit secondary antibody (Vector Laboratories, Burlingame, CA, USA) and FITC anti-mouse secondary antibody (Jackson ImmunoResearch Laboratories, West Grove, PA, USA). .. The sections were then mounted on gelatin-coated glass slides, and the coverslips were mounted using fluorescent mounting medium (DakoCytomation, Carpinteria, CA, USA).

    Blocking Assay:

    Article Title: NeuN/Rbfox3 nuclear and cytoplasmic isoforms differentially regulate alternative splicing and nonsense-mediated decay of Rbfox2.
    Article Snippet: .. Cells were incubated with anti-NeuN 1:150 in block for 1 hour, washed 3 times with PBST, then incubated with FITC anti-mouse secondary antibody (Jackson Immunolabs) 1:200 in block. .. Cells were washed 3 times with PBST, with 0.5 mg/ml DAPI included in the second wash and mounted with ProLong Gold (Invitrogen).

    Purification:

    Article Title: Modification of the telomerase gene with human regulatory sequences resets mouse telomeres to human length
    Article Snippet: .. Novel plant genotypes Seed stocks Authentication Plants anti-gH2A.X (Ser139) (IF, Cell Signaling Technology, Cat. 2577S); anti-TRF2 (IF, NOVUS, Cat. NB100-56506); anti-E-Cadherin (IF, BD Biosciences, Cat. 610181); FITC anti-mouse secondary antibody (IF, Jackson Immunoresearch, Cat.115-095-003); Goat anti-Rabbit IgG (H+L) Cross-Adsorbed Secondary Antibody, Alexa FluorTM 594 (IF, Invitrogen, Cat. A-11012); PE anti-mouse CD4 (FC, Biolegend, 100407); FITC anti-mouse CD8a (FC, Biolegend, 100706); APC anti-mouse CD19 (FC, Biolegend, 115512); PE/Cyannine7 anti-mouse CD45 (FC, Biolegend, 147703); Ultra-LEAFTM Purified anti-mouse CD3! .. Antibody (T cells activation, Biolegend, 100359); purified antimouse CD28 (activation, Biolegend, 102101) Mouse anti-E-Cadherin antibody was reference published from the publication in Nature Communications (2023) 14:6708.



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    ( A ) Representative Western blot analysis of GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells ( n = 3 independent experiments). ( B ) Quantitation of relative GPC3 protein level from (A). Data are presented as means ± SD ( n = 3). Statistical analysis was performed using one-way ANOVA with a Tukey’s post hoc test. ( C ) Flow cytometry analysis of surface GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells. ( D ) Representative IHC images of GPC3 expression (brown) in tumor tissues from orthotopic Hepa1-6 tumor-bearing mice. Scale bar, 50 μm. ( E ) CLSM images of HepG2 and Hepa1-6 cells treated with SPD1 or SPD2 nanoparticles (50 μM; red fluorescence) for 6 hours. Scale bars, 20 μm. ( F ) Time-dependent CLSM imaging of HepG2 cells treated with SPD1 nanoparticles (50 μM) showing membrane-localized fibrillar transformation. Scale bars, 20 μm. ( G ) CLSM analysis of HepG2 cells sequentially incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours; red) and <t>FITC-labeled</t> anti-GPC3 antibody (green; 1:200; Abcam, #ab207080). Colocalization (yellow) indicates specific binding of SPD1 to membrane-bound GPC3. Fluorescence intensity and colocalization were quantified using MATLAB. Data are presented as means ± SD ( n = 3); n.s., not significant (one-way ANOVA with Tukey’s post hoc test). Scale bars, 20 μm. ( H ) SEM images of untreated HepG2 and WRL-68 cells or incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours). Magnified insets highlight membrane-associated fibrillar structures. ( I ) TEM images of untreated HepG2 cells (top) and those treated with SPD1 nanoparticles (50 μM, 24 hours; bottom). Red arrows indicate membrane-associated nanofibers. Scale bars, 500 nm. ( J ) SEM images showing the persistence of SPD1-derived fibrillar networks on HepG2 cells at 6, 24, and 72 hours posttreatment (50 μM). Scale bars, 2 μm. All experiments were independently repeated three times with consistent and reproducible results.
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    ( A ) Representative Western blot analysis of GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells ( n = 3 independent experiments). ( B ) Quantitation of relative GPC3 protein level from (A). Data are presented as means ± SD ( n = 3). Statistical analysis was performed using one-way ANOVA with a Tukey’s post hoc test. ( C ) Flow cytometry analysis of surface GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells. ( D ) Representative IHC images of GPC3 expression (brown) in tumor tissues from orthotopic Hepa1-6 tumor-bearing mice. Scale bar, 50 μm. ( E ) CLSM images of HepG2 and Hepa1-6 cells treated with SPD1 or SPD2 nanoparticles (50 μM; red fluorescence) for 6 hours. Scale bars, 20 μm. ( F ) Time-dependent CLSM imaging of HepG2 cells treated with SPD1 nanoparticles (50 μM) showing membrane-localized fibrillar transformation. Scale bars, 20 μm. ( G ) CLSM analysis of HepG2 cells sequentially incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours; red) and <t>FITC-labeled</t> anti-GPC3 antibody (green; 1:200; Abcam, #ab207080). Colocalization (yellow) indicates specific binding of SPD1 to membrane-bound GPC3. Fluorescence intensity and colocalization were quantified using MATLAB. Data are presented as means ± SD ( n = 3); n.s., not significant (one-way ANOVA with Tukey’s post hoc test). Scale bars, 20 μm. ( H ) SEM images of untreated HepG2 and WRL-68 cells or incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours). Magnified insets highlight membrane-associated fibrillar structures. ( I ) TEM images of untreated HepG2 cells (top) and those treated with SPD1 nanoparticles (50 μM, 24 hours; bottom). Red arrows indicate membrane-associated nanofibers. Scale bars, 500 nm. ( J ) SEM images showing the persistence of SPD1-derived fibrillar networks on HepG2 cells at 6, 24, and 72 hours posttreatment (50 μM). Scale bars, 2 μm. All experiments were independently repeated three times with consistent and reproducible results.
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    ( A ) Representative Western blot analysis of GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells ( n = 3 independent experiments). ( B ) Quantitation of relative GPC3 protein level from (A). Data are presented as means ± SD ( n = 3). Statistical analysis was performed using one-way ANOVA with a Tukey’s post hoc test. ( C ) Flow cytometry analysis of surface GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells. ( D ) Representative IHC images of GPC3 expression (brown) in tumor tissues from orthotopic Hepa1-6 tumor-bearing mice. Scale bar, 50 μm. ( E ) CLSM images of HepG2 and Hepa1-6 cells treated with SPD1 or SPD2 nanoparticles (50 μM; red fluorescence) for 6 hours. Scale bars, 20 μm. ( F ) Time-dependent CLSM imaging of HepG2 cells treated with SPD1 nanoparticles (50 μM) showing membrane-localized fibrillar transformation. Scale bars, 20 μm. ( G ) CLSM analysis of HepG2 cells sequentially incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours; red) and <t>FITC-labeled</t> anti-GPC3 antibody (green; 1:200; Abcam, #ab207080). Colocalization (yellow) indicates specific binding of SPD1 to membrane-bound GPC3. Fluorescence intensity and colocalization were quantified using MATLAB. Data are presented as means ± SD ( n = 3); n.s., not significant (one-way ANOVA with Tukey’s post hoc test). Scale bars, 20 μm. ( H ) SEM images of untreated HepG2 and WRL-68 cells or incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours). Magnified insets highlight membrane-associated fibrillar structures. ( I ) TEM images of untreated HepG2 cells (top) and those treated with SPD1 nanoparticles (50 μM, 24 hours; bottom). Red arrows indicate membrane-associated nanofibers. Scale bars, 500 nm. ( J ) SEM images showing the persistence of SPD1-derived fibrillar networks on HepG2 cells at 6, 24, and 72 hours posttreatment (50 μM). Scale bars, 2 μm. All experiments were independently repeated three times with consistent and reproducible results.
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    ( A ) Representative Western blot analysis of GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells ( n = 3 independent experiments). ( B ) Quantitation of relative GPC3 protein level from (A). Data are presented as means ± SD ( n = 3). Statistical analysis was performed using one-way ANOVA with a Tukey’s post hoc test. ( C ) Flow cytometry analysis of surface GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells. ( D ) Representative IHC images of GPC3 expression (brown) in tumor tissues from orthotopic Hepa1-6 tumor-bearing mice. Scale bar, 50 μm. ( E ) CLSM images of HepG2 and Hepa1-6 cells treated with SPD1 or SPD2 nanoparticles (50 μM; red fluorescence) for 6 hours. Scale bars, 20 μm. ( F ) Time-dependent CLSM imaging of HepG2 cells treated with SPD1 nanoparticles (50 μM) showing membrane-localized fibrillar transformation. Scale bars, 20 μm. ( G ) CLSM analysis of HepG2 cells sequentially incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours; red) and <t>FITC-labeled</t> anti-GPC3 antibody (green; 1:200; Abcam, #ab207080). Colocalization (yellow) indicates specific binding of SPD1 to membrane-bound GPC3. Fluorescence intensity and colocalization were quantified using MATLAB. Data are presented as means ± SD ( n = 3); n.s., not significant (one-way ANOVA with Tukey’s post hoc test). Scale bars, 20 μm. ( H ) SEM images of untreated HepG2 and WRL-68 cells or incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours). Magnified insets highlight membrane-associated fibrillar structures. ( I ) TEM images of untreated HepG2 cells (top) and those treated with SPD1 nanoparticles (50 μM, 24 hours; bottom). Red arrows indicate membrane-associated nanofibers. Scale bars, 500 nm. ( J ) SEM images showing the persistence of SPD1-derived fibrillar networks on HepG2 cells at 6, 24, and 72 hours posttreatment (50 μM). Scale bars, 2 μm. All experiments were independently repeated three times with consistent and reproducible results.
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    Jackson Immuno anti mouse secondary antibodies
    ( A ) Representative Western blot analysis of GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells ( n = 3 independent experiments). ( B ) Quantitation of relative GPC3 protein level from (A). Data are presented as means ± SD ( n = 3). Statistical analysis was performed using one-way ANOVA with a Tukey’s post hoc test. ( C ) Flow cytometry analysis of surface GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells. ( D ) Representative IHC images of GPC3 expression (brown) in tumor tissues from orthotopic Hepa1-6 tumor-bearing mice. Scale bar, 50 μm. ( E ) CLSM images of HepG2 and Hepa1-6 cells treated with SPD1 or SPD2 nanoparticles (50 μM; red fluorescence) for 6 hours. Scale bars, 20 μm. ( F ) Time-dependent CLSM imaging of HepG2 cells treated with SPD1 nanoparticles (50 μM) showing membrane-localized fibrillar transformation. Scale bars, 20 μm. ( G ) CLSM analysis of HepG2 cells sequentially incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours; red) and <t>FITC-labeled</t> anti-GPC3 antibody (green; 1:200; Abcam, #ab207080). Colocalization (yellow) indicates specific binding of SPD1 to membrane-bound GPC3. Fluorescence intensity and colocalization were quantified using MATLAB. Data are presented as means ± SD ( n = 3); n.s., not significant (one-way ANOVA with Tukey’s post hoc test). Scale bars, 20 μm. ( H ) SEM images of untreated HepG2 and WRL-68 cells or incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours). Magnified insets highlight membrane-associated fibrillar structures. ( I ) TEM images of untreated HepG2 cells (top) and those treated with SPD1 nanoparticles (50 μM, 24 hours; bottom). Red arrows indicate membrane-associated nanofibers. Scale bars, 500 nm. ( J ) SEM images showing the persistence of SPD1-derived fibrillar networks on HepG2 cells at 6, 24, and 72 hours posttreatment (50 μM). Scale bars, 2 μm. All experiments were independently repeated three times with consistent and reproducible results.
    Anti Mouse Secondary Antibodies, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    ( A ) Representative Western blot analysis of GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells ( n = 3 independent experiments). ( B ) Quantitation of relative GPC3 protein level from (A). Data are presented as means ± SD ( n = 3). Statistical analysis was performed using one-way ANOVA with a Tukey’s post hoc test. ( C ) Flow cytometry analysis of surface GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells. ( D ) Representative IHC images of GPC3 expression (brown) in tumor tissues from orthotopic Hepa1-6 tumor-bearing mice. Scale bar, 50 μm. ( E ) CLSM images of HepG2 and Hepa1-6 cells treated with SPD1 or SPD2 nanoparticles (50 μM; red fluorescence) for 6 hours. Scale bars, 20 μm. ( F ) Time-dependent CLSM imaging of HepG2 cells treated with SPD1 nanoparticles (50 μM) showing membrane-localized fibrillar transformation. Scale bars, 20 μm. ( G ) CLSM analysis of HepG2 cells sequentially incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours; red) and FITC-labeled anti-GPC3 antibody (green; 1:200; Abcam, #ab207080). Colocalization (yellow) indicates specific binding of SPD1 to membrane-bound GPC3. Fluorescence intensity and colocalization were quantified using MATLAB. Data are presented as means ± SD ( n = 3); n.s., not significant (one-way ANOVA with Tukey’s post hoc test). Scale bars, 20 μm. ( H ) SEM images of untreated HepG2 and WRL-68 cells or incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours). Magnified insets highlight membrane-associated fibrillar structures. ( I ) TEM images of untreated HepG2 cells (top) and those treated with SPD1 nanoparticles (50 μM, 24 hours; bottom). Red arrows indicate membrane-associated nanofibers. Scale bars, 500 nm. ( J ) SEM images showing the persistence of SPD1-derived fibrillar networks on HepG2 cells at 6, 24, and 72 hours posttreatment (50 μM). Scale bars, 2 μm. All experiments were independently repeated three times with consistent and reproducible results.

    Journal: Science Advances

    Article Title: In vivo membrane engineering traps Gd-based MRI contrast agents for detecting microhepatocellular carcinoma

    doi: 10.1126/sciadv.aec9913

    Figure Lengend Snippet: ( A ) Representative Western blot analysis of GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells ( n = 3 independent experiments). ( B ) Quantitation of relative GPC3 protein level from (A). Data are presented as means ± SD ( n = 3). Statistical analysis was performed using one-way ANOVA with a Tukey’s post hoc test. ( C ) Flow cytometry analysis of surface GPC3 expression in HepG2, WRL-68, and Hepa1-6 cells. ( D ) Representative IHC images of GPC3 expression (brown) in tumor tissues from orthotopic Hepa1-6 tumor-bearing mice. Scale bar, 50 μm. ( E ) CLSM images of HepG2 and Hepa1-6 cells treated with SPD1 or SPD2 nanoparticles (50 μM; red fluorescence) for 6 hours. Scale bars, 20 μm. ( F ) Time-dependent CLSM imaging of HepG2 cells treated with SPD1 nanoparticles (50 μM) showing membrane-localized fibrillar transformation. Scale bars, 20 μm. ( G ) CLSM analysis of HepG2 cells sequentially incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours; red) and FITC-labeled anti-GPC3 antibody (green; 1:200; Abcam, #ab207080). Colocalization (yellow) indicates specific binding of SPD1 to membrane-bound GPC3. Fluorescence intensity and colocalization were quantified using MATLAB. Data are presented as means ± SD ( n = 3); n.s., not significant (one-way ANOVA with Tukey’s post hoc test). Scale bars, 20 μm. ( H ) SEM images of untreated HepG2 and WRL-68 cells or incubated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours). Magnified insets highlight membrane-associated fibrillar structures. ( I ) TEM images of untreated HepG2 cells (top) and those treated with SPD1 nanoparticles (50 μM, 24 hours; bottom). Red arrows indicate membrane-associated nanofibers. Scale bars, 500 nm. ( J ) SEM images showing the persistence of SPD1-derived fibrillar networks on HepG2 cells at 6, 24, and 72 hours posttreatment (50 μM). Scale bars, 2 μm. All experiments were independently repeated three times with consistent and reproducible results.

    Article Snippet: After three PBS washes (5 min each), cells were incubated with an FITC-conjugated goat anti-mouse IgG secondary antibody (1:1000; Abbkine, A23210) for 1 hour at room temperature in the dark.

    Techniques: Western Blot, Expressing, Quantitation Assay, Flow Cytometry, Fluorescence, Imaging, Membrane, Transformation Assay, Incubation, Labeling, Binding Assay, Derivative Assay

    ( A ) Representative CLSM images of HepG2 cells incubated with SPD1 nanoparticles (red, 50 μM) for 6 hours, followed by treatment with FITC-N 3 (10 to 50 μM, green) for an additional 6 hours. Merged yellow fluorescence indicates successful copper-free click conjugation between DBCO and N 3 on the cell membrane. Cells treated with SPD2+FITC-N 3 (50 μM) served as the nontargeted controls, showing minimal colocalization. Scale bars, 20 μm. ( B ) Time-dependent kinetics of bioorthogonal conjugation quantified by BCA protein assay and ICP-MS. HepG2 cells were pretreated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours), followed by incubation with Gd-DOTA-N 3 (50 μM) for 0.5, 1, 6, or 12 hours. Cells treated with Gd-DOTA-N 3 alone served as baseline controls. ( C ) T 1 -weighted MR images of HepG2 cells treated with Gd-DOTA (50 μM), Gd-DOTA-N 3 (50 μM), or sequentially with SPD1 or SPD2 (50 μM, 6 hours) followed by Gd-DOTA-N 3 (50 μM, 6 hours). ( D ) Quantitative r 1 relaxivity under the corresponding treatment conditions in (C). ( E ) r 1 relaxivity of HepG2 cells preblocked with anti-GPC3 antibody (5 μg/ml, 12 hours; Abcam, #ab207080) before SPD1 treatment (50 μM, 6 hours) followed by Gd-DOTA-N 3 (50 μM, 6 hours). ( F to H ) Cell viability of WRL-68 (F), HepG2 (G), and Hepa1-6 (H) cells after sequential treatment with SPD1 or SPD2 for 6 hours followed by Gd-DOTA-N 3 (50 μM, 6 hours). Cell viability was quantified using the CCK-8 assay. Data are presented as means ± SD { n = 3 for [(A) to (E)]; n = 6 for [(F) to (H)]}. Statistical significance was performed using one-way ANOVA followed by Tukey’s post hoc test. P < 0.05 was considered statistically significant; n.s., not significant. All experiments were independently repeated three times with consistent results.

    Journal: Science Advances

    Article Title: In vivo membrane engineering traps Gd-based MRI contrast agents for detecting microhepatocellular carcinoma

    doi: 10.1126/sciadv.aec9913

    Figure Lengend Snippet: ( A ) Representative CLSM images of HepG2 cells incubated with SPD1 nanoparticles (red, 50 μM) for 6 hours, followed by treatment with FITC-N 3 (10 to 50 μM, green) for an additional 6 hours. Merged yellow fluorescence indicates successful copper-free click conjugation between DBCO and N 3 on the cell membrane. Cells treated with SPD2+FITC-N 3 (50 μM) served as the nontargeted controls, showing minimal colocalization. Scale bars, 20 μm. ( B ) Time-dependent kinetics of bioorthogonal conjugation quantified by BCA protein assay and ICP-MS. HepG2 cells were pretreated with SPD1 or SPD2 nanoparticles (50 μM, 6 hours), followed by incubation with Gd-DOTA-N 3 (50 μM) for 0.5, 1, 6, or 12 hours. Cells treated with Gd-DOTA-N 3 alone served as baseline controls. ( C ) T 1 -weighted MR images of HepG2 cells treated with Gd-DOTA (50 μM), Gd-DOTA-N 3 (50 μM), or sequentially with SPD1 or SPD2 (50 μM, 6 hours) followed by Gd-DOTA-N 3 (50 μM, 6 hours). ( D ) Quantitative r 1 relaxivity under the corresponding treatment conditions in (C). ( E ) r 1 relaxivity of HepG2 cells preblocked with anti-GPC3 antibody (5 μg/ml, 12 hours; Abcam, #ab207080) before SPD1 treatment (50 μM, 6 hours) followed by Gd-DOTA-N 3 (50 μM, 6 hours). ( F to H ) Cell viability of WRL-68 (F), HepG2 (G), and Hepa1-6 (H) cells after sequential treatment with SPD1 or SPD2 for 6 hours followed by Gd-DOTA-N 3 (50 μM, 6 hours). Cell viability was quantified using the CCK-8 assay. Data are presented as means ± SD { n = 3 for [(A) to (E)]; n = 6 for [(F) to (H)]}. Statistical significance was performed using one-way ANOVA followed by Tukey’s post hoc test. P < 0.05 was considered statistically significant; n.s., not significant. All experiments were independently repeated three times with consistent results.

    Article Snippet: After three PBS washes (5 min each), cells were incubated with an FITC-conjugated goat anti-mouse IgG secondary antibody (1:1000; Abbkine, A23210) for 1 hour at room temperature in the dark.

    Techniques: Incubation, Fluorescence, Conjugation Assay, Membrane, Bicinchoninic Acid Protein Assay, CCK-8 Assay