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98
InvivoGen poly i c
Free reactive oxygen species affect basal and LPS induced gene expression on lipid modified matrices. A Bar charts represent relative gene expression of TNFA and IL1A normalized to B2M showing mean ± SD, n = 4. THP-1 cells were activated with 15 nM PMA, cultured on modified matrices for 24 h, and then stimulated for 2 h with human TNFα (100 ng/ml), Flagellin (0.25 μg/ml, TLR5), FSL-1 (0.5 μg/ml, TLR2, TLR6), and <t>Poly(I:C)</t> (20 μg/ml <t>TLR3),</t> respectively. B, C Bar charts show relative expression levels of macrophages treated with 5 mM N-Acetylcysteine (NAC) immediately after seeding and cultured for 24 h, followed by stimulation with 500 ng/ml LPS for 2 h, mean ± SD, n = 4. Asterisks indicate statistical significance (* p < 0.05; ** p < 0.01; *** p < 0.005; **** p < 0.001), and exact p-values are shown for borderline significance determined by one-way ANOVA.
Poly I C, supplied by InvivoGen, used in various techniques. Bioz Stars score: 98/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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(A-C) A549-ACE2 cells were infected with SARS-CoV-2 or DVG-B (both MOI 1) and cells were collected at 6-, 10- and 24-hours post-inoculation for epifluorescence microscopy to ( A ) compare the cellular distribution of SARS-CoV-2 and DVG-B derived dsRNA or ( B ) quantify the total area of dsRNA signal and total dsRNA signal (integrated signal density). Insets depict magnified images of cells indicated by the white boxes. Scale bars indicate 10μm and 25μm, respectively. ****p<0.0001 by two-way ANOVA with Bonferroni’s multiple comparisons test. 6hpi N=4 fields. 10hpi N=5 fields. 24hpi, SARS-CoV-2 N=16 fields, DVG-B N=34 fields. Mean±SD. ( C ) IFNB1 and Spike gene expression were calculated relative to GAPDH by RT-qPCR. IFNB1 copy number values were normalized to mock for each respective timepoint. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Turkey’s multiple comparisons test. N=3, mean±SD. ( D-E ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (MOI 1) for 24-hours post-inoculation, respectively. N=2 biological repeats ( D ) Epifluorescence images of dsRNA and IRF-3 staining in DMSO (left) and remdesivir (right) treated cells. Scale bars indicate 25μM. Arrowheads indicate dsRNA+ cells with nuclear IRF-3. Remdesivir treated SARS-CoV-2 and DVG-B infections N=3 fields, respectively. DMSO treated SARS-CoV-2 infections N=14 fields, DVG-B infections N=30 fields. ( E ) Quantification of the proportion of dsRNA+ cells with nuclear IRF-3 relative to the total number of dsRNA+ cells per field. Each point represents one field. ****p<0.0001 by unpaired two-tailed t-test. SARS-CoV-2: 412 dsRNA+ cells among N=14 fields. DVG-B: 120 dsRNA+ cells among N=30 fields. ( F ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (both MOI 1) or were transfected with low molecular weight <t>poly(I:C)</t> (PIC) for 24-hours post-inoculation, respectively. IL-29 and Spike gene copy numbers were calculated relative to GAPDH by RT-qPCR. **p<0.01, ****p<0.0001 by mixed-effects analysis with Bonferroni’s multiple comparisons test. N=3, mean±SD. ( G ) A549-ACE2 cells were treated with 10μM remdesivir or an equal volume of DMSO and infected with DVG-A (MOI 1) for 24-hours post-inoculation. DVG-A levels were quantified relative to GAPDH by RT-qPCR. N=4, mean±SD. ( H ) A549-ACE2 MAVS KO, PKR KO, or empty vector transduced control cells (parent) were infected with DVG-B (MOI 5) for 24-hours post-inoculation. IFNB1, IL-29, ISG54 and DVG-B copy numbers were quantified relative to GAPDH by RT-qPCR. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Bonferroni’s multiple comparisons test. N=4, mean±SD.
Low Molecular Weight Poly I C, supplied by InvivoGen, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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(A-C) A549-ACE2 cells were infected with SARS-CoV-2 or DVG-B (both MOI 1) and cells were collected at 6-, 10- and 24-hours post-inoculation for epifluorescence microscopy to ( A ) compare the cellular distribution of SARS-CoV-2 and DVG-B derived dsRNA or ( B ) quantify the total area of dsRNA signal and total dsRNA signal (integrated signal density). Insets depict magnified images of cells indicated by the white boxes. Scale bars indicate 10μm and 25μm, respectively. ****p<0.0001 by two-way ANOVA with Bonferroni’s multiple comparisons test. 6hpi N=4 fields. 10hpi N=5 fields. 24hpi, SARS-CoV-2 N=16 fields, DVG-B N=34 fields. Mean±SD. ( C ) IFNB1 and Spike gene expression were calculated relative to GAPDH by RT-qPCR. IFNB1 copy number values were normalized to mock for each respective timepoint. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Turkey’s multiple comparisons test. N=3, mean±SD. ( D-E ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (MOI 1) for 24-hours post-inoculation, respectively. N=2 biological repeats ( D ) Epifluorescence images of dsRNA and IRF-3 staining in DMSO (left) and remdesivir (right) treated cells. Scale bars indicate 25μM. Arrowheads indicate dsRNA+ cells with nuclear IRF-3. Remdesivir treated SARS-CoV-2 and DVG-B infections N=3 fields, respectively. DMSO treated SARS-CoV-2 infections N=14 fields, DVG-B infections N=30 fields. ( E ) Quantification of the proportion of dsRNA+ cells with nuclear IRF-3 relative to the total number of dsRNA+ cells per field. Each point represents one field. ****p<0.0001 by unpaired two-tailed t-test. SARS-CoV-2: 412 dsRNA+ cells among N=14 fields. DVG-B: 120 dsRNA+ cells among N=30 fields. ( F ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (both MOI 1) or were transfected with low molecular weight <t>poly(I:C)</t> (PIC) for 24-hours post-inoculation, respectively. IL-29 and Spike gene copy numbers were calculated relative to GAPDH by RT-qPCR. **p<0.01, ****p<0.0001 by mixed-effects analysis with Bonferroni’s multiple comparisons test. N=3, mean±SD. ( G ) A549-ACE2 cells were treated with 10μM remdesivir or an equal volume of DMSO and infected with DVG-A (MOI 1) for 24-hours post-inoculation. DVG-A levels were quantified relative to GAPDH by RT-qPCR. N=4, mean±SD. ( H ) A549-ACE2 MAVS KO, PKR KO, or empty vector transduced control cells (parent) were infected with DVG-B (MOI 5) for 24-hours post-inoculation. IFNB1, IL-29, ISG54 and DVG-B copy numbers were quantified relative to GAPDH by RT-qPCR. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Bonferroni’s multiple comparisons test. N=4, mean±SD.
High Molecular Weight Poly I C Invivogen Tlrl Pic Low Molecular Weight, supplied by InvivoGen, used in various techniques. Bioz Stars score: 98/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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InvivoGen high molecular weight poly i c
(A-C) A549-ACE2 cells were infected with SARS-CoV-2 or DVG-B (both MOI 1) and cells were collected at 6-, 10- and 24-hours post-inoculation for epifluorescence microscopy to ( A ) compare the cellular distribution of SARS-CoV-2 and DVG-B derived dsRNA or ( B ) quantify the total area of dsRNA signal and total dsRNA signal (integrated signal density). Insets depict magnified images of cells indicated by the white boxes. Scale bars indicate 10μm and 25μm, respectively. ****p<0.0001 by two-way ANOVA with Bonferroni’s multiple comparisons test. 6hpi N=4 fields. 10hpi N=5 fields. 24hpi, SARS-CoV-2 N=16 fields, DVG-B N=34 fields. Mean±SD. ( C ) IFNB1 and Spike gene expression were calculated relative to GAPDH by RT-qPCR. IFNB1 copy number values were normalized to mock for each respective timepoint. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Turkey’s multiple comparisons test. N=3, mean±SD. ( D-E ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (MOI 1) for 24-hours post-inoculation, respectively. N=2 biological repeats ( D ) Epifluorescence images of dsRNA and IRF-3 staining in DMSO (left) and remdesivir (right) treated cells. Scale bars indicate 25μM. Arrowheads indicate dsRNA+ cells with nuclear IRF-3. Remdesivir treated SARS-CoV-2 and DVG-B infections N=3 fields, respectively. DMSO treated SARS-CoV-2 infections N=14 fields, DVG-B infections N=30 fields. ( E ) Quantification of the proportion of dsRNA+ cells with nuclear IRF-3 relative to the total number of dsRNA+ cells per field. Each point represents one field. ****p<0.0001 by unpaired two-tailed t-test. SARS-CoV-2: 412 dsRNA+ cells among N=14 fields. DVG-B: 120 dsRNA+ cells among N=30 fields. ( F ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (both MOI 1) or were transfected with low molecular weight <t>poly(I:C)</t> (PIC) for 24-hours post-inoculation, respectively. IL-29 and Spike gene copy numbers were calculated relative to GAPDH by RT-qPCR. **p<0.01, ****p<0.0001 by mixed-effects analysis with Bonferroni’s multiple comparisons test. N=3, mean±SD. ( G ) A549-ACE2 cells were treated with 10μM remdesivir or an equal volume of DMSO and infected with DVG-A (MOI 1) for 24-hours post-inoculation. DVG-A levels were quantified relative to GAPDH by RT-qPCR. N=4, mean±SD. ( H ) A549-ACE2 MAVS KO, PKR KO, or empty vector transduced control cells (parent) were infected with DVG-B (MOI 5) for 24-hours post-inoculation. IFNB1, IL-29, ISG54 and DVG-B copy numbers were quantified relative to GAPDH by RT-qPCR. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Bonferroni’s multiple comparisons test. N=4, mean±SD.
High Molecular Weight Poly I C, supplied by InvivoGen, used in various techniques. Bioz Stars score: 98/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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(A-C) A549-ACE2 cells were infected with SARS-CoV-2 or DVG-B (both MOI 1) and cells were collected at 6-, 10- and 24-hours post-inoculation for epifluorescence microscopy to ( A ) compare the cellular distribution of SARS-CoV-2 and DVG-B derived dsRNA or ( B ) quantify the total area of dsRNA signal and total dsRNA signal (integrated signal density). Insets depict magnified images of cells indicated by the white boxes. Scale bars indicate 10μm and 25μm, respectively. ****p<0.0001 by two-way ANOVA with Bonferroni’s multiple comparisons test. 6hpi N=4 fields. 10hpi N=5 fields. 24hpi, SARS-CoV-2 N=16 fields, DVG-B N=34 fields. Mean±SD. ( C ) IFNB1 and Spike gene expression were calculated relative to GAPDH by RT-qPCR. IFNB1 copy number values were normalized to mock for each respective timepoint. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Turkey’s multiple comparisons test. N=3, mean±SD. ( D-E ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (MOI 1) for 24-hours post-inoculation, respectively. N=2 biological repeats ( D ) Epifluorescence images of dsRNA and IRF-3 staining in DMSO (left) and remdesivir (right) treated cells. Scale bars indicate 25μM. Arrowheads indicate dsRNA+ cells with nuclear IRF-3. Remdesivir treated SARS-CoV-2 and DVG-B infections N=3 fields, respectively. DMSO treated SARS-CoV-2 infections N=14 fields, DVG-B infections N=30 fields. ( E ) Quantification of the proportion of dsRNA+ cells with nuclear IRF-3 relative to the total number of dsRNA+ cells per field. Each point represents one field. ****p<0.0001 by unpaired two-tailed t-test. SARS-CoV-2: 412 dsRNA+ cells among N=14 fields. DVG-B: 120 dsRNA+ cells among N=30 fields. ( F ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (both MOI 1) or were transfected with low molecular weight <t>poly(I:C)</t> (PIC) for 24-hours post-inoculation, respectively. IL-29 and Spike gene copy numbers were calculated relative to GAPDH by RT-qPCR. **p<0.01, ****p<0.0001 by mixed-effects analysis with Bonferroni’s multiple comparisons test. N=3, mean±SD. ( G ) A549-ACE2 cells were treated with 10μM remdesivir or an equal volume of DMSO and infected with DVG-A (MOI 1) for 24-hours post-inoculation. DVG-A levels were quantified relative to GAPDH by RT-qPCR. N=4, mean±SD. ( H ) A549-ACE2 MAVS KO, PKR KO, or empty vector transduced control cells (parent) were infected with DVG-B (MOI 5) for 24-hours post-inoculation. IFNB1, IL-29, ISG54 and DVG-B copy numbers were quantified relative to GAPDH by RT-qPCR. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Bonferroni’s multiple comparisons test. N=4, mean±SD.
Tlrl Picw Fluorescein Labelled Poly I C Invivogen Tlrl Picf Rhodamine Labelled, supplied by InvivoGen, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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(A-C) A549-ACE2 cells were infected with SARS-CoV-2 or DVG-B (both MOI 1) and cells were collected at 6-, 10- and 24-hours post-inoculation for epifluorescence microscopy to ( A ) compare the cellular distribution of SARS-CoV-2 and DVG-B derived dsRNA or ( B ) quantify the total area of dsRNA signal and total dsRNA signal (integrated signal density). Insets depict magnified images of cells indicated by the white boxes. Scale bars indicate 10μm and 25μm, respectively. ****p<0.0001 by two-way ANOVA with Bonferroni’s multiple comparisons test. 6hpi N=4 fields. 10hpi N=5 fields. 24hpi, SARS-CoV-2 N=16 fields, DVG-B N=34 fields. Mean±SD. ( C ) IFNB1 and Spike gene expression were calculated relative to GAPDH by RT-qPCR. IFNB1 copy number values were normalized to mock for each respective timepoint. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Turkey’s multiple comparisons test. N=3, mean±SD. ( D-E ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (MOI 1) for 24-hours post-inoculation, respectively. N=2 biological repeats ( D ) Epifluorescence images of dsRNA and IRF-3 staining in DMSO (left) and remdesivir (right) treated cells. Scale bars indicate 25μM. Arrowheads indicate dsRNA+ cells with nuclear IRF-3. Remdesivir treated SARS-CoV-2 and DVG-B infections N=3 fields, respectively. DMSO treated SARS-CoV-2 infections N=14 fields, DVG-B infections N=30 fields. ( E ) Quantification of the proportion of dsRNA+ cells with nuclear IRF-3 relative to the total number of dsRNA+ cells per field. Each point represents one field. ****p<0.0001 by unpaired two-tailed t-test. SARS-CoV-2: 412 dsRNA+ cells among N=14 fields. DVG-B: 120 dsRNA+ cells among N=30 fields. ( F ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (both MOI 1) or were transfected with low molecular weight <t>poly(I:C)</t> (PIC) for 24-hours post-inoculation, respectively. IL-29 and Spike gene copy numbers were calculated relative to GAPDH by RT-qPCR. **p<0.01, ****p<0.0001 by mixed-effects analysis with Bonferroni’s multiple comparisons test. N=3, mean±SD. ( G ) A549-ACE2 cells were treated with 10μM remdesivir or an equal volume of DMSO and infected with DVG-A (MOI 1) for 24-hours post-inoculation. DVG-A levels were quantified relative to GAPDH by RT-qPCR. N=4, mean±SD. ( H ) A549-ACE2 MAVS KO, PKR KO, or empty vector transduced control cells (parent) were infected with DVG-B (MOI 5) for 24-hours post-inoculation. IFNB1, IL-29, ISG54 and DVG-B copy numbers were quantified relative to GAPDH by RT-qPCR. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Bonferroni’s multiple comparisons test. N=4, mean±SD.
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(A-C) A549-ACE2 cells were infected with SARS-CoV-2 or DVG-B (both MOI 1) and cells were collected at 6-, 10- and 24-hours post-inoculation for epifluorescence microscopy to ( A ) compare the cellular distribution of SARS-CoV-2 and DVG-B derived dsRNA or ( B ) quantify the total area of dsRNA signal and total dsRNA signal (integrated signal density). Insets depict magnified images of cells indicated by the white boxes. Scale bars indicate 10μm and 25μm, respectively. ****p<0.0001 by two-way ANOVA with Bonferroni’s multiple comparisons test. 6hpi N=4 fields. 10hpi N=5 fields. 24hpi, SARS-CoV-2 N=16 fields, DVG-B N=34 fields. Mean±SD. ( C ) IFNB1 and Spike gene expression were calculated relative to GAPDH by RT-qPCR. IFNB1 copy number values were normalized to mock for each respective timepoint. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Turkey’s multiple comparisons test. N=3, mean±SD. ( D-E ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (MOI 1) for 24-hours post-inoculation, respectively. N=2 biological repeats ( D ) Epifluorescence images of dsRNA and IRF-3 staining in DMSO (left) and remdesivir (right) treated cells. Scale bars indicate 25μM. Arrowheads indicate dsRNA+ cells with nuclear IRF-3. Remdesivir treated SARS-CoV-2 and DVG-B infections N=3 fields, respectively. DMSO treated SARS-CoV-2 infections N=14 fields, DVG-B infections N=30 fields. ( E ) Quantification of the proportion of dsRNA+ cells with nuclear IRF-3 relative to the total number of dsRNA+ cells per field. Each point represents one field. ****p<0.0001 by unpaired two-tailed t-test. SARS-CoV-2: 412 dsRNA+ cells among N=14 fields. DVG-B: 120 dsRNA+ cells among N=30 fields. ( F ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (both MOI 1) or were transfected with low molecular weight <t>poly(I:C)</t> (PIC) for 24-hours post-inoculation, respectively. IL-29 and Spike gene copy numbers were calculated relative to GAPDH by RT-qPCR. **p<0.01, ****p<0.0001 by mixed-effects analysis with Bonferroni’s multiple comparisons test. N=3, mean±SD. ( G ) A549-ACE2 cells were treated with 10μM remdesivir or an equal volume of DMSO and infected with DVG-A (MOI 1) for 24-hours post-inoculation. DVG-A levels were quantified relative to GAPDH by RT-qPCR. N=4, mean±SD. ( H ) A549-ACE2 MAVS KO, PKR KO, or empty vector transduced control cells (parent) were infected with DVG-B (MOI 5) for 24-hours post-inoculation. IFNB1, IL-29, ISG54 and DVG-B copy numbers were quantified relative to GAPDH by RT-qPCR. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Bonferroni’s multiple comparisons test. N=4, mean±SD.
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Free reactive oxygen species affect basal and LPS induced gene expression on lipid modified matrices. A Bar charts represent relative gene expression of TNFA and IL1A normalized to B2M showing mean ± SD, n = 4. THP-1 cells were activated with 15 nM PMA, cultured on modified matrices for 24 h, and then stimulated for 2 h with human TNFα (100 ng/ml), Flagellin (0.25 μg/ml, TLR5), FSL-1 (0.5 μg/ml, TLR2, TLR6), and Poly(I:C) (20 μg/ml TLR3), respectively. B, C Bar charts show relative expression levels of macrophages treated with 5 mM N-Acetylcysteine (NAC) immediately after seeding and cultured for 24 h, followed by stimulation with 500 ng/ml LPS for 2 h, mean ± SD, n = 4. Asterisks indicate statistical significance (* p < 0.05; ** p < 0.01; *** p < 0.005; **** p < 0.001), and exact p-values are shown for borderline significance determined by one-way ANOVA.

Journal: Redox Biology

Article Title: Modification of the dermal matrix by senescence associated lipids and its functional consequence

doi: 10.1016/j.redox.2026.104069

Figure Lengend Snippet: Free reactive oxygen species affect basal and LPS induced gene expression on lipid modified matrices. A Bar charts represent relative gene expression of TNFA and IL1A normalized to B2M showing mean ± SD, n = 4. THP-1 cells were activated with 15 nM PMA, cultured on modified matrices for 24 h, and then stimulated for 2 h with human TNFα (100 ng/ml), Flagellin (0.25 μg/ml, TLR5), FSL-1 (0.5 μg/ml, TLR2, TLR6), and Poly(I:C) (20 μg/ml TLR3), respectively. B, C Bar charts show relative expression levels of macrophages treated with 5 mM N-Acetylcysteine (NAC) immediately after seeding and cultured for 24 h, followed by stimulation with 500 ng/ml LPS for 2 h, mean ± SD, n = 4. Asterisks indicate statistical significance (* p < 0.05; ** p < 0.01; *** p < 0.005; **** p < 0.001), and exact p-values are shown for borderline significance determined by one-way ANOVA.

Article Snippet: In detail, human TNFα (InvitrogenThermo Fisher, USA) was added to a final concentration of 100 ng/ml, 0.25 μg/ml of Flagellin (TLR5, InvivoGen, France), 0.5 μg/ml of FSL-1 (TLR2, TLR6, InvivoGen) and 20 μg/ml of Poly(I:C) (TLR3, InvivoGen) [ , ].

Techniques: Gene Expression, Modification, Cell Culture, Expressing

(A-C) A549-ACE2 cells were infected with SARS-CoV-2 or DVG-B (both MOI 1) and cells were collected at 6-, 10- and 24-hours post-inoculation for epifluorescence microscopy to ( A ) compare the cellular distribution of SARS-CoV-2 and DVG-B derived dsRNA or ( B ) quantify the total area of dsRNA signal and total dsRNA signal (integrated signal density). Insets depict magnified images of cells indicated by the white boxes. Scale bars indicate 10μm and 25μm, respectively. ****p<0.0001 by two-way ANOVA with Bonferroni’s multiple comparisons test. 6hpi N=4 fields. 10hpi N=5 fields. 24hpi, SARS-CoV-2 N=16 fields, DVG-B N=34 fields. Mean±SD. ( C ) IFNB1 and Spike gene expression were calculated relative to GAPDH by RT-qPCR. IFNB1 copy number values were normalized to mock for each respective timepoint. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Turkey’s multiple comparisons test. N=3, mean±SD. ( D-E ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (MOI 1) for 24-hours post-inoculation, respectively. N=2 biological repeats ( D ) Epifluorescence images of dsRNA and IRF-3 staining in DMSO (left) and remdesivir (right) treated cells. Scale bars indicate 25μM. Arrowheads indicate dsRNA+ cells with nuclear IRF-3. Remdesivir treated SARS-CoV-2 and DVG-B infections N=3 fields, respectively. DMSO treated SARS-CoV-2 infections N=14 fields, DVG-B infections N=30 fields. ( E ) Quantification of the proportion of dsRNA+ cells with nuclear IRF-3 relative to the total number of dsRNA+ cells per field. Each point represents one field. ****p<0.0001 by unpaired two-tailed t-test. SARS-CoV-2: 412 dsRNA+ cells among N=14 fields. DVG-B: 120 dsRNA+ cells among N=30 fields. ( F ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (both MOI 1) or were transfected with low molecular weight poly(I:C) (PIC) for 24-hours post-inoculation, respectively. IL-29 and Spike gene copy numbers were calculated relative to GAPDH by RT-qPCR. **p<0.01, ****p<0.0001 by mixed-effects analysis with Bonferroni’s multiple comparisons test. N=3, mean±SD. ( G ) A549-ACE2 cells were treated with 10μM remdesivir or an equal volume of DMSO and infected with DVG-A (MOI 1) for 24-hours post-inoculation. DVG-A levels were quantified relative to GAPDH by RT-qPCR. N=4, mean±SD. ( H ) A549-ACE2 MAVS KO, PKR KO, or empty vector transduced control cells (parent) were infected with DVG-B (MOI 5) for 24-hours post-inoculation. IFNB1, IL-29, ISG54 and DVG-B copy numbers were quantified relative to GAPDH by RT-qPCR. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Bonferroni’s multiple comparisons test. N=4, mean±SD.

Journal: bioRxiv

Article Title: SARS-CoV-2 Defective Viral Genomes from Distinct Genomic Regions Drive Divergent Interferon Responses

doi: 10.64898/2026.03.19.712870

Figure Lengend Snippet: (A-C) A549-ACE2 cells were infected with SARS-CoV-2 or DVG-B (both MOI 1) and cells were collected at 6-, 10- and 24-hours post-inoculation for epifluorescence microscopy to ( A ) compare the cellular distribution of SARS-CoV-2 and DVG-B derived dsRNA or ( B ) quantify the total area of dsRNA signal and total dsRNA signal (integrated signal density). Insets depict magnified images of cells indicated by the white boxes. Scale bars indicate 10μm and 25μm, respectively. ****p<0.0001 by two-way ANOVA with Bonferroni’s multiple comparisons test. 6hpi N=4 fields. 10hpi N=5 fields. 24hpi, SARS-CoV-2 N=16 fields, DVG-B N=34 fields. Mean±SD. ( C ) IFNB1 and Spike gene expression were calculated relative to GAPDH by RT-qPCR. IFNB1 copy number values were normalized to mock for each respective timepoint. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Turkey’s multiple comparisons test. N=3, mean±SD. ( D-E ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (MOI 1) for 24-hours post-inoculation, respectively. N=2 biological repeats ( D ) Epifluorescence images of dsRNA and IRF-3 staining in DMSO (left) and remdesivir (right) treated cells. Scale bars indicate 25μM. Arrowheads indicate dsRNA+ cells with nuclear IRF-3. Remdesivir treated SARS-CoV-2 and DVG-B infections N=3 fields, respectively. DMSO treated SARS-CoV-2 infections N=14 fields, DVG-B infections N=30 fields. ( E ) Quantification of the proportion of dsRNA+ cells with nuclear IRF-3 relative to the total number of dsRNA+ cells per field. Each point represents one field. ****p<0.0001 by unpaired two-tailed t-test. SARS-CoV-2: 412 dsRNA+ cells among N=14 fields. DVG-B: 120 dsRNA+ cells among N=30 fields. ( F ) A549-ACE2 cells treated with 10μM remdesivir or an equal volume of DMSO were infected with SARS-CoV-2 or DVG-B (both MOI 1) or were transfected with low molecular weight poly(I:C) (PIC) for 24-hours post-inoculation, respectively. IL-29 and Spike gene copy numbers were calculated relative to GAPDH by RT-qPCR. **p<0.01, ****p<0.0001 by mixed-effects analysis with Bonferroni’s multiple comparisons test. N=3, mean±SD. ( G ) A549-ACE2 cells were treated with 10μM remdesivir or an equal volume of DMSO and infected with DVG-A (MOI 1) for 24-hours post-inoculation. DVG-A levels were quantified relative to GAPDH by RT-qPCR. N=4, mean±SD. ( H ) A549-ACE2 MAVS KO, PKR KO, or empty vector transduced control cells (parent) were infected with DVG-B (MOI 5) for 24-hours post-inoculation. IFNB1, IL-29, ISG54 and DVG-B copy numbers were quantified relative to GAPDH by RT-qPCR. *p<0.05, **p<0.01, ***p<0.001 by two-way ANOVA with Bonferroni’s multiple comparisons test. N=4, mean±SD.

Article Snippet: Alternatively, cells were transfected with 1 μg/mL low molecular weight poly(I:C) (InvivoGen catalog no. tlrl-picw) via lipofectamine 2000.

Techniques: Infection, Epifluorescence Microscopy, Derivative Assay, Gene Expression, Quantitative RT-PCR, Staining, Two Tailed Test, Transfection, Molecular Weight, Plasmid Preparation, Control