recombinant human α-syn monomer Search Results


human recombinant α-syn monomers  (Merck & Co)
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Merck & Co human recombinant α-syn monomers
SARS-CoV-2 differently affects SNCA mRNA expression and <t>α-syn</t> protein levels at 24 and 48 h post-infection in A549-hACE2 and CaLu-3 epithelial lung cells. A qPCR-based relative quantification of SNCA mRNA in Mock- and SARS-CoV-2-infected cells (MOI 0.05), either in the presence or absence of <t>exogenous</t> <t>IFN-β,</t> 24 and 48 h post-infection. Results are shown as mean ± SEM from n = 3 independent experiments. Statistical analysis was performed by applying Two-way ANOVA, followed by multiple testing correction. To avoid graphs overcrowding, p values are shown for statistically significant groups of interest only. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. B and C Representative immunofluorescence images for SARS-CoV-2 Spike (S) and α-syn proteins in Mock- and SARS-CoV-2-infected (MOI 0.05) in A549-hACE2 ( B ) and CaLu-3 ( C ) epithelial lung cells 24 and 48 h post-infection, in the absence (CTR) or presence of IFN-β. Cells were fixed in 4% Formaldehyde solution for 15 min and permeabilized with 0.1% of Triton X-100 for 15 min. Images are representative of n = 3 independent experiments. Bars correspond to 20 μm
Human Recombinant α Syn Monomers, supplied by Merck & Co, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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SARS-CoV-2 differently affects SNCA mRNA expression and α-syn protein levels at 24 and 48 h post-infection in A549-hACE2 and CaLu-3 epithelial lung cells. A qPCR-based relative quantification of SNCA mRNA in Mock- and SARS-CoV-2-infected cells (MOI 0.05), either in the presence or absence of exogenous IFN-β, 24 and 48 h post-infection. Results are shown as mean ± SEM from n = 3 independent experiments. Statistical analysis was performed by applying Two-way ANOVA, followed by multiple testing correction. To avoid graphs overcrowding, p values are shown for statistically significant groups of interest only. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. B and C Representative immunofluorescence images for SARS-CoV-2 Spike (S) and α-syn proteins in Mock- and SARS-CoV-2-infected (MOI 0.05) in A549-hACE2 ( B ) and CaLu-3 ( C ) epithelial lung cells 24 and 48 h post-infection, in the absence (CTR) or presence of IFN-β. Cells were fixed in 4% Formaldehyde solution for 15 min and permeabilized with 0.1% of Triton X-100 for 15 min. Images are representative of n = 3 independent experiments. Bars correspond to 20 μm

Journal: Biological Research

Article Title: Alpha-synuclein dynamics bridge Type-I Interferon response and SARS-CoV-2 replication in peripheral cells

doi: 10.1186/s40659-023-00482-x

Figure Lengend Snippet: SARS-CoV-2 differently affects SNCA mRNA expression and α-syn protein levels at 24 and 48 h post-infection in A549-hACE2 and CaLu-3 epithelial lung cells. A qPCR-based relative quantification of SNCA mRNA in Mock- and SARS-CoV-2-infected cells (MOI 0.05), either in the presence or absence of exogenous IFN-β, 24 and 48 h post-infection. Results are shown as mean ± SEM from n = 3 independent experiments. Statistical analysis was performed by applying Two-way ANOVA, followed by multiple testing correction. To avoid graphs overcrowding, p values are shown for statistically significant groups of interest only. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. B and C Representative immunofluorescence images for SARS-CoV-2 Spike (S) and α-syn proteins in Mock- and SARS-CoV-2-infected (MOI 0.05) in A549-hACE2 ( B ) and CaLu-3 ( C ) epithelial lung cells 24 and 48 h post-infection, in the absence (CTR) or presence of IFN-β. Cells were fixed in 4% Formaldehyde solution for 15 min and permeabilized with 0.1% of Triton X-100 for 15 min. Images are representative of n = 3 independent experiments. Bars correspond to 20 μm

Article Snippet: Type-I IFN (human recombinant IFN-β, 500 IU/ml, BEI resources) and human recombinant α-syn monomers (1 μM, Merck-Sigma), were exogenously pre-administered to cells, both alone and in combination, at 24 h pre-infection.

Techniques: Expressing, Infection, Immunofluorescence

A1 and A2 Exogenous administration of α-syn monomers increases SARS-CoV-2 replication in A549-hACE2 and CaLu-3 epithelial cells, which is prevented by IFN-β. Quantification of viral replication from cell supernatants through RT-qPCR for the SARS-CoV-2 N gene in infected A549-hACE2 ( A1 ) and CaLu-3 cells ( A2 ), 24 and 48 h post-infection (MOI 0.05). A1 In A549-hACE2 cells, exogenous α-syn monomers significantly increased viral replication compared with infected CTR both at 24 and 48 h post-infection. A2 In CaLu-3 cells the pro-viral effect of exogenous α-syn monomers was maximally detected at 24 h, while it declined at 48 h post-infection. In both cases, IFN-β, either alone or in combination with exogenous α-syn, completely prevented SARS-CoV-2 replication. Results show SARS-CoV-2 N gene copy numbers quantified from the RNA isolated from 100 μl of cell supernatants. Results are shown as mean ± SEM from n ≥ 5 independent experiments. Values for 24 and 48 h post-infection were analyzed by applying One-Way ANOVA followed by multiple testing correction. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. B1 and B2 Representative immunofluorescence images of SARS-CoV-2 Nucleocapsid (N) and α-syn proteins in Mock- and SARS-CoV-2-Infected A549-hACE2 epithelial lung cells 48 h post-infection (MOI 0.05), in the absence and presence of IFN-β, exogenous α-syn monomers, or both. SARS-CoV-2 replication is enhanced in the presence of exogenous α-syn monomers, which impair accumulation of permeabilization-resistant α-syn species that are instead increased by IFN-β. B1 For visualization of large, permeabilization-resistant α-syn species, that potentially correspond to multimers or intracellular membrane-bound species, cells were fixed for 15 min in 4% Formaldehyde solution and permeabilized with 0.3% of Triton X-100 for 15 min. Bars correspond to 20 μm. B2 For better preservation, and visualization of small, highly-soluble α-syn species potentially corresponding to monomers, cells were fixed for 15 min in 4% Paraformaldehyde (PFA) and permeabilized with 0.1% of Triton X-100 for 10 min. Bars correspond to 20 μm. Images are representative of n = 3 independent experiments. C Quantification of SARS-CoV-2-N-positive cells in SARS-CoV-2-infected CTR vs α-syn-treated SARS-CoV-2-infected cells. Values are expressed as the percentage of N-positive cells ± SD calculated as the number of N-positive cells/total cells per microscopic field from n = 12 microscopic fields per experimental group, from n = 3 independent experiments. Data were analyzed by applying the Student’s t-test for comparison between VIRUS CTR and VIRUS α-syn groups. **p < 0.01

Journal: Biological Research

Article Title: Alpha-synuclein dynamics bridge Type-I Interferon response and SARS-CoV-2 replication in peripheral cells

doi: 10.1186/s40659-023-00482-x

Figure Lengend Snippet: A1 and A2 Exogenous administration of α-syn monomers increases SARS-CoV-2 replication in A549-hACE2 and CaLu-3 epithelial cells, which is prevented by IFN-β. Quantification of viral replication from cell supernatants through RT-qPCR for the SARS-CoV-2 N gene in infected A549-hACE2 ( A1 ) and CaLu-3 cells ( A2 ), 24 and 48 h post-infection (MOI 0.05). A1 In A549-hACE2 cells, exogenous α-syn monomers significantly increased viral replication compared with infected CTR both at 24 and 48 h post-infection. A2 In CaLu-3 cells the pro-viral effect of exogenous α-syn monomers was maximally detected at 24 h, while it declined at 48 h post-infection. In both cases, IFN-β, either alone or in combination with exogenous α-syn, completely prevented SARS-CoV-2 replication. Results show SARS-CoV-2 N gene copy numbers quantified from the RNA isolated from 100 μl of cell supernatants. Results are shown as mean ± SEM from n ≥ 5 independent experiments. Values for 24 and 48 h post-infection were analyzed by applying One-Way ANOVA followed by multiple testing correction. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. B1 and B2 Representative immunofluorescence images of SARS-CoV-2 Nucleocapsid (N) and α-syn proteins in Mock- and SARS-CoV-2-Infected A549-hACE2 epithelial lung cells 48 h post-infection (MOI 0.05), in the absence and presence of IFN-β, exogenous α-syn monomers, or both. SARS-CoV-2 replication is enhanced in the presence of exogenous α-syn monomers, which impair accumulation of permeabilization-resistant α-syn species that are instead increased by IFN-β. B1 For visualization of large, permeabilization-resistant α-syn species, that potentially correspond to multimers or intracellular membrane-bound species, cells were fixed for 15 min in 4% Formaldehyde solution and permeabilized with 0.3% of Triton X-100 for 15 min. Bars correspond to 20 μm. B2 For better preservation, and visualization of small, highly-soluble α-syn species potentially corresponding to monomers, cells were fixed for 15 min in 4% Paraformaldehyde (PFA) and permeabilized with 0.1% of Triton X-100 for 10 min. Bars correspond to 20 μm. Images are representative of n = 3 independent experiments. C Quantification of SARS-CoV-2-N-positive cells in SARS-CoV-2-infected CTR vs α-syn-treated SARS-CoV-2-infected cells. Values are expressed as the percentage of N-positive cells ± SD calculated as the number of N-positive cells/total cells per microscopic field from n = 12 microscopic fields per experimental group, from n = 3 independent experiments. Data were analyzed by applying the Student’s t-test for comparison between VIRUS CTR and VIRUS α-syn groups. **p < 0.01

Article Snippet: Type-I IFN (human recombinant IFN-β, 500 IU/ml, BEI resources) and human recombinant α-syn monomers (1 μM, Merck-Sigma), were exogenously pre-administered to cells, both alone and in combination, at 24 h pre-infection.

Techniques: Quantitative RT-PCR, Infection, Isolation, Immunofluorescence, Membrane, Preserving, Comparison, Virus

In HUVECS, absence of productive SARS-CoV-2 infection is associated with α-syn accumulation. A Quantification of SARS-CoV-2 replication in HUVECs in the absence and presence of α-syn and IFN-β (MOI 1). Results are shown as mean ± SEM from n = 4 independent experiments. Data were analyzed by applying Two-Way ANOVA. **p < 0.01, ***p < 0.001. B In the absence of productive viral replication, overall α-syn immunostaining in HUVECs is not decreased by either SARS-CoV-2 infection or the addition of exogenous α-syn monomers. Representative immunofluorescence images for SARS-CoV-2-Nucleocapsid (N), and α-syn in Mock- and SARS-CoV-2-Infected HUVECs 72 h post-infection (MOI 1), in the absence and presence of IFN-β, or exogenous α-syn monomers. Cells were fixed for 15 min in Formaldehyde solution, followed by 15 min permeabilization with 0.1% Triton X-100. Bars correspond to 20 μm. C In HUVECs, absence of productive SARS-CoV-2 infection is associated with the occurrence of juxtanuclear α-syn foci. 3D reconstruction of immunofluorescence images for SARS-CoV-2-Nucleocapsid (N), and α-syn proteins in Mock- and SARS-CoV-2-Infected HUVECs 72 h post-infection (MOI 1) shows the presence of juxtanuclear α-syn foci (arrows) which are induced by SARS-CoV-2 and potentiated by IFN-β and exogenous α-syn monomers

Journal: Biological Research

Article Title: Alpha-synuclein dynamics bridge Type-I Interferon response and SARS-CoV-2 replication in peripheral cells

doi: 10.1186/s40659-023-00482-x

Figure Lengend Snippet: In HUVECS, absence of productive SARS-CoV-2 infection is associated with α-syn accumulation. A Quantification of SARS-CoV-2 replication in HUVECs in the absence and presence of α-syn and IFN-β (MOI 1). Results are shown as mean ± SEM from n = 4 independent experiments. Data were analyzed by applying Two-Way ANOVA. **p < 0.01, ***p < 0.001. B In the absence of productive viral replication, overall α-syn immunostaining in HUVECs is not decreased by either SARS-CoV-2 infection or the addition of exogenous α-syn monomers. Representative immunofluorescence images for SARS-CoV-2-Nucleocapsid (N), and α-syn in Mock- and SARS-CoV-2-Infected HUVECs 72 h post-infection (MOI 1), in the absence and presence of IFN-β, or exogenous α-syn monomers. Cells were fixed for 15 min in Formaldehyde solution, followed by 15 min permeabilization with 0.1% Triton X-100. Bars correspond to 20 μm. C In HUVECs, absence of productive SARS-CoV-2 infection is associated with the occurrence of juxtanuclear α-syn foci. 3D reconstruction of immunofluorescence images for SARS-CoV-2-Nucleocapsid (N), and α-syn proteins in Mock- and SARS-CoV-2-Infected HUVECs 72 h post-infection (MOI 1) shows the presence of juxtanuclear α-syn foci (arrows) which are induced by SARS-CoV-2 and potentiated by IFN-β and exogenous α-syn monomers

Article Snippet: Type-I IFN (human recombinant IFN-β, 500 IU/ml, BEI resources) and human recombinant α-syn monomers (1 μM, Merck-Sigma), were exogenously pre-administered to cells, both alone and in combination, at 24 h pre-infection.

Techniques: Infection, Immunostaining, Immunofluorescence

SARS-CoV-2 synergizes with an excess of exogenously added α-syn monomers to reduce α-syn multimer:monomer ratio, which is prevented by IFN-β. A Representative western blot, and B quantification of total α-syn, α-syn monomers, α-syn multimers, and C α-syn multimer:monomer ratio in A549-hACE2 cells 48 h post- Mock and SARS-CoV-2 infection in the presence of exogenously added α-syn. Quantification was performed by normalizing to total protein (Loading control). Results show raw normalized values presented as mean ± SEM from n = 3 independent experiments. Multimer:monomer ratio is expressed as absolute value calculated as “multimers/monomers”. Data were analyzed by applying Two-Way ANOVA (for total, multimer, and monomer α-syn quantification) or One Way ANOVA (for multimer:monomer ratio). *p < 0.05, ***p < 0.001, ****p < 0.0001

Journal: Biological Research

Article Title: Alpha-synuclein dynamics bridge Type-I Interferon response and SARS-CoV-2 replication in peripheral cells

doi: 10.1186/s40659-023-00482-x

Figure Lengend Snippet: SARS-CoV-2 synergizes with an excess of exogenously added α-syn monomers to reduce α-syn multimer:monomer ratio, which is prevented by IFN-β. A Representative western blot, and B quantification of total α-syn, α-syn monomers, α-syn multimers, and C α-syn multimer:monomer ratio in A549-hACE2 cells 48 h post- Mock and SARS-CoV-2 infection in the presence of exogenously added α-syn. Quantification was performed by normalizing to total protein (Loading control). Results show raw normalized values presented as mean ± SEM from n = 3 independent experiments. Multimer:monomer ratio is expressed as absolute value calculated as “multimers/monomers”. Data were analyzed by applying Two-Way ANOVA (for total, multimer, and monomer α-syn quantification) or One Way ANOVA (for multimer:monomer ratio). *p < 0.05, ***p < 0.001, ****p < 0.0001

Article Snippet: Type-I IFN (human recombinant IFN-β, 500 IU/ml, BEI resources) and human recombinant α-syn monomers (1 μM, Merck-Sigma), were exogenously pre-administered to cells, both alone and in combination, at 24 h pre-infection.

Techniques: Western Blot, Infection

A Gene expression changes associated with the pro-viral and anti-viral effects of exogenous α-syn monomers and IFN-β, respectively . qPCR-based transcriptional analyses were performed in A549-hACE2 cells 48 h post-infection. Results are expressed as mean ± SEM from n = 3 independent experiments. Data for each gene were analyzed by One Way ANOVA. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. SNCA synclein alpha; IFNB Interferon beta; STAT1 Signal transducer and activator of transcription 1; MX1 Myxovirus Resistance Protein 1; MX2 Myxovirus Resistance Protein 2; OAS1 2'-5'-oligoadenylate synthetase 1, RIG-I retinoic acid-inducible gene I; TNFA tumor necrosis factor alpha, TNFRSF1A tumor necrosis factor receptor superfamily 1A, TLR8 Toll-like receptor 8; Toll-like receptor 9 (TLR9). B SNCA mRNA expression is negatively correlated with viral titers at 3 and 5d post-infection of epithelial lung cells . The analyses were performed in CaLu-3 and A549-hACE2 cells at 3 and 5d post-infection with SARS-CoV-2 at 3 different MOI (0.01, 0.005, 0.001). Correlation between relative SNCA mRNA expression and SARS-CoV-2 N gene mRNA expression in A549-hACE2 and CaLu-3 cells was calculated through the Pearson’s r coefficient. Statistical significance was calculated through the two-tailed, paired Student’s t test . n = 12 for A549-hACE2 cells; n = 9 for CaLu3 cells. C1 and C2 SNCA mRNA expression is increased in human PBMCs stimulated with SARS-CoV-2 and is negatively correlated with SARS-CoV-2 replication in co-cultured CaLu-3 cells. C1 qPCR-based quantification of SNCA mRNA expression in in vitro SARS-CoV-2-exposed human PBMCs compared with Mock CTR. Statistical significance was calculated through the two-tailed, paired Student’s t test . The graph shows individual values (n = 8) with mean ± SD. ** p < 0.01. C2 Correlation analysis between relative SNCA mRNA expression within SARS-CoV-2-exposed PBMCs, and SARS-CoV-2 replication (N gene copy number) in co-cultured CaLu-3 epithelial cells. Correlation was calculated through the Pearson’s r coefficient. Statistical significance was calculated through the two-tailed, paired Student’s t test. n = 8

Journal: Biological Research

Article Title: Alpha-synuclein dynamics bridge Type-I Interferon response and SARS-CoV-2 replication in peripheral cells

doi: 10.1186/s40659-023-00482-x

Figure Lengend Snippet: A Gene expression changes associated with the pro-viral and anti-viral effects of exogenous α-syn monomers and IFN-β, respectively . qPCR-based transcriptional analyses were performed in A549-hACE2 cells 48 h post-infection. Results are expressed as mean ± SEM from n = 3 independent experiments. Data for each gene were analyzed by One Way ANOVA. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. SNCA synclein alpha; IFNB Interferon beta; STAT1 Signal transducer and activator of transcription 1; MX1 Myxovirus Resistance Protein 1; MX2 Myxovirus Resistance Protein 2; OAS1 2'-5'-oligoadenylate synthetase 1, RIG-I retinoic acid-inducible gene I; TNFA tumor necrosis factor alpha, TNFRSF1A tumor necrosis factor receptor superfamily 1A, TLR8 Toll-like receptor 8; Toll-like receptor 9 (TLR9). B SNCA mRNA expression is negatively correlated with viral titers at 3 and 5d post-infection of epithelial lung cells . The analyses were performed in CaLu-3 and A549-hACE2 cells at 3 and 5d post-infection with SARS-CoV-2 at 3 different MOI (0.01, 0.005, 0.001). Correlation between relative SNCA mRNA expression and SARS-CoV-2 N gene mRNA expression in A549-hACE2 and CaLu-3 cells was calculated through the Pearson’s r coefficient. Statistical significance was calculated through the two-tailed, paired Student’s t test . n = 12 for A549-hACE2 cells; n = 9 for CaLu3 cells. C1 and C2 SNCA mRNA expression is increased in human PBMCs stimulated with SARS-CoV-2 and is negatively correlated with SARS-CoV-2 replication in co-cultured CaLu-3 cells. C1 qPCR-based quantification of SNCA mRNA expression in in vitro SARS-CoV-2-exposed human PBMCs compared with Mock CTR. Statistical significance was calculated through the two-tailed, paired Student’s t test . The graph shows individual values (n = 8) with mean ± SD. ** p < 0.01. C2 Correlation analysis between relative SNCA mRNA expression within SARS-CoV-2-exposed PBMCs, and SARS-CoV-2 replication (N gene copy number) in co-cultured CaLu-3 epithelial cells. Correlation was calculated through the Pearson’s r coefficient. Statistical significance was calculated through the two-tailed, paired Student’s t test. n = 8

Article Snippet: Type-I IFN (human recombinant IFN-β, 500 IU/ml, BEI resources) and human recombinant α-syn monomers (1 μM, Merck-Sigma), were exogenously pre-administered to cells, both alone and in combination, at 24 h pre-infection.

Techniques: Expressing, Infection, Two Tailed Test, Cell Culture, In Vitro

Summary cartoon depicting the opposite effects of extracellular α-syn monomers and IFN-β upon cellular internalization, and related α-syn multimer:monomer dynamics during SARS-CoV-2 infection of susceptible epithelial lung cells

Journal: Biological Research

Article Title: Alpha-synuclein dynamics bridge Type-I Interferon response and SARS-CoV-2 replication in peripheral cells

doi: 10.1186/s40659-023-00482-x

Figure Lengend Snippet: Summary cartoon depicting the opposite effects of extracellular α-syn monomers and IFN-β upon cellular internalization, and related α-syn multimer:monomer dynamics during SARS-CoV-2 infection of susceptible epithelial lung cells

Article Snippet: Type-I IFN (human recombinant IFN-β, 500 IU/ml, BEI resources) and human recombinant α-syn monomers (1 μM, Merck-Sigma), were exogenously pre-administered to cells, both alone and in combination, at 24 h pre-infection.

Techniques: Infection