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  • 99
    Millipore elisa kit
    Effect of Imm124-E on serum levels of IL-6, <t>GLP-1,</t> and adiponectin. All sera were measured using <t>ELISA</t> kits at days 1 and 30 of the trial in all treated patients. Graphs show ( A ) Serum levels of IL-6 from six patients; ( B ) Serum levels of GLP-1 post–glucose tolerance test from five patients; ( C ) Serum levels of adiponectin from eight patients. Note: Graphs indicate means ± SD. Abbreviations: IL-6, interleukin-6; GLP-1, glucagon-like peptide 1; ELISA, enzyme-linked immunosorbent assay; SD, standard deviation.
    Elisa Kit, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2068 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/elisa kit/product/Millipore
    Average 99 stars, based on 2068 article reviews
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    95
    Thermo Fisher elisa kits
    Antibody responses, virus titers, and inflammatory cytokine levels in BALF and lungs after challenge. Levels of IgG antibodies were determined in samples collected from mice at day 5 p.i. with A/Philippines/2/82 (H3N2) virus ( n = 5). (A) IgG antibody levels in BALF. The antibody level was determined by <t>ELISA</t> using M2e peptide as a coating antigen. Lung viral titers (B), IFN-γ (C), and IL-6 (D) <t>cytokines</t> in BALF were determined at day 5 p.i. Lung viral titers were determined by an egg infection assay. IFN-γ and IL-6 were determined by a cytokine ELISA. Data represent mean ± SEM. Statistical significances were determined by 1-way ANOVA. Asterisks indicate significant differences (* p
    Elisa Kits, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 11232 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Becton Dickinson elisa kits
    Effects of ASWP and three ASEPs on cell viability; phagocytic activity; and the production of nitric oxide (NO), tumor necrosis factor <t>(TNF)-α,</t> interleukin (IL)-6, and IL-12 in RAW264.7 macrophages. ASWP was obtained from Aster scaber using hot water extraction and ASEP-A, ASEP-C, and ASEP-P were obtained from Aster scaber using α-amylase, cellulase, and pectinase, respectively, in an enzyme-assisted extraction. RAW264.7 macrophages were stimulated with the polysaccharide samples at 1, 10, or 100 μg/mL. LPS (1 μg/mL) was used as the positive control (PC). ( A ) Cell viability was determined by the cell counting kit-8 (CCK-8) assay; ( B ) Phagocytic activity was determined using a phagocytosis assay; ( C ) NO levels in the culture media were determined by measuring nitrite accumulation, and the secretion levels of TNF-α, IL-6, and IL-12 were measured by <t>ELISA.</t> ( D ) mRNA expression levels of iNOS, TNF-α, IL-6, and IL-12 in macrophages were quantified by qRT-PCR analysis. Lowercase letters (a–c) indicate significant differences ( p
    Elisa Kits, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 94/100, based on 3147 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Abcam elisa kit
    qRT-PCR and <t>ELISA</t> testing of expression in sheep monocytes with LPS stimulation. A, B and C show the expression of TLR4 , Myd88 and NF-κB , at 0min, 5 min, 30 min and 60 min after LPS (100 ng/mL) stimulation, respectively. D, E and F show the phosphorylation levels of <t>p38</t> , JNK , and ERK signaling at 0min, 5 min, 30 min and 60 min after LPS (100 ng/mL) stimulation, respectively. Tg: transgenic sheep; NTg: non-transgenic sheep. All data are presented as the mean ± SEM from three experiments, *P
    Elisa Kit, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 1611 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Cayman Chemical elisa kit
    Acetylation and expression of StAR in response to HDAC inhibitors, and their correlation to <t>E2</t> synthesis in MCF7 cells. Cells (1 × 10 6 per dish) were plated in 100-mm dishes 24h before treatments. Cells were then treated without or with panobinostat (10nM) for 0-180min (A-C) or other HDAC inhibitors for 45min (D-F). Following treatments, cells were collected, extracted with lysis buffer, and processed for either immunoprecipitation or immunoblotting studies, as described in Materials and methods . Representative immunoblots illustrate acetylation (Ac-StAR) and expression of StAR (T-StAR) in different treatment groups. IgG heavy chain (IgG-Hc) and β-actin expression were assessed for loading controls in immunoprecipitation and immunoblotting, respectively. Integrated optical density (IOD) values of Ac-StAR and T-StAR in each band were quantified and normalized with corresponding IgG-Hc and β-actin expression, and presented as fold response (C,F). E2 levels in media at each time point were determined by <t>ELISA</t> and presented as pg/mg protein (C). Data are representative of 3-4 independent experiments. *, p
    Elisa Kit, supplied by Cayman Chemical, used in various techniques. Bioz Stars score: 94/100, based on 1503 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Crystal Chem Inc ultra sensitive mouse insulin elisa kit
    Impaired insulin secretion from isolated Pick1 cKO islets. (a) <t>GSIS</t> was performed on isolated islets stimulated with 4.8 or 30 mM KCl or 2.8 or 16.7 mM glucose (Glc). (b) The stimulated index was also calculated following KCl or Glc treatment. (c) Insulin content in the isolated islets was measured by <t>ELISA.</t> (d) Isolated WT and Pick1 cKO islets were incubated in the presence of 16.7 mM Glc at 2-min intervals for 20 min. (e) Same data as shown in d, but plotted as release per 2 min. (f, g) Secreted insulin was tested from isolated WT and Pick1 cKO islets incubated with or without E4. Data are represented as mean ± SEM, n = 3, three to four groups per type of mouse, 10–0 islets per group. * p
    Ultra Sensitive Mouse Insulin Elisa Kit, supplied by Crystal Chem Inc, used in various techniques. Bioz Stars score: 93/100, based on 1190 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ultra sensitive mouse insulin elisa kit/product/Crystal Chem Inc
    Average 93 stars, based on 1190 article reviews
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    93
    Millipore elisa kits
    Effect of EA on <t>Glu</t> content and expression of NMDAR2A and NMDAR2B protein. (A) Glu content in the CA1 area of the hippocampus in each group, measured by <t>ELISA.</t> (B) Protein expression of NMDAR2A and NMDAR2B in the CA1 area of the hippocampus in each group, examined by Western blotting. (C,D) Quantitative indices of protein expression of NR2A and NR2B, derived from figure 2B . #P
    Elisa Kits, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 1417 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Enzo Biochem elisa kit
    Regulation of <t>TNF-</t> α production in RAW 264.7 cells. Cells were treated with the steam (a) or 50% ethanol (b) GE at various concentrations for 24 h. The culture media were collected and cytokine concentrations were measured by <t>ELISA.</t> The data were expressed as mean ± SEM. ∗ p
    Elisa Kit, supplied by Enzo Biochem, used in various techniques. Bioz Stars score: 93/100, based on 841 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    USCN Life elisa kit
    Moxibustion modulated the hypothalamic-pituitary-adrenal axis of CFS rats. Data are presented as the mean ± SEM ( n = 8 per group; one-way analysis of variance followed by the least significant difference post hoc test). (A) CRH mRNA expression in the hypothalamus was semi-quantified by real-time PCR. (B, C) ACTH (B) and <t>CORT</t> (C) concentration in plasma was detected by enzyme-linked immunosorbent assay. ** P
    Elisa Kit, supplied by USCN Life, used in various techniques. Bioz Stars score: 94/100, based on 1303 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    ALPCO elisa kit
    Impaired insulin secretion and <t>proinsulin</t> maturation in XBP1-deficient β-cells. ( A ) Glucose-stimulated insulin secretion assays. Serum insulin levels were measured in control heterozygous Xbp1 f/+ ;RIP-cre and Xbp1 f/f ;RIP-cre mice at indicated time points after a bolus glucose injection. Each line represents an individual mouse. ( B ) Min6 cells stably expressing shRNAs targeting control luciferase or XBP1 mRNAs were tested for XBP1s expression by Western blot. ( C ) Cells were pretreated with 2.5 mM glucose for 2 h and then cultured in 5 mM or 25 mM glucose media for 2 h. Culture media were collected to measure insulin content by <t>ELISA.</t> ( D ) Serum proinsulin levels relative to total insulin in WT and Xbp1 f/f ;RIP-cre mice were determined. Data from both males and females were combined, because sex difference was not significant. The graphs display proinsulin:insulin ratio ( Left ) or proinsulin and insulin concentrations (pmol per liter) ( Right ). Each dot represents an individual mouse (4- to 6-mo-old). ( E ) Min6 cells expressing control or XBP1 shRNA were pulse-labeled for 30 min with [ 35 S]Met/Cys and then cultured in chase media for the indicated time. Cells and culture supernatants were harvested for immunoprecipitation of radiolabeled insulin and proinsulin, which were revealed by SDS/PAGE followed by autoradiography.
    Elisa Kit, supplied by ALPCO, used in various techniques. Bioz Stars score: 94/100, based on 904 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    MyBiosource elisa kit
    Effects of the <t>β2-GPI</t> DNA vaccine and FK506 treatment on β2-GPI-specific Treg cell response. Spleen cells were purified from the different mouse groups on day 56 and stimulated with recombinant β2-GPI protein (10 μg/mL). After 96 h, (A) the percentage of Foxp3-expressing CD4+ T cells was determined by flow cytometry. The dot plot shows data from one representative mouse from each group. The bar graph represents the mean ± standard deviation (SD) of six mice from three independent experiments. (B) The culture supernatants were collected, and IL-10 and TGF-β production was analyzed in triplicate by <t>ELISA.</t> The data are presented as the means ± SD of triplicate assays from six mice/group. ns p > 0.05, **p
    Elisa Kit, supplied by MyBiosource, used in various techniques. Bioz Stars score: 94/100, based on 932 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Cloud-Clone elisa kit
    IL-25 is highly expressed in HCC patients. a The level of <t>NMU</t> were detected by <t>ELISA</t> in serum of hepatic hemangioma patients ( n = 10) and HCC patients ( n = 10). b Immunohistochemistry (IHC) staining was performed in a tissue microarray consisted of 228 HCC peri- and intra-tumor tissue, NMU representative IHC images are shown. Bar, 20 μm. ** p
    Elisa Kit, supplied by Cloud-Clone, used in various techniques. Bioz Stars score: 94/100, based on 860 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/elisa kit/product/Cloud-Clone
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    94
    IDEXX elisa kit
    Correlation between <t>cELISA</t> and the HI test and the commercial <t>ELISA</t> kit (IDEXX). a Serum titers for antibodies against NDV detected between the cELISA (PI) and HI test (log2). b Titres of antibodies against NDV in different dilution serum samples detected between the cELISA (PI) and commercial ELISA kit test (S/P)
    Elisa Kit, supplied by IDEXX, used in various techniques. Bioz Stars score: 94/100, based on 814 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/elisa kit/product/IDEXX
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    93
    Thermo Fisher amyloid beta 42 human elisa kit
    Correlation between <t>cELISA</t> and the HI test and the commercial <t>ELISA</t> kit (IDEXX). a Serum titers for antibodies against NDV detected between the cELISA (PI) and HI test (log2). b Titres of antibodies against NDV in different dilution serum samples detected between the cELISA (PI) and commercial ELISA kit test (S/P)
    Amyloid Beta 42 Human Elisa Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 793 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/amyloid beta 42 human elisa kit/product/Thermo Fisher
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    93
    Crystal Chem Inc elisa kit
    EREG stimulates leptin secretion and lipolysis in mice with diet-induced obesity (DIO). (A) DIO WT male mice (n=7/ group) were injected with 100μL PBS (Veh) with and without EREG (1.5ng/g body weight or 20ng/per <t>iAb</t> depot) into both epididymal iAb fat pads every other day for 2 weeks. Total mRNA was isolated from one whole iAb fad pad. Expression of Lep was measured using NanoString assay. Normalized data represent mean±SD, n=5; Mann-Whitney U test. (B) Expression of LEP protein levels were measured in plasma in the same mice group by <t>ELISA.</t> Data (mean±SD, n=5/group). Independent Student’s t -test. (C) Non-esterified fatty acids (NEFA) and (D) TG concentrations in plasma are shown as mean ± SD, n=5. Independent Student’s t -test.
    Elisa Kit, supplied by Crystal Chem Inc, used in various techniques. Bioz Stars score: 93/100, based on 585 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/elisa kit/product/Crystal Chem Inc
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    99
    Cell Signaling Technology Inc β actin
    Western blot analysis of the effect of Sanguinarine on CEM/ADR5000 leukemia cells. Evaluation of the P-gp, NFκB, and IκBα expressions. <t>β-actin</t> was used as loading control. Bands were normalized to β-actin in order to obtain numerical values (Mean ± SD ).
    β Actin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 38293 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β actin/product/Cell Signaling Technology Inc
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    94
    Phoenix Pharmaceuticals elisa kit
    Enho mRNA and protein levels are downregulated in the aged rat brain and are associated with decreased <t>adropin</t> levels in plasma A) Enho mRNA expression was significantly decreased in the cerebral cortex of aged rats compared to young rats. B) Cortical homogenates (50 µg of total protein) from rat brains were subjected to SDS-polyacrylamide gel electrophoresis and endogenous adropin protein levels were detected by Western blot. Representative immunoblots and densitometric data showed that adropin protein levels in the cerebral cortex were dramatically decreased in aged rats compared to young rats. C) Adropin antibody is highly specific. Cortical brain homogenates (samples 1 and 2) were denatured at 70°C for 10 min in 2X Laemmli sample buffer containing 4% β-mercaptoethanol, separated on 4-20% SDS-polyacrylamide gels, and then transferred onto nitrocellulose membranes. Before incubation with the membrane, the adropin monoclonal antibody was pre-absorbed with different concentrations of synthetic adropin 34-76 peptide (0-300 ng/ml), which resulted in a concentration-dependent loss of the intensity of the band corresponding to endogenous brain adropin. D) Plasma adropin levels were measured by a commercial adropin <t>ELISA</t> kit. A significant decrease in adropin levels was observed in aged rats compared with young controls. * P
    Elisa Kit, supplied by Phoenix Pharmaceuticals, used in various techniques. Bioz Stars score: 94/100, based on 452 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Effect of Imm124-E on serum levels of IL-6, GLP-1, and adiponectin. All sera were measured using ELISA kits at days 1 and 30 of the trial in all treated patients. Graphs show ( A ) Serum levels of IL-6 from six patients; ( B ) Serum levels of GLP-1 post–glucose tolerance test from five patients; ( C ) Serum levels of adiponectin from eight patients. Note: Graphs indicate means ± SD. Abbreviations: IL-6, interleukin-6; GLP-1, glucagon-like peptide 1; ELISA, enzyme-linked immunosorbent assay; SD, standard deviation.

    Journal: Journal of Inflammation Research

    Article Title: Alleviation of insulin resistance and liver damage by oral administration of Imm124-E is mediated by increased Tregs and associated with increased serum GLP-1 and adiponectin: results of a phase I/II clinical trial in NASH

    doi: 10.2147/JIR.S35227

    Figure Lengend Snippet: Effect of Imm124-E on serum levels of IL-6, GLP-1, and adiponectin. All sera were measured using ELISA kits at days 1 and 30 of the trial in all treated patients. Graphs show ( A ) Serum levels of IL-6 from six patients; ( B ) Serum levels of GLP-1 post–glucose tolerance test from five patients; ( C ) Serum levels of adiponectin from eight patients. Note: Graphs indicate means ± SD. Abbreviations: IL-6, interleukin-6; GLP-1, glucagon-like peptide 1; ELISA, enzyme-linked immunosorbent assay; SD, standard deviation.

    Article Snippet: The circulating level of human GLP-1 was quantified using a commercial ELISA kit from Millipore Corp (Billerica, MA) according to the manufacturer’s instructions.

    Techniques: Enzyme-linked Immunosorbent Assay, Standard Deviation

    Antibody responses, virus titers, and inflammatory cytokine levels in BALF and lungs after challenge. Levels of IgG antibodies were determined in samples collected from mice at day 5 p.i. with A/Philippines/2/82 (H3N2) virus ( n = 5). (A) IgG antibody levels in BALF. The antibody level was determined by ELISA using M2e peptide as a coating antigen. Lung viral titers (B), IFN-γ (C), and IL-6 (D) cytokines in BALF were determined at day 5 p.i. Lung viral titers were determined by an egg infection assay. IFN-γ and IL-6 were determined by a cytokine ELISA. Data represent mean ± SEM. Statistical significances were determined by 1-way ANOVA. Asterisks indicate significant differences (* p

    Journal: PLoS ONE

    Article Title: Cross Protection against Influenza A Virus by Yeast-Expressed Heterologous Tandem Repeat M2 Extracellular Proteins

    doi: 10.1371/journal.pone.0137822

    Figure Lengend Snippet: Antibody responses, virus titers, and inflammatory cytokine levels in BALF and lungs after challenge. Levels of IgG antibodies were determined in samples collected from mice at day 5 p.i. with A/Philippines/2/82 (H3N2) virus ( n = 5). (A) IgG antibody levels in BALF. The antibody level was determined by ELISA using M2e peptide as a coating antigen. Lung viral titers (B), IFN-γ (C), and IL-6 (D) cytokines in BALF were determined at day 5 p.i. Lung viral titers were determined by an egg infection assay. IFN-γ and IL-6 were determined by a cytokine ELISA. Data represent mean ± SEM. Statistical significances were determined by 1-way ANOVA. Asterisks indicate significant differences (* p

    Article Snippet: Cytokines in BALF were assayed with ELISA kits in duplicate against a standard curve according to the manufacturers’ instructions (eBioscience, San Diego, CA).

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay, Infection

    Effect of IFN-γ cell preconditioning (1 ng/ml for 24 h) singly and with HRV-14 or HRV-1b infection (TCID 50 10 2.5 for 90 mins) on sICAM-1 protein release (A) and sICAM-1 gene expression (B) in NHBE cells over the study period . sICAM-1 in cell culture supernatants was assayed using ELISA. Data are mean ± S.E. of three separate experiments (Fig 5A,*p

    Journal: Journal of Inflammation (London, England)

    Article Title: IFN-gamma regulation of ICAM-1 receptors in bronchial epithelial cells: soluble ICAM-1 release inhibits human rhinovirus infection

    doi: 10.1186/1476-9255-5-8

    Figure Lengend Snippet: Effect of IFN-γ cell preconditioning (1 ng/ml for 24 h) singly and with HRV-14 or HRV-1b infection (TCID 50 10 2.5 for 90 mins) on sICAM-1 protein release (A) and sICAM-1 gene expression (B) in NHBE cells over the study period . sICAM-1 in cell culture supernatants was assayed using ELISA. Data are mean ± S.E. of three separate experiments (Fig 5A,*p

    Article Snippet: sICAM-1 protein ELISA Soluble ICAM-1 assays were performed with a commercially available ELISA kit (BioSource International, California USA).

    Techniques: Infection, Expressing, Cell Culture, Enzyme-linked Immunosorbent Assay

    Effect of protease inhibitors on sICAM-1 release (A) and mICAM-1 levels (B) . Cell cultures were pre-incubated with 1 complete™ mini tablet/10 ml of media; sICAM-1 in associated supernatants was assayed using ELISA. Data are mean ± S.E. of three separate experiments (Fig. 7A, *p

    Journal: Journal of Inflammation (London, England)

    Article Title: IFN-gamma regulation of ICAM-1 receptors in bronchial epithelial cells: soluble ICAM-1 release inhibits human rhinovirus infection

    doi: 10.1186/1476-9255-5-8

    Figure Lengend Snippet: Effect of protease inhibitors on sICAM-1 release (A) and mICAM-1 levels (B) . Cell cultures were pre-incubated with 1 complete™ mini tablet/10 ml of media; sICAM-1 in associated supernatants was assayed using ELISA. Data are mean ± S.E. of three separate experiments (Fig. 7A, *p

    Article Snippet: sICAM-1 protein ELISA Soluble ICAM-1 assays were performed with a commercially available ELISA kit (BioSource International, California USA).

    Techniques: Incubation, Enzyme-linked Immunosorbent Assay

    Interleukin-6 concentration in wild-type MEFs and ATF3 KO cells. Wild-type MEFs and ATF3 KO cells were treated with 1, 2, 4, or 8 µM doxorubicin, for 24 h. The concentration of interleukin-6 (IL-6) in the cell culture supernatant was measured using an enzyme-linked immunosorbent assay kit. The IL-6 concentration was calculated using a standard curve. Experiments were independently performed 5 times. **P

    Journal: PLoS ONE

    Article Title: Doxorubicin Induces Cytotoxicity through Upregulation of pERK-Dependent ATF3

    doi: 10.1371/journal.pone.0044990

    Figure Lengend Snippet: Interleukin-6 concentration in wild-type MEFs and ATF3 KO cells. Wild-type MEFs and ATF3 KO cells were treated with 1, 2, 4, or 8 µM doxorubicin, for 24 h. The concentration of interleukin-6 (IL-6) in the cell culture supernatant was measured using an enzyme-linked immunosorbent assay kit. The IL-6 concentration was calculated using a standard curve. Experiments were independently performed 5 times. **P

    Article Snippet: Cytokine Analysis The concentration of each cytokine in the supernatant of the culture media and serum was determined using commercially available enzyme-linked immunosorbent assay (ELISA) kits (eBioscience).

    Techniques: Concentration Assay, Cell Culture, Enzyme-linked Immunosorbent Assay

    Pam2CSK 4 (P2C) and Pam3CSK 4 (P3C)‐stimulated responses of CD14 + monocytes. CD14 + CD16 – monocytes were derived by negative selection from the whole blood of multiple sclerosis (MS) patients (15 patients;16 total responses) and healthy controls (13 individuals; 15 total responses), cultured at 1 × 10 5 /ml (0·2ml/well), stimulated with 2 μg/ml of P2C or P3C for 4 h, and tumour necrosis factor (TNF)‐α in the culture supernatants measured by enzyme‐linked immunosorbent assay (ELISA). The level of TNF‐α representing the upper threshold of control responses [based on the control interquartile range (IQR)] is depicted by the dashed line. The percentage of responses above the threshold is depicted for controls and total MS patients. Medians are depicted. (a) Red icons represent P2C stimulation and black icons represent P3C stimulation; (b) same results as in (a), but green icons represent patients with progressive MS. The mean TNF‐α level for the control cohort was 104 pg/ml and the mean TNF‐α level for the total MS cohort was 247 pg/ml; P = 0·0712 via Mann–Whitney. The percentage of responses above threshold: 57% of Prog MS patients, 33% of relapsing–remitting MS (RRMS) patients.

    Journal: Clinical and Experimental Immunology

    Article Title: Enhanced TLR2 responses in multiple sclerosis

    doi: 10.1111/cei.13150

    Figure Lengend Snippet: Pam2CSK 4 (P2C) and Pam3CSK 4 (P3C)‐stimulated responses of CD14 + monocytes. CD14 + CD16 – monocytes were derived by negative selection from the whole blood of multiple sclerosis (MS) patients (15 patients;16 total responses) and healthy controls (13 individuals; 15 total responses), cultured at 1 × 10 5 /ml (0·2ml/well), stimulated with 2 μg/ml of P2C or P3C for 4 h, and tumour necrosis factor (TNF)‐α in the culture supernatants measured by enzyme‐linked immunosorbent assay (ELISA). The level of TNF‐α representing the upper threshold of control responses [based on the control interquartile range (IQR)] is depicted by the dashed line. The percentage of responses above the threshold is depicted for controls and total MS patients. Medians are depicted. (a) Red icons represent P2C stimulation and black icons represent P3C stimulation; (b) same results as in (a), but green icons represent patients with progressive MS. The mean TNF‐α level for the control cohort was 104 pg/ml and the mean TNF‐α level for the total MS cohort was 247 pg/ml; P = 0·0712 via Mann–Whitney. The percentage of responses above threshold: 57% of Prog MS patients, 33% of relapsing–remitting MS (RRMS) patients.

    Article Snippet: Human tumour necrosis factor (TNF)‐α was measured in the supernatants using the Ready‐SET‐Go!® enzyme‐linked immunosorbent assay (ELISA) kit from eBiosciences/Affymetrix Inc. (Santa Clara, CA, USA).

    Techniques: Derivative Assay, Selection, Mass Spectrometry, Cell Culture, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

    Effects of ASWP and three ASEPs on cell viability; phagocytic activity; and the production of nitric oxide (NO), tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-12 in RAW264.7 macrophages. ASWP was obtained from Aster scaber using hot water extraction and ASEP-A, ASEP-C, and ASEP-P were obtained from Aster scaber using α-amylase, cellulase, and pectinase, respectively, in an enzyme-assisted extraction. RAW264.7 macrophages were stimulated with the polysaccharide samples at 1, 10, or 100 μg/mL. LPS (1 μg/mL) was used as the positive control (PC). ( A ) Cell viability was determined by the cell counting kit-8 (CCK-8) assay; ( B ) Phagocytic activity was determined using a phagocytosis assay; ( C ) NO levels in the culture media were determined by measuring nitrite accumulation, and the secretion levels of TNF-α, IL-6, and IL-12 were measured by ELISA. ( D ) mRNA expression levels of iNOS, TNF-α, IL-6, and IL-12 in macrophages were quantified by qRT-PCR analysis. Lowercase letters (a–c) indicate significant differences ( p

    Journal: International Journal of Molecular Sciences

    Article Title: The Effect of Pectinase-Assisted Extraction on the Physicochemical and Biological Properties of Polysaccharides from Aster scaber

    doi: 10.3390/ijms19092839

    Figure Lengend Snippet: Effects of ASWP and three ASEPs on cell viability; phagocytic activity; and the production of nitric oxide (NO), tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-12 in RAW264.7 macrophages. ASWP was obtained from Aster scaber using hot water extraction and ASEP-A, ASEP-C, and ASEP-P were obtained from Aster scaber using α-amylase, cellulase, and pectinase, respectively, in an enzyme-assisted extraction. RAW264.7 macrophages were stimulated with the polysaccharide samples at 1, 10, or 100 μg/mL. LPS (1 μg/mL) was used as the positive control (PC). ( A ) Cell viability was determined by the cell counting kit-8 (CCK-8) assay; ( B ) Phagocytic activity was determined using a phagocytosis assay; ( C ) NO levels in the culture media were determined by measuring nitrite accumulation, and the secretion levels of TNF-α, IL-6, and IL-12 were measured by ELISA. ( D ) mRNA expression levels of iNOS, TNF-α, IL-6, and IL-12 in macrophages were quantified by qRT-PCR analysis. Lowercase letters (a–c) indicate significant differences ( p

    Article Snippet: Meanwhile, the secretion of TNF-α, IL-6, and IL-12 from the macrophages were assessed using corresponding ELISA kits (BD Biosciences, Pharmingen, San Diego, CA, USA).

    Techniques: Activity Assay, Positive Control, Cell Counting, CCK-8 Assay, Phagocytosis Assay, Enzyme-linked Immunosorbent Assay, Expressing, Quantitative RT-PCR

    UPS13 is essential for Paxillin-mediated NLRP3 deubiquitination upon ATP treatment. A HEK293T cells were co-transfected with pHA-Paxillin and pFlag-BRCC3, pFlag-EIF3S5, pFlag-UPS13 or pFlag-OTUB1. Lysates were prepared and subjected to IP (top) or subjected to Western blot (as input) (bottom). B Hela cells stably expressing sh-UPS13 or sh-EIF3S5 were generated and analyzed. C The stable Hela cells were co-transfected with pFlag-NLRP3, pHA-UB, or pMyc-Paxillin. Lysates were prepared and subjected to denature-IP (top) or subjected to Western blot (bottom). D, E HEK293T cells were transfected with pFlag-UPS13 and pHA-Paxillin. F, G HEK293T cells were transfected with pFlag-UPS13 and pHA-NLRP3. Lysates were prepared and subjected to IP (top) or subjected to Western blot (bottom) (D–G). H HEK293T cells were co-transfected with pFlag-UPS13 and pHA-NLRP3 or pHA-NLRP3 mutants. I The diagrams of UPS13 and its mutants, as indicated. J HEK293T cells were co-transfected with pHA-NLRP3 and pFlag-UPS13 or pFlag-UPS13 mutants. K HEK293T cells were co-transfected with pHA-Paxillin and pFlag-UPS13 or pFlag-UPS13 mutants. L HEK293T cells were co-transfected with pFlag-UPS13 and pHA-Paxillin or pHA-Paxillin mutants. Lysates were prepared and subjected to IP (top) or subjected to Western blot (bottom) (H, j –l). M HEK293T cells stably expressing sh-UPS13 were generated. The stable cells were co-transfected with pFlag-NLRP3 and pHA-UB. N The stable HEK293T cells were co-transfected with pFlag-NLRP3, or pHA-UB with or without Spautin-1 (20 μM) for 6 h. O The stable HEK293T cells wre co-transfected with pFlag-NLRP3, pHA-UB, or pMyc-Paxillin with or without Spautin-1 (20 μM) for 6 h. Lysates were prepared and subjected to denature-IP (top) or subjected to Western blot (bottom) (m–o). P , Q TPA-differentiated THP-1 macrophages were treated with ATP (5 mM) for 2 h with/without Spautin-1 (5, 10, and 20 μM). IL-1β in supernatants was determined by ELISA (P). Mature IL-1β(p17) and cleaved Caspase-1(p22/p20) in supernatants and Paxillin, pro-IL-1β, and pro-Casp-1 in lysates were determined by Western blot (Q). Data shown are means ± SEMs; ***p

    Journal: bioRxiv

    Article Title: Paxillin Promotes ATP-induced Activation of P2X7 Receptor and NLRP3 Inflammasome

    doi: 10.1101/2020.04.03.023721

    Figure Lengend Snippet: UPS13 is essential for Paxillin-mediated NLRP3 deubiquitination upon ATP treatment. A HEK293T cells were co-transfected with pHA-Paxillin and pFlag-BRCC3, pFlag-EIF3S5, pFlag-UPS13 or pFlag-OTUB1. Lysates were prepared and subjected to IP (top) or subjected to Western blot (as input) (bottom). B Hela cells stably expressing sh-UPS13 or sh-EIF3S5 were generated and analyzed. C The stable Hela cells were co-transfected with pFlag-NLRP3, pHA-UB, or pMyc-Paxillin. Lysates were prepared and subjected to denature-IP (top) or subjected to Western blot (bottom). D, E HEK293T cells were transfected with pFlag-UPS13 and pHA-Paxillin. F, G HEK293T cells were transfected with pFlag-UPS13 and pHA-NLRP3. Lysates were prepared and subjected to IP (top) or subjected to Western blot (bottom) (D–G). H HEK293T cells were co-transfected with pFlag-UPS13 and pHA-NLRP3 or pHA-NLRP3 mutants. I The diagrams of UPS13 and its mutants, as indicated. J HEK293T cells were co-transfected with pHA-NLRP3 and pFlag-UPS13 or pFlag-UPS13 mutants. K HEK293T cells were co-transfected with pHA-Paxillin and pFlag-UPS13 or pFlag-UPS13 mutants. L HEK293T cells were co-transfected with pFlag-UPS13 and pHA-Paxillin or pHA-Paxillin mutants. Lysates were prepared and subjected to IP (top) or subjected to Western blot (bottom) (H, j –l). M HEK293T cells stably expressing sh-UPS13 were generated. The stable cells were co-transfected with pFlag-NLRP3 and pHA-UB. N The stable HEK293T cells were co-transfected with pFlag-NLRP3, or pHA-UB with or without Spautin-1 (20 μM) for 6 h. O The stable HEK293T cells wre co-transfected with pFlag-NLRP3, pHA-UB, or pMyc-Paxillin with or without Spautin-1 (20 μM) for 6 h. Lysates were prepared and subjected to denature-IP (top) or subjected to Western blot (bottom) (m–o). P , Q TPA-differentiated THP-1 macrophages were treated with ATP (5 mM) for 2 h with/without Spautin-1 (5, 10, and 20 μM). IL-1β in supernatants was determined by ELISA (P). Mature IL-1β(p17) and cleaved Caspase-1(p22/p20) in supernatants and Paxillin, pro-IL-1β, and pro-Casp-1 in lysates were determined by Western blot (Q). Data shown are means ± SEMs; ***p

    Article Snippet: Enzyme-linked immunosorbent assay (ELISA) The concentrations of human IL-1β in culture supernatants were measured by ELISA kit (BD Biosciences, San Jose, CA, USA).

    Techniques: Transfection, Western Blot, Stable Transfection, Expressing, Generated, Enzyme-linked Immunosorbent Assay

    ATP promotes Paxillin phosphorylation and Paxillin-NLRP3 interaction. A–D THP-1 cells stably expressing control lentivirus or Paxillin lentivirus were generated and differentiated to macrophages, which were then treated with ATP (5 mM) or Nigericin (2 μM) for 2 h (A, B), treated with DMSO, YVAD (50 μM), Glybenclamide (25 μg/ml), A438079 (100 μM), AZ10606120 (100 μM), and BAPTA-AM (30 μM) for 1 h before the treatment with ATP (5 mM) for 2 h (C, D). IL-1β levels in supernatants were determined by ELISA (A, C). Mature IL-1β (p17) and cleaved Casp-1 (p22/p20) in supernatants or Paxillin, pro-IL-1β, and pro-Casp-1 in lysates were determined by Western blot (B, D). E TPA-differentiated THP-1 macrophages were treated with DMSO or ATP (5 mM) for 2 h. Lysates were prepared and subjected to IP (top) or subjected to Western blot (as input) (bottom). F BMDM cells prepared from C57BL/6 mice bone marrow were treated with LPS (1 μg/ml) for 6 h, and the primed BMDMs were stimulated by DMSO or ATP (5 mM) for 30 min. Lysates were prepared and subjected to IP (top) or subjected to Western blot (as input) (bottom). G, H TPA-differentiated THP-1 macrophages were treated with ATP (5 mM) for 30, 60, and 120 min (G). LPS-primed BMDMs were stimulated by ATP (5 mM) for 5, 15, and 30 min (H). The protein level of p-Paxillin(Y 118), p-Paxillin(Y31), Paxillin, and GAPDH was determined by Western blot. I HEK293T cells were co-transfected with pFlag-NLRP3 and pHA-Vector, pHA-Paxillin, pHA-Paxillin-(Y31A), and pHA-Paxillin-(Y118A). Lysates were prepared and subjected to IP (top) or subjected to Western blot (as input) (bottom). J TPA-differentiated THP-1 macrophages were treated with DMSO, A438079 (100 μM), AZ10606120 (100 μM) for 1 h before the treatment with ATP (5 mM) for 2 h. The protein level of p-Paxillin(Y118), Paxillin, and GAPDH was determined by Western blot. Data shown are means ± SEMs; ***p

    Journal: bioRxiv

    Article Title: Paxillin Promotes ATP-induced Activation of P2X7 Receptor and NLRP3 Inflammasome

    doi: 10.1101/2020.04.03.023721

    Figure Lengend Snippet: ATP promotes Paxillin phosphorylation and Paxillin-NLRP3 interaction. A–D THP-1 cells stably expressing control lentivirus or Paxillin lentivirus were generated and differentiated to macrophages, which were then treated with ATP (5 mM) or Nigericin (2 μM) for 2 h (A, B), treated with DMSO, YVAD (50 μM), Glybenclamide (25 μg/ml), A438079 (100 μM), AZ10606120 (100 μM), and BAPTA-AM (30 μM) for 1 h before the treatment with ATP (5 mM) for 2 h (C, D). IL-1β levels in supernatants were determined by ELISA (A, C). Mature IL-1β (p17) and cleaved Casp-1 (p22/p20) in supernatants or Paxillin, pro-IL-1β, and pro-Casp-1 in lysates were determined by Western blot (B, D). E TPA-differentiated THP-1 macrophages were treated with DMSO or ATP (5 mM) for 2 h. Lysates were prepared and subjected to IP (top) or subjected to Western blot (as input) (bottom). F BMDM cells prepared from C57BL/6 mice bone marrow were treated with LPS (1 μg/ml) for 6 h, and the primed BMDMs were stimulated by DMSO or ATP (5 mM) for 30 min. Lysates were prepared and subjected to IP (top) or subjected to Western blot (as input) (bottom). G, H TPA-differentiated THP-1 macrophages were treated with ATP (5 mM) for 30, 60, and 120 min (G). LPS-primed BMDMs were stimulated by ATP (5 mM) for 5, 15, and 30 min (H). The protein level of p-Paxillin(Y 118), p-Paxillin(Y31), Paxillin, and GAPDH was determined by Western blot. I HEK293T cells were co-transfected with pFlag-NLRP3 and pHA-Vector, pHA-Paxillin, pHA-Paxillin-(Y31A), and pHA-Paxillin-(Y118A). Lysates were prepared and subjected to IP (top) or subjected to Western blot (as input) (bottom). J TPA-differentiated THP-1 macrophages were treated with DMSO, A438079 (100 μM), AZ10606120 (100 μM) for 1 h before the treatment with ATP (5 mM) for 2 h. The protein level of p-Paxillin(Y118), Paxillin, and GAPDH was determined by Western blot. Data shown are means ± SEMs; ***p

    Article Snippet: Enzyme-linked immunosorbent assay (ELISA) The concentrations of human IL-1β in culture supernatants were measured by ELISA kit (BD Biosciences, San Jose, CA, USA).

    Techniques: Stable Transfection, Expressing, Generated, Enzyme-linked Immunosorbent Assay, Western Blot, Mouse Assay, Transfection, Plasmid Preparation

    Paxillin is required for ATP- and Nigericin-induced NLRP3 inflammasome activation. A, B BMDMs prepared from C57BL/6 mice bone marrow were infected by lentivirus that expressing sh-Paxillin for 3 days. LPS-primed BMDMs were stimulated by MDP (50 μg/ml) for 6 h, ATP (5 mM) for 30 min, poly(dA:dT) (5 μg/ml) for 6 h, or Salmonella for 4 h. C, D LPS-primed BMDMs were stimulated by ATP (5 mM) for 30 min, Nigericin (2 μM) for 2 h, MSU (100 μg/ml) for 6 h, or Alum crystals (200 mg/ml) for 6 h. E, F BMDCs prepared from C57BL/6 mice bone marrow were infected by lentivirus that express sh-Paxillin for 3 days. LPS-primed BMDCs were stimulated by ATP (5 mM) for 30 min, Nigericin (2 μM) for 2 h, or MSU (100 μg/ml) for 6 h. IL-1β levels in supernatants were determined by ELISA (A, C, and E). Mature IL-1β (p17) and cleaved Casp-1 (p10) in supernatants as well as Paxillin, pro-IL-1β, and pro-Casp-1 in lysates were determined by Western blot (B, D, and F). G, H PBMCs isolated from healthy individuals were infected by lentivirus that expresses sh-Paxillin for 2 days. Before stimulation, PBMCs were treated with LPS (1 μg/ml) for 6 h, and PBMCs were then stimulated by ATP (5 mM) for 30 min, Nigericin (2 μM) for 2 h, MSU (100 μg/ml) for 6 h, Alum crystals (200 mg/ml) for 6 h, or MDP (50 μg/ml) for 6 h. Paxillin in lysates was determined by Western blot (G). IL-1β levels in supernatants were determined by ELISA (H). I, J THP-1 cells stably expressing sh-NC or sh-Paxillin were generated and differentiated to macrophages, which were then treated with ATP (5 mM) for 2 h, Nigericin (2 μM) for 2 h, or MSU (100 μg/ml) for 6 h. IL-1β levels in supernatants were determined by ELISA (I). Mature IL-1β (p17) and cleaved Casp-1 (p22/p20) in supernatants and Paxillin, pro-IL-1β and pro-Casp-1 in lysates were determined by Western blot (J). Data shown are means ± SEMs; ***p

    Journal: bioRxiv

    Article Title: Paxillin Promotes ATP-induced Activation of P2X7 Receptor and NLRP3 Inflammasome

    doi: 10.1101/2020.04.03.023721

    Figure Lengend Snippet: Paxillin is required for ATP- and Nigericin-induced NLRP3 inflammasome activation. A, B BMDMs prepared from C57BL/6 mice bone marrow were infected by lentivirus that expressing sh-Paxillin for 3 days. LPS-primed BMDMs were stimulated by MDP (50 μg/ml) for 6 h, ATP (5 mM) for 30 min, poly(dA:dT) (5 μg/ml) for 6 h, or Salmonella for 4 h. C, D LPS-primed BMDMs were stimulated by ATP (5 mM) for 30 min, Nigericin (2 μM) for 2 h, MSU (100 μg/ml) for 6 h, or Alum crystals (200 mg/ml) for 6 h. E, F BMDCs prepared from C57BL/6 mice bone marrow were infected by lentivirus that express sh-Paxillin for 3 days. LPS-primed BMDCs were stimulated by ATP (5 mM) for 30 min, Nigericin (2 μM) for 2 h, or MSU (100 μg/ml) for 6 h. IL-1β levels in supernatants were determined by ELISA (A, C, and E). Mature IL-1β (p17) and cleaved Casp-1 (p10) in supernatants as well as Paxillin, pro-IL-1β, and pro-Casp-1 in lysates were determined by Western blot (B, D, and F). G, H PBMCs isolated from healthy individuals were infected by lentivirus that expresses sh-Paxillin for 2 days. Before stimulation, PBMCs were treated with LPS (1 μg/ml) for 6 h, and PBMCs were then stimulated by ATP (5 mM) for 30 min, Nigericin (2 μM) for 2 h, MSU (100 μg/ml) for 6 h, Alum crystals (200 mg/ml) for 6 h, or MDP (50 μg/ml) for 6 h. Paxillin in lysates was determined by Western blot (G). IL-1β levels in supernatants were determined by ELISA (H). I, J THP-1 cells stably expressing sh-NC or sh-Paxillin were generated and differentiated to macrophages, which were then treated with ATP (5 mM) for 2 h, Nigericin (2 μM) for 2 h, or MSU (100 μg/ml) for 6 h. IL-1β levels in supernatants were determined by ELISA (I). Mature IL-1β (p17) and cleaved Casp-1 (p22/p20) in supernatants and Paxillin, pro-IL-1β and pro-Casp-1 in lysates were determined by Western blot (J). Data shown are means ± SEMs; ***p

    Article Snippet: Enzyme-linked immunosorbent assay (ELISA) The concentrations of human IL-1β in culture supernatants were measured by ELISA kit (BD Biosciences, San Jose, CA, USA).

    Techniques: Activation Assay, Mouse Assay, Infection, Expressing, Enzyme-linked Immunosorbent Assay, Western Blot, Isolation, Stable Transfection, Generated

    qRT-PCR and ELISA testing of expression in sheep monocytes with LPS stimulation. A, B and C show the expression of TLR4 , Myd88 and NF-κB , at 0min, 5 min, 30 min and 60 min after LPS (100 ng/mL) stimulation, respectively. D, E and F show the phosphorylation levels of p38 , JNK , and ERK signaling at 0min, 5 min, 30 min and 60 min after LPS (100 ng/mL) stimulation, respectively. Tg: transgenic sheep; NTg: non-transgenic sheep. All data are presented as the mean ± SEM from three experiments, *P

    Journal: International Journal of Biological Sciences

    Article Title: Overexpression of Toll-like Receptor 4-linked Mitogen-activated Protein Kinase Signaling Contributes to Internalization of Escherichia coli in Sheep

    doi: 10.7150/ijbs.25275

    Figure Lengend Snippet: qRT-PCR and ELISA testing of expression in sheep monocytes with LPS stimulation. A, B and C show the expression of TLR4 , Myd88 and NF-κB , at 0min, 5 min, 30 min and 60 min after LPS (100 ng/mL) stimulation, respectively. D, E and F show the phosphorylation levels of p38 , JNK , and ERK signaling at 0min, 5 min, 30 min and 60 min after LPS (100 ng/mL) stimulation, respectively. Tg: transgenic sheep; NTg: non-transgenic sheep. All data are presented as the mean ± SEM from three experiments, *P

    Article Snippet: Phosphorylation levels of p38, JNK and ERK were analyzed using ELISA kit (Abcam) according to the manufacturer's instructions.

    Techniques: Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Expressing, Transgenic Assay

    Antitumor potency of the SA-IL-2 MCSCs vaccine in the pulmonary model mice. a In the cytotoxicity assay, the portion of CTL in the experimental group was significantly higher than in the four control groups. b In ELISA assay, the expression of serum IgG antibodies in the experimental group was significantly higher than in the four control groups. c FCM analysis showed that the portion of DCs (CD11c + CD80 + ) in the experimental group was significantly larger than in the four control groups. d FCM analysis showed that the portion of CD8 + and CD4 + T cells in the experimental group was significantly larger than in the four control groups. *P

    Journal: Stem Cell Research & Therapy

    Article Title: Development of a therapy against metastatic bladder cancer using an interleukin-2 surface-modified MB49 bladder cancer stem cells vaccine

    doi: 10.1186/s13287-015-0211-1

    Figure Lengend Snippet: Antitumor potency of the SA-IL-2 MCSCs vaccine in the pulmonary model mice. a In the cytotoxicity assay, the portion of CTL in the experimental group was significantly higher than in the four control groups. b In ELISA assay, the expression of serum IgG antibodies in the experimental group was significantly higher than in the four control groups. c FCM analysis showed that the portion of DCs (CD11c + CD80 + ) in the experimental group was significantly larger than in the four control groups. d FCM analysis showed that the portion of CD8 + and CD4 + T cells in the experimental group was significantly larger than in the four control groups. *P

    Article Snippet: The concentrations of IgG were measured using ELISA kits (Abcam) according to the manufacturer’s protocol.

    Techniques: Mouse Assay, Cytotoxicity Assay, CTL Assay, Enzyme-linked Immunosorbent Assay, Expressing

    Antitumor potency of the SA-IL-2 MCSCs vaccine in the subcutaneous model mice. a In the cytotoxicity assay, the portion of CTL in the experimental group was significantly higher than in the four control groups. b In ELISA assay, the expression of serum IgG antibodies in the experimental group was significantly higher than in the four control groups. c FCM analysis showed that the portion of DCs (CD11c + CD80 + ) in the experimental group was significantly larger than in the four control groups. d FCM analysis showed that the portion of CD8 + and CD4 + T cells in the experimental group was significantly larger than in the four control groups. *P

    Journal: Stem Cell Research & Therapy

    Article Title: Development of a therapy against metastatic bladder cancer using an interleukin-2 surface-modified MB49 bladder cancer stem cells vaccine

    doi: 10.1186/s13287-015-0211-1

    Figure Lengend Snippet: Antitumor potency of the SA-IL-2 MCSCs vaccine in the subcutaneous model mice. a In the cytotoxicity assay, the portion of CTL in the experimental group was significantly higher than in the four control groups. b In ELISA assay, the expression of serum IgG antibodies in the experimental group was significantly higher than in the four control groups. c FCM analysis showed that the portion of DCs (CD11c + CD80 + ) in the experimental group was significantly larger than in the four control groups. d FCM analysis showed that the portion of CD8 + and CD4 + T cells in the experimental group was significantly larger than in the four control groups. *P

    Article Snippet: The concentrations of IgG were measured using ELISA kits (Abcam) according to the manufacturer’s protocol.

    Techniques: Mouse Assay, Cytotoxicity Assay, CTL Assay, Enzyme-linked Immunosorbent Assay, Expressing

    Acetylation and expression of StAR in response to HDAC inhibitors, and their correlation to E2 synthesis in MCF7 cells. Cells (1 × 10 6 per dish) were plated in 100-mm dishes 24h before treatments. Cells were then treated without or with panobinostat (10nM) for 0-180min (A-C) or other HDAC inhibitors for 45min (D-F). Following treatments, cells were collected, extracted with lysis buffer, and processed for either immunoprecipitation or immunoblotting studies, as described in Materials and methods . Representative immunoblots illustrate acetylation (Ac-StAR) and expression of StAR (T-StAR) in different treatment groups. IgG heavy chain (IgG-Hc) and β-actin expression were assessed for loading controls in immunoprecipitation and immunoblotting, respectively. Integrated optical density (IOD) values of Ac-StAR and T-StAR in each band were quantified and normalized with corresponding IgG-Hc and β-actin expression, and presented as fold response (C,F). E2 levels in media at each time point were determined by ELISA and presented as pg/mg protein (C). Data are representative of 3-4 independent experiments. *, p

    Journal: Biochemical and biophysical research communications

    Article Title: Overexpression of the steroidogenic acute regulatory protein in breast cancer: Regulation by histone deacetylase inhibition

    doi: 10.1016/j.bbrc.2018.12.145

    Figure Lengend Snippet: Acetylation and expression of StAR in response to HDAC inhibitors, and their correlation to E2 synthesis in MCF7 cells. Cells (1 × 10 6 per dish) were plated in 100-mm dishes 24h before treatments. Cells were then treated without or with panobinostat (10nM) for 0-180min (A-C) or other HDAC inhibitors for 45min (D-F). Following treatments, cells were collected, extracted with lysis buffer, and processed for either immunoprecipitation or immunoblotting studies, as described in Materials and methods . Representative immunoblots illustrate acetylation (Ac-StAR) and expression of StAR (T-StAR) in different treatment groups. IgG heavy chain (IgG-Hc) and β-actin expression were assessed for loading controls in immunoprecipitation and immunoblotting, respectively. Integrated optical density (IOD) values of Ac-StAR and T-StAR in each band were quantified and normalized with corresponding IgG-Hc and β-actin expression, and presented as fold response (C,F). E2 levels in media at each time point were determined by ELISA and presented as pg/mg protein (C). Data are representative of 3-4 independent experiments. *, p

    Article Snippet: 17β-estradiol (E2) levels in cell culture media were determined using ELISA Kit from Cayman Chemical (Ann Arbor, MI), according to manufacturer’s protocol [ ], optimized further for better efficacy.

    Techniques: Expressing, Lysis, Immunoprecipitation, Western Blot, Enzyme-linked Immunosorbent Assay

    Relative expression of StAR and aromatase proteins, and E2 synthesis, in human normal mammary epithelial (MCF10A and MCF12F), ER+ breast cancer (MCF7, MBA-MD-361, and T-47D), TNBC (MBA-MD-468, MBA-MD-231, and BT-549), and H295R, cell lines. These cells were cultured with appropriate media, and were harvested in RIPA buffer at 75-80% confluence. Cells and media were then processed for whole cell extract preparation and E2 extraction by diethyl ether, respectively. Representative immunoblots illustrate total StAR (T-StAR) and aromatase protein levels in different cell lines (A). Immunoblots shown are representative of 4 independent experiments. β-actin expression was assessed as a loading control. B, E2 levels in media from different cell lines were determined by ELISA and presented as pg/mg protein (n=4, ±SE). Different letters above the bars indicate that these groups differ significantly from each other at least at p

    Journal: Biochemical and biophysical research communications

    Article Title: Overexpression of the steroidogenic acute regulatory protein in breast cancer: Regulation by histone deacetylase inhibition

    doi: 10.1016/j.bbrc.2018.12.145

    Figure Lengend Snippet: Relative expression of StAR and aromatase proteins, and E2 synthesis, in human normal mammary epithelial (MCF10A and MCF12F), ER+ breast cancer (MCF7, MBA-MD-361, and T-47D), TNBC (MBA-MD-468, MBA-MD-231, and BT-549), and H295R, cell lines. These cells were cultured with appropriate media, and were harvested in RIPA buffer at 75-80% confluence. Cells and media were then processed for whole cell extract preparation and E2 extraction by diethyl ether, respectively. Representative immunoblots illustrate total StAR (T-StAR) and aromatase protein levels in different cell lines (A). Immunoblots shown are representative of 4 independent experiments. β-actin expression was assessed as a loading control. B, E2 levels in media from different cell lines were determined by ELISA and presented as pg/mg protein (n=4, ±SE). Different letters above the bars indicate that these groups differ significantly from each other at least at p

    Article Snippet: 17β-estradiol (E2) levels in cell culture media were determined using ELISA Kit from Cayman Chemical (Ann Arbor, MI), according to manufacturer’s protocol [ ], optimized further for better efficacy.

    Techniques: Expressing, Cell Culture, Western Blot, Enzyme-linked Immunosorbent Assay

    The effects of ambient temperature on IFN-γ, IL-4, IL-13 and serum OVA specific IgE. The BALF and sera were collected from the mice and analyzed by ELISA. The bars indicate the cytokine levels in the BALF ( A ) and the antigen-specific IgE in the serum ( B ). ST: standard temperature (20 °C); TT: thermoneutral temperature (30 °C). Control: Naïve mice. Asthma: Asthma mice. ( C ) Spleen cells were cultured at the indicated temperature for 48 h in the presence of ionomycin. The data are presented as mean ± SEM. *P

    Journal: Scientific Reports

    Article Title: Thermoneutral housing temperature regulates T-regulatory cell function and inhibits ovabumin-induced asthma development in mice

    doi: 10.1038/s41598-017-07471-7

    Figure Lengend Snippet: The effects of ambient temperature on IFN-γ, IL-4, IL-13 and serum OVA specific IgE. The BALF and sera were collected from the mice and analyzed by ELISA. The bars indicate the cytokine levels in the BALF ( A ) and the antigen-specific IgE in the serum ( B ). ST: standard temperature (20 °C); TT: thermoneutral temperature (30 °C). Control: Naïve mice. Asthma: Asthma mice. ( C ) Spleen cells were cultured at the indicated temperature for 48 h in the presence of ionomycin. The data are presented as mean ± SEM. *P

    Article Snippet: The levels of IL-4, IL-13 and IFN-γ in spleen homogenate, BALF and the culture supernatants, OVA specific IgE in the serum were assessed using commercial ELISA kits from Cayman Chemical Co. (Ann Arbor, MI, USA) according to the manufacturer’s instructions.

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay, Cell Culture

    Impaired insulin secretion from isolated Pick1 cKO islets. (a) GSIS was performed on isolated islets stimulated with 4.8 or 30 mM KCl or 2.8 or 16.7 mM glucose (Glc). (b) The stimulated index was also calculated following KCl or Glc treatment. (c) Insulin content in the isolated islets was measured by ELISA. (d) Isolated WT and Pick1 cKO islets were incubated in the presence of 16.7 mM Glc at 2-min intervals for 20 min. (e) Same data as shown in d, but plotted as release per 2 min. (f, g) Secreted insulin was tested from isolated WT and Pick1 cKO islets incubated with or without E4. Data are represented as mean ± SEM, n = 3, three to four groups per type of mouse, 10–0 islets per group. * p

    Journal: Molecular Biology of the Cell

    Article Title: PICK1 is essential for insulin production and the maintenance of glucose homeostasis

    doi: 10.1091/mbc.E17-03-0204

    Figure Lengend Snippet: Impaired insulin secretion from isolated Pick1 cKO islets. (a) GSIS was performed on isolated islets stimulated with 4.8 or 30 mM KCl or 2.8 or 16.7 mM glucose (Glc). (b) The stimulated index was also calculated following KCl or Glc treatment. (c) Insulin content in the isolated islets was measured by ELISA. (d) Isolated WT and Pick1 cKO islets were incubated in the presence of 16.7 mM Glc at 2-min intervals for 20 min. (e) Same data as shown in d, but plotted as release per 2 min. (f, g) Secreted insulin was tested from isolated WT and Pick1 cKO islets incubated with or without E4. Data are represented as mean ± SEM, n = 3, three to four groups per type of mouse, 10–0 islets per group. * p

    Article Snippet: The serum insulin and proinsulin and the insulin from the medium and islet supernatants collected from the GSIS were tested using the Ultra-sensitive mouse insulin ELISA Kit (Crystal Chem) or the Proinsulin rat/mouse ELISA Kit (Mercodia Insulin/Proinsulin).

    Techniques: Isolation, Gas Chromatography, Enzyme-linked Immunosorbent Assay, Incubation

    Effect of EA on Glu content and expression of NMDAR2A and NMDAR2B protein. (A) Glu content in the CA1 area of the hippocampus in each group, measured by ELISA. (B) Protein expression of NMDAR2A and NMDAR2B in the CA1 area of the hippocampus in each group, examined by Western blotting. (C,D) Quantitative indices of protein expression of NR2A and NR2B, derived from figure 2B . #P

    Journal: Acupuncture in Medicine

    Article Title: Electroacupuncture ameliorates cognitive impairment through inhibition of Ca2+-mediated neurotoxicity in a rat model of cerebral ischaemia–reperfusion injury

    doi: 10.1136/acupmed-2016-011353

    Figure Lengend Snippet: Effect of EA on Glu content and expression of NMDAR2A and NMDAR2B protein. (A) Glu content in the CA1 area of the hippocampus in each group, measured by ELISA. (B) Protein expression of NMDAR2A and NMDAR2B in the CA1 area of the hippocampus in each group, examined by Western blotting. (C,D) Quantitative indices of protein expression of NR2A and NR2B, derived from figure 2B . #P

    Article Snippet: The hippocampus was separated quickly and stored in liquid nitrogen pending measurement of the concentration of Glu using ELISA kits (Sigma, San Francisco, California, USA).

    Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Western Blot, Derivative Assay

    Regulation of TNF- α production in RAW 264.7 cells. Cells were treated with the steam (a) or 50% ethanol (b) GE at various concentrations for 24 h. The culture media were collected and cytokine concentrations were measured by ELISA. The data were expressed as mean ± SEM. ∗ p

    Journal: Evidence-based Complementary and Alternative Medicine : eCAM

    Article Title: Immunomodulatory Efficacy of Standardized Annona muricata (Graviola) Leaf Extract via Activation of Mitogen-Activated Protein Kinase Pathways in RAW 264.7 Macrophages

    doi: 10.1155/2016/2905127

    Figure Lengend Snippet: Regulation of TNF- α production in RAW 264.7 cells. Cells were treated with the steam (a) or 50% ethanol (b) GE at various concentrations for 24 h. The culture media were collected and cytokine concentrations were measured by ELISA. The data were expressed as mean ± SEM. ∗ p

    Article Snippet: The concentration of TNF-α and IL-1β was measured using ELISA kit (Enzo Life Sciences, Ann Arbor, MI, USA).

    Techniques: Serial Time-encoded Amplified Microscopy, Enzyme-linked Immunosorbent Assay

    Moxibustion modulated the hypothalamic-pituitary-adrenal axis of CFS rats. Data are presented as the mean ± SEM ( n = 8 per group; one-way analysis of variance followed by the least significant difference post hoc test). (A) CRH mRNA expression in the hypothalamus was semi-quantified by real-time PCR. (B, C) ACTH (B) and CORT (C) concentration in plasma was detected by enzyme-linked immunosorbent assay. ** P

    Journal: Neural Regeneration Research

    Article Title: Moxibustion upregulates hippocampal progranulin expression

    doi: 10.4103/1673-5374.180746

    Figure Lengend Snippet: Moxibustion modulated the hypothalamic-pituitary-adrenal axis of CFS rats. Data are presented as the mean ± SEM ( n = 8 per group; one-way analysis of variance followed by the least significant difference post hoc test). (A) CRH mRNA expression in the hypothalamus was semi-quantified by real-time PCR. (B, C) ACTH (B) and CORT (C) concentration in plasma was detected by enzyme-linked immunosorbent assay. ** P

    Article Snippet: Enzyme-linked immunosorbent assay (ELISA) Plasma levels of adrenocorticotropic hormone (ACTH) and corticosterone (CORT) were measured 2 days after blood was taken using a commercial ELISA kit (USCN-LIFE™, Wuhan, China), following the manufacturer's instructions.

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Concentration Assay, Enzyme-linked Immunosorbent Assay

    Impaired insulin secretion and proinsulin maturation in XBP1-deficient β-cells. ( A ) Glucose-stimulated insulin secretion assays. Serum insulin levels were measured in control heterozygous Xbp1 f/+ ;RIP-cre and Xbp1 f/f ;RIP-cre mice at indicated time points after a bolus glucose injection. Each line represents an individual mouse. ( B ) Min6 cells stably expressing shRNAs targeting control luciferase or XBP1 mRNAs were tested for XBP1s expression by Western blot. ( C ) Cells were pretreated with 2.5 mM glucose for 2 h and then cultured in 5 mM or 25 mM glucose media for 2 h. Culture media were collected to measure insulin content by ELISA. ( D ) Serum proinsulin levels relative to total insulin in WT and Xbp1 f/f ;RIP-cre mice were determined. Data from both males and females were combined, because sex difference was not significant. The graphs display proinsulin:insulin ratio ( Left ) or proinsulin and insulin concentrations (pmol per liter) ( Right ). Each dot represents an individual mouse (4- to 6-mo-old). ( E ) Min6 cells expressing control or XBP1 shRNA were pulse-labeled for 30 min with [ 35 S]Met/Cys and then cultured in chase media for the indicated time. Cells and culture supernatants were harvested for immunoprecipitation of radiolabeled insulin and proinsulin, which were revealed by SDS/PAGE followed by autoradiography.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Dual and opposing roles of the unfolded protein response regulated by IRE1? and XBP1 in proinsulin processing and insulin secretion

    doi: 10.1073/pnas.1105564108

    Figure Lengend Snippet: Impaired insulin secretion and proinsulin maturation in XBP1-deficient β-cells. ( A ) Glucose-stimulated insulin secretion assays. Serum insulin levels were measured in control heterozygous Xbp1 f/+ ;RIP-cre and Xbp1 f/f ;RIP-cre mice at indicated time points after a bolus glucose injection. Each line represents an individual mouse. ( B ) Min6 cells stably expressing shRNAs targeting control luciferase or XBP1 mRNAs were tested for XBP1s expression by Western blot. ( C ) Cells were pretreated with 2.5 mM glucose for 2 h and then cultured in 5 mM or 25 mM glucose media for 2 h. Culture media were collected to measure insulin content by ELISA. ( D ) Serum proinsulin levels relative to total insulin in WT and Xbp1 f/f ;RIP-cre mice were determined. Data from both males and females were combined, because sex difference was not significant. The graphs display proinsulin:insulin ratio ( Left ) or proinsulin and insulin concentrations (pmol per liter) ( Right ). Each dot represents an individual mouse (4- to 6-mo-old). ( E ) Min6 cells expressing control or XBP1 shRNA were pulse-labeled for 30 min with [ 35 S]Met/Cys and then cultured in chase media for the indicated time. Cells and culture supernatants were harvested for immunoprecipitation of radiolabeled insulin and proinsulin, which were revealed by SDS/PAGE followed by autoradiography.

    Article Snippet: Serum proinsulin levels were measured using an ELISA kit (ALPCO Diagnostics).

    Techniques: Mouse Assay, Injection, Stable Transfection, Expressing, Luciferase, Western Blot, Cell Culture, Enzyme-linked Immunosorbent Assay, shRNA, Labeling, Immunoprecipitation, SDS Page, Autoradiography

    Pregnancy increases lung tissue expression of proinflammatory cytokines and chemokines while dampening the upregulation of progesterone and PGF2α following infection. Hormone expression in lung lysates was quantified via ELISA in uninfected (N = 6) and infected (N = 5) non-pregnant and pregnant mice for A. progesterone; B. prostaglandin F2α (PGF2α); and C. prostaglandin E2 (PGE2). Cytokine and chemokine expression in sera was quantified in lung lysates of D. uninfected pregnant (E16) (N = 8) and non-pregnant mice (N = 6); E. infected pregnant (4 d.p.i., E16) (N = 5) and non-pregnant mice (N = 5). Student’s t-test was performed between selected groups and significance noted above asterisk brackets. Abbreviations for groups are as described in Fig 3 .

    Journal: PLoS Pathogens

    Article Title: H1N1 influenza virus infection results in adverse pregnancy outcomes by disrupting tissue-specific hormonal regulation

    doi: 10.1371/journal.ppat.1006757

    Figure Lengend Snippet: Pregnancy increases lung tissue expression of proinflammatory cytokines and chemokines while dampening the upregulation of progesterone and PGF2α following infection. Hormone expression in lung lysates was quantified via ELISA in uninfected (N = 6) and infected (N = 5) non-pregnant and pregnant mice for A. progesterone; B. prostaglandin F2α (PGF2α); and C. prostaglandin E2 (PGE2). Cytokine and chemokine expression in sera was quantified in lung lysates of D. uninfected pregnant (E16) (N = 8) and non-pregnant mice (N = 6); E. infected pregnant (4 d.p.i., E16) (N = 5) and non-pregnant mice (N = 5). Student’s t-test was performed between selected groups and significance noted above asterisk brackets. Abbreviations for groups are as described in Fig 3 .

    Article Snippet: Determination of hormone levels Progesterone, prostaglandin F2α (PGF2α) and prostaglandin E2 (PGE2) were quantified via ELISA kits from ALPCO (Salem, NH).

    Techniques: Expressing, Infection, Enzyme-linked Immunosorbent Assay, Mouse Assay

    Infection increases placental expression of contraction-inducing PGF2α and reduces pregnancy-supportive progesterone. Placental lysates were isolated from uninfected and infected pregnant mice (E16). Hormone expression was quantified via ELISA for A. progesterone (N = 6 per group); B. prostaglandin F2α (PGF2α) (N = 9, uninfected, N = 7, infected) and C. prostaglandin E2 (PGE2) (N = 9 per group). Student’s t-test was performed between selected groups and significance noted above asterisk brackets.

    Journal: PLoS Pathogens

    Article Title: H1N1 influenza virus infection results in adverse pregnancy outcomes by disrupting tissue-specific hormonal regulation

    doi: 10.1371/journal.ppat.1006757

    Figure Lengend Snippet: Infection increases placental expression of contraction-inducing PGF2α and reduces pregnancy-supportive progesterone. Placental lysates were isolated from uninfected and infected pregnant mice (E16). Hormone expression was quantified via ELISA for A. progesterone (N = 6 per group); B. prostaglandin F2α (PGF2α) (N = 9, uninfected, N = 7, infected) and C. prostaglandin E2 (PGE2) (N = 9 per group). Student’s t-test was performed between selected groups and significance noted above asterisk brackets.

    Article Snippet: Determination of hormone levels Progesterone, prostaglandin F2α (PGF2α) and prostaglandin E2 (PGE2) were quantified via ELISA kits from ALPCO (Salem, NH).

    Techniques: Infection, Expressing, Isolation, Mouse Assay, Enzyme-linked Immunosorbent Assay

    Viral infection disproportionately reduces cytokine and hormone expression in the sera and lungs during pregnancy. Hormone expression in sera was quantified via ELISA in uninfected and infected non-pregnant mice for A. progesterone; C. prostaglandin F2α (PGF2α); and D. prostaglandin E2 (PGE2). Cytokine and chemokine expression in sera was quantified via Bio-Rad 23-plex assay in B. uninfected pregnant (E16) (N = 5) and non-pregnant mice (N = 5); E. uninfected and infected pregnant (E16; 4 d.p.i) (N = 5 per group); and F. infected (4 d.p.i) non-pregnant and pregnant (E16) mice (N = 5 per group). Student’s t-test was performed between selected groups and significance noted above asterisk brackets. Inf.: infected; Uninf: uninfected; NP: non-pregnant; P: pregnant mice.

    Journal: PLoS Pathogens

    Article Title: H1N1 influenza virus infection results in adverse pregnancy outcomes by disrupting tissue-specific hormonal regulation

    doi: 10.1371/journal.ppat.1006757

    Figure Lengend Snippet: Viral infection disproportionately reduces cytokine and hormone expression in the sera and lungs during pregnancy. Hormone expression in sera was quantified via ELISA in uninfected and infected non-pregnant mice for A. progesterone; C. prostaglandin F2α (PGF2α); and D. prostaglandin E2 (PGE2). Cytokine and chemokine expression in sera was quantified via Bio-Rad 23-plex assay in B. uninfected pregnant (E16) (N = 5) and non-pregnant mice (N = 5); E. uninfected and infected pregnant (E16; 4 d.p.i) (N = 5 per group); and F. infected (4 d.p.i) non-pregnant and pregnant (E16) mice (N = 5 per group). Student’s t-test was performed between selected groups and significance noted above asterisk brackets. Inf.: infected; Uninf: uninfected; NP: non-pregnant; P: pregnant mice.

    Article Snippet: Determination of hormone levels Progesterone, prostaglandin F2α (PGF2α) and prostaglandin E2 (PGE2) were quantified via ELISA kits from ALPCO (Salem, NH).

    Techniques: Infection, Expressing, Enzyme-linked Immunosorbent Assay, Mouse Assay, Plex Assay

    Effects of the β2-GPI DNA vaccine and FK506 treatment on β2-GPI-specific Treg cell response. Spleen cells were purified from the different mouse groups on day 56 and stimulated with recombinant β2-GPI protein (10 μg/mL). After 96 h, (A) the percentage of Foxp3-expressing CD4+ T cells was determined by flow cytometry. The dot plot shows data from one representative mouse from each group. The bar graph represents the mean ± standard deviation (SD) of six mice from three independent experiments. (B) The culture supernatants were collected, and IL-10 and TGF-β production was analyzed in triplicate by ELISA. The data are presented as the means ± SD of triplicate assays from six mice/group. ns p > 0.05, **p

    Journal: PLoS ONE

    Article Title: Tolerogenic β2-glycoprotein I DNA vaccine and FK506 as an adjuvant attenuates experimental obstetric antiphospholipid syndrome

    doi: 10.1371/journal.pone.0198821

    Figure Lengend Snippet: Effects of the β2-GPI DNA vaccine and FK506 treatment on β2-GPI-specific Treg cell response. Spleen cells were purified from the different mouse groups on day 56 and stimulated with recombinant β2-GPI protein (10 μg/mL). After 96 h, (A) the percentage of Foxp3-expressing CD4+ T cells was determined by flow cytometry. The dot plot shows data from one representative mouse from each group. The bar graph represents the mean ± standard deviation (SD) of six mice from three independent experiments. (B) The culture supernatants were collected, and IL-10 and TGF-β production was analyzed in triplicate by ELISA. The data are presented as the means ± SD of triplicate assays from six mice/group. ns p > 0.05, **p

    Article Snippet: Anti-β2-GPI IgG concentrations were measured with an ELISA kit (MyBioSource, San Diego, CA) according to the manufacturer's protocol.

    Techniques: Purification, Recombinant, Expressing, Flow Cytometry, Cytometry, Standard Deviation, Mouse Assay, Enzyme-linked Immunosorbent Assay

    Effects of the β2-GPI DNA vaccine and FK506 treatment on IgG anti-β2-GPI antibody levels. Serum samples were obtained from each mouse group on day 56, and IgG anti-β2-GPI antibody levels were analyzed by ELISA. The data are presented as the means ± standard deviation of triplicate assays from six mice/group. ns p > 0.05, *p

    Journal: PLoS ONE

    Article Title: Tolerogenic β2-glycoprotein I DNA vaccine and FK506 as an adjuvant attenuates experimental obstetric antiphospholipid syndrome

    doi: 10.1371/journal.pone.0198821

    Figure Lengend Snippet: Effects of the β2-GPI DNA vaccine and FK506 treatment on IgG anti-β2-GPI antibody levels. Serum samples were obtained from each mouse group on day 56, and IgG anti-β2-GPI antibody levels were analyzed by ELISA. The data are presented as the means ± standard deviation of triplicate assays from six mice/group. ns p > 0.05, *p

    Article Snippet: Anti-β2-GPI IgG concentrations were measured with an ELISA kit (MyBioSource, San Diego, CA) according to the manufacturer's protocol.

    Techniques: Enzyme-linked Immunosorbent Assay, Standard Deviation, Mouse Assay

    Effects of the β2-GPI DNA vaccine and FK506 treatment on cytokine production. Spleen cells were purified from the different mouse groups on day 56 and stimulated with recombinant β2-GPI protein (10 μg/mL). After 96 h, (A) the culture supernatants were collected, and interferon (IFN)- γ, interleukin (IL)-4 and IL-17A production was analyzed in triplicate by ELISA. The data are presented as the means ± standard deviation (SD) of triplicate assays from six mice/group. (B) The percentages of IFN-γ-expressing CD4+ T cells, IL-4-expressing CD4+ T cells and IL-17A-expressing CD4+ T cells were determined by flow cytometry. The dot plot shows data from one representative mouse from each group. The bar graph represents the mean ± SD of six mice from three independent experiments. ns p > 0.05, *p

    Journal: PLoS ONE

    Article Title: Tolerogenic β2-glycoprotein I DNA vaccine and FK506 as an adjuvant attenuates experimental obstetric antiphospholipid syndrome

    doi: 10.1371/journal.pone.0198821

    Figure Lengend Snippet: Effects of the β2-GPI DNA vaccine and FK506 treatment on cytokine production. Spleen cells were purified from the different mouse groups on day 56 and stimulated with recombinant β2-GPI protein (10 μg/mL). After 96 h, (A) the culture supernatants were collected, and interferon (IFN)- γ, interleukin (IL)-4 and IL-17A production was analyzed in triplicate by ELISA. The data are presented as the means ± standard deviation (SD) of triplicate assays from six mice/group. (B) The percentages of IFN-γ-expressing CD4+ T cells, IL-4-expressing CD4+ T cells and IL-17A-expressing CD4+ T cells were determined by flow cytometry. The dot plot shows data from one representative mouse from each group. The bar graph represents the mean ± SD of six mice from three independent experiments. ns p > 0.05, *p

    Article Snippet: Anti-β2-GPI IgG concentrations were measured with an ELISA kit (MyBioSource, San Diego, CA) according to the manufacturer's protocol.

    Techniques: Purification, Recombinant, Enzyme-linked Immunosorbent Assay, Standard Deviation, Mouse Assay, Expressing, Flow Cytometry, Cytometry

    IL-25 is highly expressed in HCC patients. a The level of NMU were detected by ELISA in serum of hepatic hemangioma patients ( n = 10) and HCC patients ( n = 10). b Immunohistochemistry (IHC) staining was performed in a tissue microarray consisted of 228 HCC peri- and intra-tumor tissue, NMU representative IHC images are shown. Bar, 20 μm. ** p

    Journal: BMC Cancer

    Article Title: The prognostic value of neuromedin U in patients with hepatocellular carcinoma

    doi: 10.1186/s12885-020-6532-1

    Figure Lengend Snippet: IL-25 is highly expressed in HCC patients. a The level of NMU were detected by ELISA in serum of hepatic hemangioma patients ( n = 10) and HCC patients ( n = 10). b Immunohistochemistry (IHC) staining was performed in a tissue microarray consisted of 228 HCC peri- and intra-tumor tissue, NMU representative IHC images are shown. Bar, 20 μm. ** p

    Article Snippet: NMU protein was then detected in the serum using an ELISA kit (Cloud-Clone Corp.), according to the manufacturer’s instructions.

    Techniques: Enzyme-linked Immunosorbent Assay, Immunohistochemistry, Staining, Microarray

    Levels of HNP-1–4 in nasal aspirates from children with naturally occurring adenovirus infection (symptomatic) and when well (asymptomatic). HNP-1–4 were measured by ELISA in nasal aspirates. Statistical analysis and Mann–Whitney U test were perfomed.

    Journal: Canadian Respiratory Journal

    Article Title: Human Neutrophil Defensin-1, -3, and -4 Are Elevated in Nasal Aspirates from Children with Naturally Occurring Adenovirus Infection

    doi: 10.1155/2018/1038593

    Figure Lengend Snippet: Levels of HNP-1–4 in nasal aspirates from children with naturally occurring adenovirus infection (symptomatic) and when well (asymptomatic). HNP-1–4 were measured by ELISA in nasal aspirates. Statistical analysis and Mann–Whitney U test were perfomed.

    Article Snippet: HNP-1–4 Measurements α -Defensins were measured in nasal aspirates using specific ELISA kits purchased from Cloud-Clone Corp (alpha-defensins-1 and -3) and MyBioSource (MBS) (alpha-defensins-2 and -4).

    Techniques: Infection, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

    Correlation between cELISA and the HI test and the commercial ELISA kit (IDEXX). a Serum titers for antibodies against NDV detected between the cELISA (PI) and HI test (log2). b Titres of antibodies against NDV in different dilution serum samples detected between the cELISA (PI) and commercial ELISA kit test (S/P)

    Journal: Journal of Nanobiotechnology

    Article Title: Nanobody-horseradish peroxidase fusion protein as an ultrasensitive probe to detect antibodies against Newcastle disease virus in the immunoassay

    doi: 10.1186/s12951-019-0468-0

    Figure Lengend Snippet: Correlation between cELISA and the HI test and the commercial ELISA kit (IDEXX). a Serum titers for antibodies against NDV detected between the cELISA (PI) and HI test (log2). b Titres of antibodies against NDV in different dilution serum samples detected between the cELISA (PI) and commercial ELISA kit test (S/P)

    Article Snippet: Agreements of competitive ELISA with HI test and with commercial ELISA kit After testing the 368 clinical chicken sera separately using the cELISA, HI test, and commercial ELISA kit, the positive rates of each method were 97.83% (360/368), 100% (368/368), and 99.18% (365/368) (Table ), respectively.

    Techniques: Enzyme-linked Immunosorbent Assay

    EREG stimulates leptin secretion and lipolysis in mice with diet-induced obesity (DIO). (A) DIO WT male mice (n=7/ group) were injected with 100μL PBS (Veh) with and without EREG (1.5ng/g body weight or 20ng/per iAb depot) into both epididymal iAb fat pads every other day for 2 weeks. Total mRNA was isolated from one whole iAb fad pad. Expression of Lep was measured using NanoString assay. Normalized data represent mean±SD, n=5; Mann-Whitney U test. (B) Expression of LEP protein levels were measured in plasma in the same mice group by ELISA. Data (mean±SD, n=5/group). Independent Student’s t -test. (C) Non-esterified fatty acids (NEFA) and (D) TG concentrations in plasma are shown as mean ± SD, n=5. Independent Student’s t -test.

    Journal: The Journal of endocrinology

    Article Title: Epiregulin induces leptin secretion and energy expenditure in high-fat diet-fed mice

    doi: 10.1530/JOE-18-0289

    Figure Lengend Snippet: EREG stimulates leptin secretion and lipolysis in mice with diet-induced obesity (DIO). (A) DIO WT male mice (n=7/ group) were injected with 100μL PBS (Veh) with and without EREG (1.5ng/g body weight or 20ng/per iAb depot) into both epididymal iAb fat pads every other day for 2 weeks. Total mRNA was isolated from one whole iAb fad pad. Expression of Lep was measured using NanoString assay. Normalized data represent mean±SD, n=5; Mann-Whitney U test. (B) Expression of LEP protein levels were measured in plasma in the same mice group by ELISA. Data (mean±SD, n=5/group). Independent Student’s t -test. (C) Non-esterified fatty acids (NEFA) and (D) TG concentrations in plasma are shown as mean ± SD, n=5. Independent Student’s t -test.

    Article Snippet: Levels of LEP in iAb fat lysates produced and secreted into the medium were determined using ELISA kit (Crystal Chem, Elk Grove Village, IL).

    Techniques: Mouse Assay, Injection, Isolation, Expressing, MANN-WHITNEY, Enzyme-linked Immunosorbent Assay

    EREG/EGFR/MAPK axes induced leptin (LEP) secretion from adipose tissues ex vivo. Five iAb epididymal fat explants were excised from WT mice and stimulated. After 2h of stimulation LEP levels were measured in the media by ELISA. This experiment was repeated in three different WT mice. (A) Concentration-dependent secretion of LEP in the media from iAb fat explants stimulated with different concentrations of EREG. Data (mean±SD, n=5) are shown as percent to control (Veh, 100 %). Kruskal Wallis test. (B, C) LEP concentrations in the iAb fat lysates (B) and media (C) released from iAb fat before (triangles) and after (circles) stimulation with EREG (black bar, 50 ng/ml). Dashed and solid lines show mean value (n=3 mice) before and after stimulation. (D) Expression levels of Egfr and Erbb4 in epididymal iAb fat of lean WT mice (mean±SD, n=5) were measured by RT-PCR and normalized by TBP. (E) LEP concentrations in the iAb fat stimulated with EREG (black bar, 50 ng/ml) in the presence and absence of inhibitors of EGFR (10 μM) and MAPK (10 μM). Data (mean±SD, n=5) are shown as percent to control (Veh, white bar, 100%, dashed line). Between subject ANOVA with post-hoc Fisher LSD group comparison (α=0.05). (F) LEP in the media released following stimulation of mouse explants with and without EREG (50 ng/mL) in the presence and absence of inhibitors of HSL (10 μM), PPARα (10 μM), and PI3K (100 nM). Data (mean±SD, n=5) are shown as percent of control (Veh, 100%, dashed line). Between subject ANOVA. ns : not significant.

    Journal: The Journal of endocrinology

    Article Title: Epiregulin induces leptin secretion and energy expenditure in high-fat diet-fed mice

    doi: 10.1530/JOE-18-0289

    Figure Lengend Snippet: EREG/EGFR/MAPK axes induced leptin (LEP) secretion from adipose tissues ex vivo. Five iAb epididymal fat explants were excised from WT mice and stimulated. After 2h of stimulation LEP levels were measured in the media by ELISA. This experiment was repeated in three different WT mice. (A) Concentration-dependent secretion of LEP in the media from iAb fat explants stimulated with different concentrations of EREG. Data (mean±SD, n=5) are shown as percent to control (Veh, 100 %). Kruskal Wallis test. (B, C) LEP concentrations in the iAb fat lysates (B) and media (C) released from iAb fat before (triangles) and after (circles) stimulation with EREG (black bar, 50 ng/ml). Dashed and solid lines show mean value (n=3 mice) before and after stimulation. (D) Expression levels of Egfr and Erbb4 in epididymal iAb fat of lean WT mice (mean±SD, n=5) were measured by RT-PCR and normalized by TBP. (E) LEP concentrations in the iAb fat stimulated with EREG (black bar, 50 ng/ml) in the presence and absence of inhibitors of EGFR (10 μM) and MAPK (10 μM). Data (mean±SD, n=5) are shown as percent to control (Veh, white bar, 100%, dashed line). Between subject ANOVA with post-hoc Fisher LSD group comparison (α=0.05). (F) LEP in the media released following stimulation of mouse explants with and without EREG (50 ng/mL) in the presence and absence of inhibitors of HSL (10 μM), PPARα (10 μM), and PI3K (100 nM). Data (mean±SD, n=5) are shown as percent of control (Veh, 100%, dashed line). Between subject ANOVA. ns : not significant.

    Article Snippet: Levels of LEP in iAb fat lysates produced and secreted into the medium were determined using ELISA kit (Crystal Chem, Elk Grove Village, IL).

    Techniques: Ex Vivo, Mouse Assay, Enzyme-linked Immunosorbent Assay, Concentration Assay, Expressing, Reverse Transcription Polymerase Chain Reaction

    Western blot analysis of the effect of Sanguinarine on CEM/ADR5000 leukemia cells. Evaluation of the P-gp, NFκB, and IκBα expressions. β-actin was used as loading control. Bands were normalized to β-actin in order to obtain numerical values (Mean ± SD ).

    Journal: Frontiers in Pharmacology

    Article Title: Molecular Determinants of Sensitivity or Resistance of Cancer Cells Toward Sanguinarine

    doi: 10.3389/fphar.2018.00136

    Figure Lengend Snippet: Western blot analysis of the effect of Sanguinarine on CEM/ADR5000 leukemia cells. Evaluation of the P-gp, NFκB, and IκBα expressions. β-actin was used as loading control. Bands were normalized to β-actin in order to obtain numerical values (Mean ± SD ).

    Article Snippet: Membranes were incubated at 4°C overnight with primary antibodies including: P-gp (1:1000) (Thermoscientific, Darmstadt, Germany), NFκB (1:1000) (Cell Signaling Technology, Frankfurt, Germany), IκBα (1:1000) (Cell Signaling Technology, Frankfurt, Germany), and β-actin (1:2000) (Cell Signaling Technology, Frankfurt, Germany).

    Techniques: Western Blot

    Enho mRNA and protein levels are downregulated in the aged rat brain and are associated with decreased adropin levels in plasma A) Enho mRNA expression was significantly decreased in the cerebral cortex of aged rats compared to young rats. B) Cortical homogenates (50 µg of total protein) from rat brains were subjected to SDS-polyacrylamide gel electrophoresis and endogenous adropin protein levels were detected by Western blot. Representative immunoblots and densitometric data showed that adropin protein levels in the cerebral cortex were dramatically decreased in aged rats compared to young rats. C) Adropin antibody is highly specific. Cortical brain homogenates (samples 1 and 2) were denatured at 70°C for 10 min in 2X Laemmli sample buffer containing 4% β-mercaptoethanol, separated on 4-20% SDS-polyacrylamide gels, and then transferred onto nitrocellulose membranes. Before incubation with the membrane, the adropin monoclonal antibody was pre-absorbed with different concentrations of synthetic adropin 34-76 peptide (0-300 ng/ml), which resulted in a concentration-dependent loss of the intensity of the band corresponding to endogenous brain adropin. D) Plasma adropin levels were measured by a commercial adropin ELISA kit. A significant decrease in adropin levels was observed in aged rats compared with young controls. * P

    Journal: Aging and Disease

    Article Title: Age-Dependent Decrease in Adropin is Associated with Reduced Levels of Endothelial Nitric Oxide Synthase and Increased Oxidative Stress in the Rat Brain

    doi: 10.14336/AD.2017.0523

    Figure Lengend Snippet: Enho mRNA and protein levels are downregulated in the aged rat brain and are associated with decreased adropin levels in plasma A) Enho mRNA expression was significantly decreased in the cerebral cortex of aged rats compared to young rats. B) Cortical homogenates (50 µg of total protein) from rat brains were subjected to SDS-polyacrylamide gel electrophoresis and endogenous adropin protein levels were detected by Western blot. Representative immunoblots and densitometric data showed that adropin protein levels in the cerebral cortex were dramatically decreased in aged rats compared to young rats. C) Adropin antibody is highly specific. Cortical brain homogenates (samples 1 and 2) were denatured at 70°C for 10 min in 2X Laemmli sample buffer containing 4% β-mercaptoethanol, separated on 4-20% SDS-polyacrylamide gels, and then transferred onto nitrocellulose membranes. Before incubation with the membrane, the adropin monoclonal antibody was pre-absorbed with different concentrations of synthetic adropin 34-76 peptide (0-300 ng/ml), which resulted in a concentration-dependent loss of the intensity of the band corresponding to endogenous brain adropin. D) Plasma adropin levels were measured by a commercial adropin ELISA kit. A significant decrease in adropin levels was observed in aged rats compared with young controls. * P

    Article Snippet: Measurement of adropin levels in rat plasma Adropin levels in rat plasma were quantified using an ELISA kit (Cat. No. EK-032-35, Phoenix Pharmaceuticals, Inc., Burlingame, CA) as recommended by the manufacturer’s protocol.

    Techniques: Expressing, Polyacrylamide Gel Electrophoresis, Western Blot, Incubation, Concentration Assay, Enzyme-linked Immunosorbent Assay