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    Santa Cruz Biotechnology horseradish peroxidase hrp linked goat anti mouse igg
    Horseradish Peroxidase Hrp Linked Goat Anti Mouse Igg, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 42 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology horseradish peroxidase linked anti mouse igg
    Microfilament inhibitors suppress the AngII-mediated increase in COX-2 expression by preventing HuR binding to the corresponding mRNA. ( A ). HMC cells were treated for 2 h with either vehicle (−) or with AngII, or together with different microfilament inhibitors as indicated. Inhibitors were preincubated for 30 min (Lat.A) or 4 h (Blebbistat.) before the addition of AngII. To achieve transcriptional induction of different HuR target genes, cells were stimulated for 16 h with a cytokine mix containing IL-1β and TNFα (both at 2 nM) before AngII treatment. Steady-state mRNA levels of COX-2, cyclin A and cyclin D 1 were quantified by qRT-PCR using β-actin as a normalization control. Results are depicted as relative mRNA levels (%) in comparison with cytokine-stimulated conditions (vehicle), which were set to 100% and are means ± SD (n=3). ** P ≤ 0.01, *** P ≤ 0.005 compared with cytokine-induced conditions ## P ≤ 0.01 and ### P ≤ 0.005, compared with cytokine plus AngII-treated conditions. ( B ). Reduction of AngII-mediated stabilization of cytokine-triggered COX- 2 mRNA by blebbistatin (left panel) or latrunculin A (right panel). Quiescent HMC were treated for 20 h with a cytokine mixture containing IL-1β and TNFα (both at 2 nM) before the administration of Act D, which was added 30 min before AngII (filled triangles). Importantly, the indicated cytoskeletal inhibitors were given 4.5 h before the addition of AngII and remained for the indicated times with vehicle (open circles) or with 0.1 µM AngII (open squares) before cells were harvested and extracted for total cellular RNA. Steady-state mRNA levels of COX-2 were determined by qRT-PCR using β-actin mRNA as a normalization control. Graphs show means ± SD (n = 3) and depict the percentage of remaining COX-2 mRNA levels compared with the content of steady-state COX-2 mRNA measured immediately before the addition of Act D (0 h). ( C ). HMC were treated similarly as described in (A) before total cell lysates were assayed by IP using an anti-HuR antibody (gray bars), or the same amount of isotypic <t>IgG</t> (open bars). HuR-bound COX-2 mRNA was determined by pulldown RT assays. Data shown are means of two independent experiments and are depicted as fold induction versus cytokine-treated cells.
    Horseradish Peroxidase Linked Anti Mouse Igg, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 81/100, based on 119 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology horseradish peroxidase linked anti mouse immunoglobulin g
    Phospho-ezrin and ezrin expression in NPC metastatic nodes. Phospho-ezrin and ezrin were detected in the primary and metastatic NPC samples using immunohistochemistry. ( a , b ) primary NPC and metastatic tumor sections were stained with hematoxylin and eosin; ( c , d ) stained with antibodies against ezrin; ( e , f ) stained with phospho-ezrin at Thr567; ( g , h ) stained with <t>IgG,</t> served as a blank control. Arrow, positive cells. Original magnification, ×400. Scale bar, 5 μm. H E, hematoxylin and eosin.
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    Santa Cruz Biotechnology anti mouse igg horseradish peroxidase hrp linked
    Phospho-ezrin and ezrin expression in NPC metastatic nodes. Phospho-ezrin and ezrin were detected in the primary and metastatic NPC samples using immunohistochemistry. ( a , b ) primary NPC and metastatic tumor sections were stained with hematoxylin and eosin; ( c , d ) stained with antibodies against ezrin; ( e , f ) stained with phospho-ezrin at Thr567; ( g , h ) stained with <t>IgG,</t> served as a blank control. Arrow, positive cells. Original magnification, ×400. Scale bar, 5 μm. H E, hematoxylin and eosin.
    Anti Mouse Igg Horseradish Peroxidase Hrp Linked, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 77/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology horseradish peroxidase hrp linked antibodies
    Phospho-ezrin and ezrin expression in NPC metastatic nodes. Phospho-ezrin and ezrin were detected in the primary and metastatic NPC samples using immunohistochemistry. ( a , b ) primary NPC and metastatic tumor sections were stained with hematoxylin and eosin; ( c , d ) stained with antibodies against ezrin; ( e , f ) stained with phospho-ezrin at Thr567; ( g , h ) stained with <t>IgG,</t> served as a blank control. Arrow, positive cells. Original magnification, ×400. Scale bar, 5 μm. H E, hematoxylin and eosin.
    Horseradish Peroxidase Hrp Linked Antibodies, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 91/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology horseradish peroxidase linked secondary goat anti mouse igg
    Identification of a potent small molecule inhibitor of Plk1 PBD A. The chemical structure of T521: 5-(ethylsulfonyl)-2-(4-fluorophenyl)-4-(phenylsulfonyl) oxazole. B, C, D. The inhibitory effect of T521, Poloxin and TQ on the binding of the PBDs of Plk1, 2 and 3 to the FITC-labeled phosphopeptide was analyzed by FP assay. Briefly, T521, Poloxin or TQ was incubated with Plk1-3 PBD for 1 hr at room temperature (RT) prior to addition of FITC-labeled phosphopeptide. The inhibitory effect was calculated as described in the Materials and Methods. Error bars represent SD. E. ELISA-based PBD binding inhibition assay to determine the inhibitory effect of T521 on the interaction between Plk1 PBD and its binding target Map205 PBM . His-tagged PBD of Plk1 and different concentrations of T521 were added into GST-Map205 PBM -coated plates. After incubation, the plate was washed and then probed with mouse anti-His primary antibody, followed by goat anti-mouse <t>IgG-HRP</t> secondary antibody. After washing, the plate was incubated with TMB solution and the OD 450 was measured. Detailed procedure was described in Methods and Materials. DMSO was used as control. F. T521 inhibits Bub1-Plk1 interaction in vivo using Co-immunoprecipitation (IP) assay. HeLa cells synchronized by double-thymidine block were released into medium containing DMSO or T521 at indicated concentrations for 10 hrs, and then lysed and subject to Bub1 IP. The proteins in the precipitates were separated by 10% SDS-PAGE and probed with the anti-hPlk1 and anti-Bub1 antibodies. Moreover, the Bub1 and Plk1 levels were also determined in the lysates of HeLa cells before IP. β-actin served as the loading control.
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    Santa Cruz Biotechnology hrp linked anti mouse rabbit igg
    Immunoreactivity of various preparations by indirect ELISA Filled circles: 2C5 antibody, Circles: 2C5-modified doxorubicin-loaded micelles, Filled squares: Isotype-matched <t>IgG,</t> Squares: IgG-modified doxorubicin-loaded micelles . The binding of doxorubicin-loaded PEG-PE micelles harboring 2C5 antibody or isotype antibody to a monolayer of the antigen (nucleohistones) was evaluated by ELISA and detected using an <t>HRP</t> conjugated antibody. Data represent the mean ± SD, n=3.
    Hrp Linked Anti Mouse Rabbit Igg, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology hrp linked donkey anti mouse igg
    Immunoreactivity of various preparations by indirect ELISA Filled circles: 2C5 antibody, Circles: 2C5-modified doxorubicin-loaded micelles, Filled squares: Isotype-matched <t>IgG,</t> Squares: IgG-modified doxorubicin-loaded micelles . The binding of doxorubicin-loaded PEG-PE micelles harboring 2C5 antibody or isotype antibody to a monolayer of the antigen (nucleohistones) was evaluated by ELISA and detected using an <t>HRP</t> conjugated antibody. Data represent the mean ± SD, n=3.
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    Santa Cruz Biotechnology anti mouse igg hrp linked
    Immunoreactivity of various preparations by indirect ELISA Filled circles: 2C5 antibody, Circles: 2C5-modified doxorubicin-loaded micelles, Filled squares: Isotype-matched <t>IgG,</t> Squares: IgG-modified doxorubicin-loaded micelles . The binding of doxorubicin-loaded PEG-PE micelles harboring 2C5 antibody or isotype antibody to a monolayer of the antigen (nucleohistones) was evaluated by ELISA and detected using an <t>HRP</t> conjugated antibody. Data represent the mean ± SD, n=3.
    Anti Mouse Igg Hrp Linked, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 39 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology anti mouse horseradish peroxidase linked iggs
    Immunoreactivity of various preparations by indirect ELISA Filled circles: 2C5 antibody, Circles: 2C5-modified doxorubicin-loaded micelles, Filled squares: Isotype-matched <t>IgG,</t> Squares: IgG-modified doxorubicin-loaded micelles . The binding of doxorubicin-loaded PEG-PE micelles harboring 2C5 antibody or isotype antibody to a monolayer of the antigen (nucleohistones) was evaluated by ELISA and detected using an <t>HRP</t> conjugated antibody. Data represent the mean ± SD, n=3.
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    Santa Cruz Biotechnology horseradish peroxidase linked igg secondary antibodies
    Immunoreactivity of various preparations by indirect ELISA Filled circles: 2C5 antibody, Circles: 2C5-modified doxorubicin-loaded micelles, Filled squares: Isotype-matched <t>IgG,</t> Squares: IgG-modified doxorubicin-loaded micelles . The binding of doxorubicin-loaded PEG-PE micelles harboring 2C5 antibody or isotype antibody to a monolayer of the antigen (nucleohistones) was evaluated by ELISA and detected using an <t>HRP</t> conjugated antibody. Data represent the mean ± SD, n=3.
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    Santa Cruz Biotechnology hrp linked mouse anti rabbit igg polyclonal secondary antibody sc 2357
    Immunoreactivity of various preparations by indirect ELISA Filled circles: 2C5 antibody, Circles: 2C5-modified doxorubicin-loaded micelles, Filled squares: Isotype-matched <t>IgG,</t> Squares: IgG-modified doxorubicin-loaded micelles . The binding of doxorubicin-loaded PEG-PE micelles harboring 2C5 antibody or isotype antibody to a monolayer of the antigen (nucleohistones) was evaluated by ELISA and detected using an <t>HRP</t> conjugated antibody. Data represent the mean ± SD, n=3.
    Hrp Linked Mouse Anti Rabbit Igg Polyclonal Secondary Antibody Sc 2357, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 85/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology hrp linked mouse monoclonal igg his probe antibody h 3
    Functional significance of trimeric contacts in CaiT. (a) Western blot analysis of CaiT with given amino acid replacements in E. coli JW0039 membranes. <t>HRP-linked</t> mouse anti-His <t>IgG</t> was used to detect CaiT. PageRuler Prestained Protein Ladder was used for molecular size estimation. Shown is the section of the blot that contains the CaiT band. The complete gel is presented in Fig. S6. (b) Initial rates of counterflow (black) and maximum accumulation of carnitine in E. coli JW0039 (white) were determined as described in the legend of Fig. 3 . As negative control, cells carrying pET21a ( nc ) were used. (c) Initial rates of counterflow (black) and maximum accumulation (white) of carnitine catalyzed by CaiT variants in proteoliposomes. CaiT was purified and reconstituted as described 1 . The lipid to protein ratio of the resulting proteoliposomes was 100:1 (w/w). Proteoliposomes were preloaded with 10 mM unlabeled L-carnitine overnight. Aliquots of the proteoliposome suspension (1–2 µg of protein) were then diluted into 400 µL buffer containing 4.5 µM L-[methyl- 14 C]carnitine (55 Ci/mol) resulting in a final carnitine concentration of 54.5 µM. As negative control, proteoliposomes not preloaded with unlabeled carnitine ( nc ) were used and the obtained transport rates were used for correcting the data. Carnitine counterflow with cells or proteoliposomes was assayed at 25 °C under aerobic conditions using a rapid filtration method as described 1 . (d) Michaelis-Menten kinetics of L-carnitine counterflow in proteoliposomes reconstituted with wild-type CaiT (WT) or CaiT-D288A. Initial rates of L-[methyl- 14 C]carnitine uptake were measured at given substrate concentrations and plotted using the kinetic module of GraphPad Prism. For all experiments, standard deviations were calculated from triplicate determinations of a representative experiment.
    Hrp Linked Mouse Monoclonal Igg His Probe Antibody H 3, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 89/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology hrp linked secondary antibodies
    Functional significance of trimeric contacts in CaiT. (a) Western blot analysis of CaiT with given amino acid replacements in E. coli JW0039 membranes. <t>HRP-linked</t> mouse anti-His <t>IgG</t> was used to detect CaiT. PageRuler Prestained Protein Ladder was used for molecular size estimation. Shown is the section of the blot that contains the CaiT band. The complete gel is presented in Fig. S6. (b) Initial rates of counterflow (black) and maximum accumulation of carnitine in E. coli JW0039 (white) were determined as described in the legend of Fig. 3 . As negative control, cells carrying pET21a ( nc ) were used. (c) Initial rates of counterflow (black) and maximum accumulation (white) of carnitine catalyzed by CaiT variants in proteoliposomes. CaiT was purified and reconstituted as described 1 . The lipid to protein ratio of the resulting proteoliposomes was 100:1 (w/w). Proteoliposomes were preloaded with 10 mM unlabeled L-carnitine overnight. Aliquots of the proteoliposome suspension (1–2 µg of protein) were then diluted into 400 µL buffer containing 4.5 µM L-[methyl- 14 C]carnitine (55 Ci/mol) resulting in a final carnitine concentration of 54.5 µM. As negative control, proteoliposomes not preloaded with unlabeled carnitine ( nc ) were used and the obtained transport rates were used for correcting the data. Carnitine counterflow with cells or proteoliposomes was assayed at 25 °C under aerobic conditions using a rapid filtration method as described 1 . (d) Michaelis-Menten kinetics of L-carnitine counterflow in proteoliposomes reconstituted with wild-type CaiT (WT) or CaiT-D288A. Initial rates of L-[methyl- 14 C]carnitine uptake were measured at given substrate concentrations and plotted using the kinetic module of GraphPad Prism. For all experiments, standard deviations were calculated from triplicate determinations of a representative experiment.
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    Santa Cruz Biotechnology mouse igg
    Functional significance of trimeric contacts in CaiT. (a) Western blot analysis of CaiT with given amino acid replacements in E. coli JW0039 membranes. <t>HRP-linked</t> mouse anti-His <t>IgG</t> was used to detect CaiT. PageRuler Prestained Protein Ladder was used for molecular size estimation. Shown is the section of the blot that contains the CaiT band. The complete gel is presented in Fig. S6. (b) Initial rates of counterflow (black) and maximum accumulation of carnitine in E. coli JW0039 (white) were determined as described in the legend of Fig. 3 . As negative control, cells carrying pET21a ( nc ) were used. (c) Initial rates of counterflow (black) and maximum accumulation (white) of carnitine catalyzed by CaiT variants in proteoliposomes. CaiT was purified and reconstituted as described 1 . The lipid to protein ratio of the resulting proteoliposomes was 100:1 (w/w). Proteoliposomes were preloaded with 10 mM unlabeled L-carnitine overnight. Aliquots of the proteoliposome suspension (1–2 µg of protein) were then diluted into 400 µL buffer containing 4.5 µM L-[methyl- 14 C]carnitine (55 Ci/mol) resulting in a final carnitine concentration of 54.5 µM. As negative control, proteoliposomes not preloaded with unlabeled carnitine ( nc ) were used and the obtained transport rates were used for correcting the data. Carnitine counterflow with cells or proteoliposomes was assayed at 25 °C under aerobic conditions using a rapid filtration method as described 1 . (d) Michaelis-Menten kinetics of L-carnitine counterflow in proteoliposomes reconstituted with wild-type CaiT (WT) or CaiT-D288A. Initial rates of L-[methyl- 14 C]carnitine uptake were measured at given substrate concentrations and plotted using the kinetic module of GraphPad Prism. For all experiments, standard deviations were calculated from triplicate determinations of a representative experiment.
    Mouse Igg, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 3845 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology rabbit hrp linked igg1 secondary antibodies
    Functional significance of trimeric contacts in CaiT. (a) Western blot analysis of CaiT with given amino acid replacements in E. coli JW0039 membranes. <t>HRP-linked</t> mouse anti-His <t>IgG</t> was used to detect CaiT. PageRuler Prestained Protein Ladder was used for molecular size estimation. Shown is the section of the blot that contains the CaiT band. The complete gel is presented in Fig. S6. (b) Initial rates of counterflow (black) and maximum accumulation of carnitine in E. coli JW0039 (white) were determined as described in the legend of Fig. 3 . As negative control, cells carrying pET21a ( nc ) were used. (c) Initial rates of counterflow (black) and maximum accumulation (white) of carnitine catalyzed by CaiT variants in proteoliposomes. CaiT was purified and reconstituted as described 1 . The lipid to protein ratio of the resulting proteoliposomes was 100:1 (w/w). Proteoliposomes were preloaded with 10 mM unlabeled L-carnitine overnight. Aliquots of the proteoliposome suspension (1–2 µg of protein) were then diluted into 400 µL buffer containing 4.5 µM L-[methyl- 14 C]carnitine (55 Ci/mol) resulting in a final carnitine concentration of 54.5 µM. As negative control, proteoliposomes not preloaded with unlabeled carnitine ( nc ) were used and the obtained transport rates were used for correcting the data. Carnitine counterflow with cells or proteoliposomes was assayed at 25 °C under aerobic conditions using a rapid filtration method as described 1 . (d) Michaelis-Menten kinetics of L-carnitine counterflow in proteoliposomes reconstituted with wild-type CaiT (WT) or CaiT-D288A. Initial rates of L-[methyl- 14 C]carnitine uptake were measured at given substrate concentrations and plotted using the kinetic module of GraphPad Prism. For all experiments, standard deviations were calculated from triplicate determinations of a representative experiment.
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    Santa Cruz Biotechnology hrp conjugated anti mouse igg
    Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific <t>IgG</t> antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using <t>HRP-conjugated</t> anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P
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    Santa Cruz Biotechnology peroxidase linked goat anti mouse igg
    Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific <t>IgG</t> antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using <t>HRP-conjugated</t> anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P
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    Santa Cruz Biotechnology peroxidase linked secondary antibody goat anti mouse igg hrp
    Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific <t>IgG</t> antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using <t>HRP-conjugated</t> anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P
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    Santa Cruz Biotechnology anti mouse igg2b hrp mab
    Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific <t>IgG</t> antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using <t>HRP-conjugated</t> anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P
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    Santa Cruz Biotechnology peroxidase linked anti mouse igg
    Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific <t>IgG</t> antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using <t>HRP-conjugated</t> anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P
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    Santa Cruz Biotechnology horseradish peroxidase conjugated goat anti mouse igg
    Specific humoral responses elicited by immunization of mice with microneme proteins. The reactivity of immunoglobulins anti-STAg was determined by ELISA in serum samples collected from both immunized (TgMICs) and control (PBS) mice 15 days after the last antigen injection. Each point/bar represents the average absorbance ± SD of the serum samples from 4 animals. (A) Absorbance provided by the reaction of serum <t>IgG</t> with STAg. (B) Absorbance provided by the reaction of serum <t>IgG1</t> and IgG2a (diluted 1:25) with STAg. The average absorbance ± SD generated by the reaction of serum IgG1 or IgG2a from each group of immunized mice was significantly higher than the corresponding values provided by control mice, with the exception of the TgMIC6-immunized group, whose results were not significantly different of those of the control group. Three independent experiments were performed, and data from one representative experiment is shown. Asterisks represent statistical significant differences (*p
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    Santa Cruz Biotechnology chicken anti mouse igg horse radish peroxidase linked antibodies
    Specific humoral responses elicited by immunization of mice with microneme proteins. The reactivity of immunoglobulins anti-STAg was determined by ELISA in serum samples collected from both immunized (TgMICs) and control (PBS) mice 15 days after the last antigen injection. Each point/bar represents the average absorbance ± SD of the serum samples from 4 animals. (A) Absorbance provided by the reaction of serum <t>IgG</t> with STAg. (B) Absorbance provided by the reaction of serum <t>IgG1</t> and IgG2a (diluted 1:25) with STAg. The average absorbance ± SD generated by the reaction of serum IgG1 or IgG2a from each group of immunized mice was significantly higher than the corresponding values provided by control mice, with the exception of the TgMIC6-immunized group, whose results were not significantly different of those of the control group. Three independent experiments were performed, and data from one representative experiment is shown. Asterisks represent statistical significant differences (*p
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    Santa Cruz Biotechnology goat anti mouse igg2a hrp secondary antibodies
    Specific humoral responses elicited by immunization of mice with microneme proteins. The reactivity of immunoglobulins anti-STAg was determined by ELISA in serum samples collected from both immunized (TgMICs) and control (PBS) mice 15 days after the last antigen injection. Each point/bar represents the average absorbance ± SD of the serum samples from 4 animals. (A) Absorbance provided by the reaction of serum <t>IgG</t> with STAg. (B) Absorbance provided by the reaction of serum <t>IgG1</t> and IgG2a (diluted 1:25) with STAg. The average absorbance ± SD generated by the reaction of serum IgG1 or IgG2a from each group of immunized mice was significantly higher than the corresponding values provided by control mice, with the exception of the TgMIC6-immunized group, whose results were not significantly different of those of the control group. Three independent experiments were performed, and data from one representative experiment is shown. Asterisks represent statistical significant differences (*p
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    Santa Cruz Biotechnology hrp conjugated goat anti mouse igg1
    Specific binding ELISA of hE16 to plant-derived DIII. Serial dilutions of hE16 purified from mammalian or plant cells were incubated in sample wells coated with plant-produced WNV DIII and detected with an <t>HRP-conjugated</t> anti-human gamma antibody. A commercial generic human <t>IgG</t> was used as a negative control. Mean ± SD of samples from three independent experiments is presented.
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    Santa Cruz Biotechnology peroxidase conjugated goat anti mouse immunoglobulin g igg antibody
    In vivo immunogenicity of K. pneumoniae EVs. ( a ) K. pneumoniae EV-reactive <t>IgG</t> antibody in serum from mice immunized with 10, 100, 100 ng of K. pneumoniae EVs. This antibody was measured by ELISA ( n =5; * P
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    Santa Cruz Biotechnology horseradish peroxidase hrp conjugated goat against mouse igg
    In vivo immunogenicity of K. pneumoniae EVs. ( a ) K. pneumoniae EV-reactive <t>IgG</t> antibody in serum from mice immunized with 10, 100, 100 ng of K. pneumoniae EVs. This antibody was measured by ELISA ( n =5; * P
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    Santa Cruz Biotechnology goat anti mouse igg hrp conjugate
    In vivo immunogenicity of K. pneumoniae EVs. ( a ) K. pneumoniae EV-reactive <t>IgG</t> antibody in serum from mice immunized with 10, 100, 100 ng of K. pneumoniae EVs. This antibody was measured by ELISA ( n =5; * P
    Goat Anti Mouse Igg Hrp Conjugate, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 84 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology horseradish peroxidase hrp conjugated anti mouse igg
    Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific <t>IgG</t> antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using <t>HRP-conjugated</t> anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P
    Horseradish Peroxidase Hrp Conjugated Anti Mouse Igg, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 139 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology hrp conjugated bovine anti goat igg ab
    Expression of CD23 in mouse AECs Cell lysates (50 µg) derived from primary nasal ( A ), tracheal ( B ), and lung ( B ) epithelial cells were electrophoresed on a 12 % SDS-PAGE gel under reducing condition. The separated proteins were transferred onto a nitrocellulose membrane, blotted with rat anti-mouse CD23 mAb (B3B4) followed by <t>HRP-conjugated</t> rabbit anti-rat <t>IgG</t> Ab. Proteins were visualized using the ECL method. Lysates (50 µg) from mouse spleen or CHO cells were used as a positive or negative control, respectively. β-tubulin was used as an internal control. Arrows indicate mouse CD23 and β-tubulin. ( C ) Immunohistochemical staining of mouse lung. Normal mouse lung was prepared in OCT medium and cryosectioned at 5 µm. Frozen tissue sections were fixed and permeabilized with ice-cold acetone and blocked with 10% normal goat serum. A spleen section was used as a positive control. Sections were incubated with rabbit anti-CD23 Ab or normal rabbit IgG, followed by staining with Alexa Fluor 555-conjugated goat anti-rabbit Ab. Nuclei were stained with DAPI. Images were captured using a Zeiss LSM510 confocal microscope. Samples were visualized under consisten contrast and brightness settings.
    Hrp Conjugated Bovine Anti Goat Igg Ab, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 77/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology hrp horseradish peroxidase conjugated goat anti rabbit igg antibody
    Production of autoantibodies to erythrocytes in SOD1-deficient mice ( A ) Detection of <t>IgG</t> bound to erythrocytes from SOD1 −/− mice. Collected erythrocytes were reacted with FITC-labelled anti-mouse IgG followed by FACS analysis. Relative values of bound IgG are shown. n =7 for SOD1 +/+ and n =5 for SOD1 −/− . ( B ) Cytosolic and membrane fractions of RBCs were incubated with blood plasma from SOD1 +/+ and SOD1 −/− mice (1:50 dilution) followed by reaction with an <t>HRP-conjugated</t> anti-mouse IgG. ( C ) Immunoblot analysis using blood plasma from an SOD1 −/− mouse and serum of a rabbit that had been immunized with bovine CAII. Western blot analysis of human CAII and RBCs with blood plasma from a SOD1 −/− mouse at 40 weeks of age (1:50 dilution; upper panel). While mass marker b contains CAII, mass marker a does not. Characteristics of an anti-bovine CAII antibody (1:1000 dilution) raised in rabbit by immunoblotting of the same samples (lower panel); representative of several experiments. ( D ) Immunoprecipitation of bovine CAII and lysate of mouse RBCs with an anti-bovine CAII polyclonal antibody. Immunoblotting was performed with blood plasma from SOD1 −/− mice and an anti-CAII monoclonal antibody (mAb). Representative data from several experiments. ( E ) The titres of the anti-CAII antibody in blood plasma were elevated, as judged by ELISA. The inset shows an enlargement of the results obtained for SOD1 +/+ and SOD1 −/− mice at 14 weeks of age.
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    Santa Cruz Biotechnology hrp conjugated donkey anti goat igg ab
    effect of drug treatment on Hsc70 and protein disulphide isomerase (PDI) expression in small intestinal villi isolated from ECwt-infected mice. A: flow cytometric analysis was performed on intestinal epithelial cells (IEC) isolated from villi from ECwt-infected mice (n = 2 mice for each experimental group) that had been treated with ibuprofen (IBF) (20 mg/kg/day), diclofenac (DCF) (1 mg/kg/day), pioglitazone (PGZ)(30 mg/kg/day), rosiglitazone (RGZ) (4 mg/kg/day), N-acetylcysteine (NAC) (18 mg/kg/day), ascorbic acid (AA) (20 mg/kg/day) or difenoxilate sodium (7.5 mg/kg/day) during three days post-inoculation (d.p.i.). Goat primary antibodies (Abs) against Hsc70 or PDI and fluorescein isothiocyanate (FITC)-conjugated mouse anti-goat <t>IgG</t> secondary Abs were used. Immunofluorescence analysis was performed on a Cyan (Dako) flow cytometer using a FlowJo software; B: radio immunoprecipitation assay lysates from villi isolated from ECwt-infected mice (n = 3 mice for each experimental group) that had been treated with NAC (18 mg/kg/day) during three days after 24 h post-inoculation. were added to ELISA plates coated with rabbit polyclonal Abs against Hsc70 or PDI. Goat Abs against Hsc70 or PDI were used as detection Abs and the reaction was revealed using horseradish peroxidase <t>(HRP)-conjugated</t> donkey anti-goat Abs and phenylenediamine dihydrochloride substrate before reading at optical density (OD) 492 nm . Lysates from uninfected villi were used as a control. Data are expressed as mean OD 492 nm Hsc70 or PDI antigen. Graph shows significant increase of Hsc70 and PDI expression (p
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    Image Search Results


    Microfilament inhibitors suppress the AngII-mediated increase in COX-2 expression by preventing HuR binding to the corresponding mRNA. ( A ). HMC cells were treated for 2 h with either vehicle (−) or with AngII, or together with different microfilament inhibitors as indicated. Inhibitors were preincubated for 30 min (Lat.A) or 4 h (Blebbistat.) before the addition of AngII. To achieve transcriptional induction of different HuR target genes, cells were stimulated for 16 h with a cytokine mix containing IL-1β and TNFα (both at 2 nM) before AngII treatment. Steady-state mRNA levels of COX-2, cyclin A and cyclin D 1 were quantified by qRT-PCR using β-actin as a normalization control. Results are depicted as relative mRNA levels (%) in comparison with cytokine-stimulated conditions (vehicle), which were set to 100% and are means ± SD (n=3). ** P ≤ 0.01, *** P ≤ 0.005 compared with cytokine-induced conditions ## P ≤ 0.01 and ### P ≤ 0.005, compared with cytokine plus AngII-treated conditions. ( B ). Reduction of AngII-mediated stabilization of cytokine-triggered COX- 2 mRNA by blebbistatin (left panel) or latrunculin A (right panel). Quiescent HMC were treated for 20 h with a cytokine mixture containing IL-1β and TNFα (both at 2 nM) before the administration of Act D, which was added 30 min before AngII (filled triangles). Importantly, the indicated cytoskeletal inhibitors were given 4.5 h before the addition of AngII and remained for the indicated times with vehicle (open circles) or with 0.1 µM AngII (open squares) before cells were harvested and extracted for total cellular RNA. Steady-state mRNA levels of COX-2 were determined by qRT-PCR using β-actin mRNA as a normalization control. Graphs show means ± SD (n = 3) and depict the percentage of remaining COX-2 mRNA levels compared with the content of steady-state COX-2 mRNA measured immediately before the addition of Act D (0 h). ( C ). HMC were treated similarly as described in (A) before total cell lysates were assayed by IP using an anti-HuR antibody (gray bars), or the same amount of isotypic IgG (open bars). HuR-bound COX-2 mRNA was determined by pulldown RT assays. Data shown are means of two independent experiments and are depicted as fold induction versus cytokine-treated cells.

    Journal: Nucleic Acids Research

    Article Title: RNA-dependent association with myosin IIA promotes F-actin-guided trafficking of the ELAV-like protein HuR to polysomes

    doi: 10.1093/nar/gkt663

    Figure Lengend Snippet: Microfilament inhibitors suppress the AngII-mediated increase in COX-2 expression by preventing HuR binding to the corresponding mRNA. ( A ). HMC cells were treated for 2 h with either vehicle (−) or with AngII, or together with different microfilament inhibitors as indicated. Inhibitors were preincubated for 30 min (Lat.A) or 4 h (Blebbistat.) before the addition of AngII. To achieve transcriptional induction of different HuR target genes, cells were stimulated for 16 h with a cytokine mix containing IL-1β and TNFα (both at 2 nM) before AngII treatment. Steady-state mRNA levels of COX-2, cyclin A and cyclin D 1 were quantified by qRT-PCR using β-actin as a normalization control. Results are depicted as relative mRNA levels (%) in comparison with cytokine-stimulated conditions (vehicle), which were set to 100% and are means ± SD (n=3). ** P ≤ 0.01, *** P ≤ 0.005 compared with cytokine-induced conditions ## P ≤ 0.01 and ### P ≤ 0.005, compared with cytokine plus AngII-treated conditions. ( B ). Reduction of AngII-mediated stabilization of cytokine-triggered COX- 2 mRNA by blebbistatin (left panel) or latrunculin A (right panel). Quiescent HMC were treated for 20 h with a cytokine mixture containing IL-1β and TNFα (both at 2 nM) before the administration of Act D, which was added 30 min before AngII (filled triangles). Importantly, the indicated cytoskeletal inhibitors were given 4.5 h before the addition of AngII and remained for the indicated times with vehicle (open circles) or with 0.1 µM AngII (open squares) before cells were harvested and extracted for total cellular RNA. Steady-state mRNA levels of COX-2 were determined by qRT-PCR using β-actin mRNA as a normalization control. Graphs show means ± SD (n = 3) and depict the percentage of remaining COX-2 mRNA levels compared with the content of steady-state COX-2 mRNA measured immediately before the addition of Act D (0 h). ( C ). HMC were treated similarly as described in (A) before total cell lysates were assayed by IP using an anti-HuR antibody (gray bars), or the same amount of isotypic IgG (open bars). HuR-bound COX-2 mRNA was determined by pulldown RT assays. Data shown are means of two independent experiments and are depicted as fold induction versus cytokine-treated cells.

    Article Snippet: Antibodies raised against HuR, histone deacetylase-1 (HDAC1), c-myc, anti-rabbit and anti-mouse horseradish peroxidase linked immunoglobulin G (IgG) were purchased from Santa Cruz Biotechnology (Heidelberg, Germany).

    Techniques: Expressing, Binding Assay, Quantitative RT-PCR, Activated Clotting Time Assay

    AngII promotes a physical interaction of HuR with non-muscle myosin IIA, which structurally depends on phosphorylated S318 within the RRM3. ( A , upper panel). Schematic representation of c-myc-tagged wild-type HuR and different deletion mutants bearing truncations in the specific HuR domains. (A, lower panel). HMC were transfected with the indicated c-myc-HuR coding plasmids. Seventy-two hours after transfection, cells were treated without (−) or with AngII (+) for 2 h. Thereafter, c-myc-tagged HuR proteins were immunoprecipitated with an anti-c-myc antibody (IP: c-myc), or isotype specific control (IP: IgG). HuR-bound myosin IIA was detected by Western blot analysis with an anti-myosin IIA specific antibody. Equal pull-down (input levels) of c-myc was ascertained by reincubating the blot with the same antibody used for IP. ( B ). Blot-overlay assay demonstrating that the hinge region of HuR is not relevant for AngII-induced myosin IIA binding. Pull-down of myosin IIA from vehicle (−) or AngII treated HMC was performed as described in ‘Materials and Methods’ section . Blots were probed with equal amounts of total cell lysates cells overexpressing c-myc-HuR wild-type or c-myc-HuRΔHinge. Myosin IIA-HuR interaction was assessed by reincubating the blot with an anti-c-myc antibody. Equal pull-down of myosin is ascertained by reincubating the blot with the same antibody used for the myosin IIA co-IP. ( C ). HMC were transfected with the indicated c-myc-point mutated HuR and IPs were performed similar as described for (A). Equal pull-down (input levels) of c-myc was ascertained by reincubating the blot with the same antibody used for IP. ( D , left panel). HuR binding to myosin IIA is sensitive toward RNase treatment. HMC overexpressing c-myc-HuRwild-type or c-myc-HuRΔHinge were treated for 2 h with AngII (0.1 µM) before total cell lysates from transfectants were immunoprecipitated with an anti-c-myc antibody or with control IgG. Before the IP reaction, the cell homogenates were subjected to RNase treatment and subsequently immunoblotted with an anti-myosin IIA specific antibody. Equal pull-down of c-myc is ascertained by reincubating the blot with anti-c-myc antiserum. (right panel). To test for AngII-dependent association of myosinIIA with endogenous HuR, 200 µg of total cell extracts from unstimulated (−) or AngII-stimulated (+) HMC were subjected to (IP) as described in ‘Materials and Methods’ section. The data shown are from a single experiment representative of two repeats with similar results.

    Journal: Nucleic Acids Research

    Article Title: RNA-dependent association with myosin IIA promotes F-actin-guided trafficking of the ELAV-like protein HuR to polysomes

    doi: 10.1093/nar/gkt663

    Figure Lengend Snippet: AngII promotes a physical interaction of HuR with non-muscle myosin IIA, which structurally depends on phosphorylated S318 within the RRM3. ( A , upper panel). Schematic representation of c-myc-tagged wild-type HuR and different deletion mutants bearing truncations in the specific HuR domains. (A, lower panel). HMC were transfected with the indicated c-myc-HuR coding plasmids. Seventy-two hours after transfection, cells were treated without (−) or with AngII (+) for 2 h. Thereafter, c-myc-tagged HuR proteins were immunoprecipitated with an anti-c-myc antibody (IP: c-myc), or isotype specific control (IP: IgG). HuR-bound myosin IIA was detected by Western blot analysis with an anti-myosin IIA specific antibody. Equal pull-down (input levels) of c-myc was ascertained by reincubating the blot with the same antibody used for IP. ( B ). Blot-overlay assay demonstrating that the hinge region of HuR is not relevant for AngII-induced myosin IIA binding. Pull-down of myosin IIA from vehicle (−) or AngII treated HMC was performed as described in ‘Materials and Methods’ section . Blots were probed with equal amounts of total cell lysates cells overexpressing c-myc-HuR wild-type or c-myc-HuRΔHinge. Myosin IIA-HuR interaction was assessed by reincubating the blot with an anti-c-myc antibody. Equal pull-down of myosin is ascertained by reincubating the blot with the same antibody used for the myosin IIA co-IP. ( C ). HMC were transfected with the indicated c-myc-point mutated HuR and IPs were performed similar as described for (A). Equal pull-down (input levels) of c-myc was ascertained by reincubating the blot with the same antibody used for IP. ( D , left panel). HuR binding to myosin IIA is sensitive toward RNase treatment. HMC overexpressing c-myc-HuRwild-type or c-myc-HuRΔHinge were treated for 2 h with AngII (0.1 µM) before total cell lysates from transfectants were immunoprecipitated with an anti-c-myc antibody or with control IgG. Before the IP reaction, the cell homogenates were subjected to RNase treatment and subsequently immunoblotted with an anti-myosin IIA specific antibody. Equal pull-down of c-myc is ascertained by reincubating the blot with anti-c-myc antiserum. (right panel). To test for AngII-dependent association of myosinIIA with endogenous HuR, 200 µg of total cell extracts from unstimulated (−) or AngII-stimulated (+) HMC were subjected to (IP) as described in ‘Materials and Methods’ section. The data shown are from a single experiment representative of two repeats with similar results.

    Article Snippet: Antibodies raised against HuR, histone deacetylase-1 (HDAC1), c-myc, anti-rabbit and anti-mouse horseradish peroxidase linked immunoglobulin G (IgG) were purchased from Santa Cruz Biotechnology (Heidelberg, Germany).

    Techniques: Transfection, Immunoprecipitation, Western Blot, Overlay Assay, Binding Assay, Co-Immunoprecipitation Assay

    Phospho-ezrin and ezrin expression in NPC metastatic nodes. Phospho-ezrin and ezrin were detected in the primary and metastatic NPC samples using immunohistochemistry. ( a , b ) primary NPC and metastatic tumor sections were stained with hematoxylin and eosin; ( c , d ) stained with antibodies against ezrin; ( e , f ) stained with phospho-ezrin at Thr567; ( g , h ) stained with IgG, served as a blank control. Arrow, positive cells. Original magnification, ×400. Scale bar, 5 μm. H E, hematoxylin and eosin.

    Journal: International Journal of Molecular Sciences

    Article Title: Dinitrosopiperazine-Mediated Phosphorylated-Proteins Are Involved in Nasopharyngeal Carcinoma Metastasis

    doi: 10.3390/ijms151120054

    Figure Lengend Snippet: Phospho-ezrin and ezrin expression in NPC metastatic nodes. Phospho-ezrin and ezrin were detected in the primary and metastatic NPC samples using immunohistochemistry. ( a , b ) primary NPC and metastatic tumor sections were stained with hematoxylin and eosin; ( c , d ) stained with antibodies against ezrin; ( e , f ) stained with phospho-ezrin at Thr567; ( g , h ) stained with IgG, served as a blank control. Arrow, positive cells. Original magnification, ×400. Scale bar, 5 μm. H E, hematoxylin and eosin.

    Article Snippet: The secondary antibodies, horseradish peroxidase-linked anti-mouse immunoglobulin G and anti-rabbit immunoglobulin G were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA).

    Techniques: Expressing, Immunohistochemistry, Staining

    Validation of the representative phosphoproteins by Western-blotting. Representative proteins ezrin ( A ); vimentin ( B ); stathmin ( C ); STAT3 ( D ) of the immunoprecipitated proteins were detected by Western-blotting using anti-serine or threonine phosphorylation antibody. IgG served as the control. The relative Western-blotting ratio of phosphorylation levels of every protein was normalized to its corresponding proteomic data.

    Journal: International Journal of Molecular Sciences

    Article Title: Dinitrosopiperazine-Mediated Phosphorylated-Proteins Are Involved in Nasopharyngeal Carcinoma Metastasis

    doi: 10.3390/ijms151120054

    Figure Lengend Snippet: Validation of the representative phosphoproteins by Western-blotting. Representative proteins ezrin ( A ); vimentin ( B ); stathmin ( C ); STAT3 ( D ) of the immunoprecipitated proteins were detected by Western-blotting using anti-serine or threonine phosphorylation antibody. IgG served as the control. The relative Western-blotting ratio of phosphorylation levels of every protein was normalized to its corresponding proteomic data.

    Article Snippet: The secondary antibodies, horseradish peroxidase-linked anti-mouse immunoglobulin G and anti-rabbit immunoglobulin G were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA).

    Techniques: Western Blot, Immunoprecipitation

    Identification of a potent small molecule inhibitor of Plk1 PBD A. The chemical structure of T521: 5-(ethylsulfonyl)-2-(4-fluorophenyl)-4-(phenylsulfonyl) oxazole. B, C, D. The inhibitory effect of T521, Poloxin and TQ on the binding of the PBDs of Plk1, 2 and 3 to the FITC-labeled phosphopeptide was analyzed by FP assay. Briefly, T521, Poloxin or TQ was incubated with Plk1-3 PBD for 1 hr at room temperature (RT) prior to addition of FITC-labeled phosphopeptide. The inhibitory effect was calculated as described in the Materials and Methods. Error bars represent SD. E. ELISA-based PBD binding inhibition assay to determine the inhibitory effect of T521 on the interaction between Plk1 PBD and its binding target Map205 PBM . His-tagged PBD of Plk1 and different concentrations of T521 were added into GST-Map205 PBM -coated plates. After incubation, the plate was washed and then probed with mouse anti-His primary antibody, followed by goat anti-mouse IgG-HRP secondary antibody. After washing, the plate was incubated with TMB solution and the OD 450 was measured. Detailed procedure was described in Methods and Materials. DMSO was used as control. F. T521 inhibits Bub1-Plk1 interaction in vivo using Co-immunoprecipitation (IP) assay. HeLa cells synchronized by double-thymidine block were released into medium containing DMSO or T521 at indicated concentrations for 10 hrs, and then lysed and subject to Bub1 IP. The proteins in the precipitates were separated by 10% SDS-PAGE and probed with the anti-hPlk1 and anti-Bub1 antibodies. Moreover, the Bub1 and Plk1 levels were also determined in the lysates of HeLa cells before IP. β-actin served as the loading control.

    Journal: Oncotarget

    Article Title: Identification of a novel Polo-like kinase 1 inhibitor that specifically blocks the functions of Polo-Box domain

    doi: 10.18632/oncotarget.13603

    Figure Lengend Snippet: Identification of a potent small molecule inhibitor of Plk1 PBD A. The chemical structure of T521: 5-(ethylsulfonyl)-2-(4-fluorophenyl)-4-(phenylsulfonyl) oxazole. B, C, D. The inhibitory effect of T521, Poloxin and TQ on the binding of the PBDs of Plk1, 2 and 3 to the FITC-labeled phosphopeptide was analyzed by FP assay. Briefly, T521, Poloxin or TQ was incubated with Plk1-3 PBD for 1 hr at room temperature (RT) prior to addition of FITC-labeled phosphopeptide. The inhibitory effect was calculated as described in the Materials and Methods. Error bars represent SD. E. ELISA-based PBD binding inhibition assay to determine the inhibitory effect of T521 on the interaction between Plk1 PBD and its binding target Map205 PBM . His-tagged PBD of Plk1 and different concentrations of T521 were added into GST-Map205 PBM -coated plates. After incubation, the plate was washed and then probed with mouse anti-His primary antibody, followed by goat anti-mouse IgG-HRP secondary antibody. After washing, the plate was incubated with TMB solution and the OD 450 was measured. Detailed procedure was described in Methods and Materials. DMSO was used as control. F. T521 inhibits Bub1-Plk1 interaction in vivo using Co-immunoprecipitation (IP) assay. HeLa cells synchronized by double-thymidine block were released into medium containing DMSO or T521 at indicated concentrations for 10 hrs, and then lysed and subject to Bub1 IP. The proteins in the precipitates were separated by 10% SDS-PAGE and probed with the anti-hPlk1 and anti-Bub1 antibodies. Moreover, the Bub1 and Plk1 levels were also determined in the lysates of HeLa cells before IP. β-actin served as the loading control.

    Article Snippet: The plate was further probed for 45 min at 37°C with goat anti-mouse IgG-HRP secondary antibody (Cat# sc-2005; Santa Cruz; 1:2000; 100 μL/well).

    Techniques: Binding Assay, Labeling, FP Assay, Incubation, Enzyme-linked Immunosorbent Assay, Inhibition, In Vivo, Immunoprecipitation, Blocking Assay, SDS Page

    Immunoreactivity of various preparations by indirect ELISA Filled circles: 2C5 antibody, Circles: 2C5-modified doxorubicin-loaded micelles, Filled squares: Isotype-matched IgG, Squares: IgG-modified doxorubicin-loaded micelles . The binding of doxorubicin-loaded PEG-PE micelles harboring 2C5 antibody or isotype antibody to a monolayer of the antigen (nucleohistones) was evaluated by ELISA and detected using an HRP conjugated antibody. Data represent the mean ± SD, n=3.

    Journal: Journal of controlled release : official journal of the Controlled Release Society

    Article Title: Accumulation and toxicity of antibody-targeted doxorubicin-loaded PEG-PE micelles in ovarian cancer cell spheroid model

    doi: 10.1016/j.jconrel.2012.09.003

    Figure Lengend Snippet: Immunoreactivity of various preparations by indirect ELISA Filled circles: 2C5 antibody, Circles: 2C5-modified doxorubicin-loaded micelles, Filled squares: Isotype-matched IgG, Squares: IgG-modified doxorubicin-loaded micelles . The binding of doxorubicin-loaded PEG-PE micelles harboring 2C5 antibody or isotype antibody to a monolayer of the antigen (nucleohistones) was evaluated by ELISA and detected using an HRP conjugated antibody. Data represent the mean ± SD, n=3.

    Article Snippet: Mouse monoclonal anti Bcl-2 antibody and rabbit anti-mouse IgG-HRP were from Santa Cruz Biotechnology (Santa Cruz, CA).

    Techniques: Indirect ELISA, Modification, Binding Assay, Enzyme-linked Immunosorbent Assay

    Functional significance of trimeric contacts in CaiT. (a) Western blot analysis of CaiT with given amino acid replacements in E. coli JW0039 membranes. HRP-linked mouse anti-His IgG was used to detect CaiT. PageRuler Prestained Protein Ladder was used for molecular size estimation. Shown is the section of the blot that contains the CaiT band. The complete gel is presented in Fig. S6. (b) Initial rates of counterflow (black) and maximum accumulation of carnitine in E. coli JW0039 (white) were determined as described in the legend of Fig. 3 . As negative control, cells carrying pET21a ( nc ) were used. (c) Initial rates of counterflow (black) and maximum accumulation (white) of carnitine catalyzed by CaiT variants in proteoliposomes. CaiT was purified and reconstituted as described 1 . The lipid to protein ratio of the resulting proteoliposomes was 100:1 (w/w). Proteoliposomes were preloaded with 10 mM unlabeled L-carnitine overnight. Aliquots of the proteoliposome suspension (1–2 µg of protein) were then diluted into 400 µL buffer containing 4.5 µM L-[methyl- 14 C]carnitine (55 Ci/mol) resulting in a final carnitine concentration of 54.5 µM. As negative control, proteoliposomes not preloaded with unlabeled carnitine ( nc ) were used and the obtained transport rates were used for correcting the data. Carnitine counterflow with cells or proteoliposomes was assayed at 25 °C under aerobic conditions using a rapid filtration method as described 1 . (d) Michaelis-Menten kinetics of L-carnitine counterflow in proteoliposomes reconstituted with wild-type CaiT (WT) or CaiT-D288A. Initial rates of L-[methyl- 14 C]carnitine uptake were measured at given substrate concentrations and plotted using the kinetic module of GraphPad Prism. For all experiments, standard deviations were calculated from triplicate determinations of a representative experiment.

    Journal: Scientific Reports

    Article Title: Comparison of the functional properties of trimeric and monomeric CaiT of Escherichia coli

    doi: 10.1038/s41598-019-40516-7

    Figure Lengend Snippet: Functional significance of trimeric contacts in CaiT. (a) Western blot analysis of CaiT with given amino acid replacements in E. coli JW0039 membranes. HRP-linked mouse anti-His IgG was used to detect CaiT. PageRuler Prestained Protein Ladder was used for molecular size estimation. Shown is the section of the blot that contains the CaiT band. The complete gel is presented in Fig. S6. (b) Initial rates of counterflow (black) and maximum accumulation of carnitine in E. coli JW0039 (white) were determined as described in the legend of Fig. 3 . As negative control, cells carrying pET21a ( nc ) were used. (c) Initial rates of counterflow (black) and maximum accumulation (white) of carnitine catalyzed by CaiT variants in proteoliposomes. CaiT was purified and reconstituted as described 1 . The lipid to protein ratio of the resulting proteoliposomes was 100:1 (w/w). Proteoliposomes were preloaded with 10 mM unlabeled L-carnitine overnight. Aliquots of the proteoliposome suspension (1–2 µg of protein) were then diluted into 400 µL buffer containing 4.5 µM L-[methyl- 14 C]carnitine (55 Ci/mol) resulting in a final carnitine concentration of 54.5 µM. As negative control, proteoliposomes not preloaded with unlabeled carnitine ( nc ) were used and the obtained transport rates were used for correcting the data. Carnitine counterflow with cells or proteoliposomes was assayed at 25 °C under aerobic conditions using a rapid filtration method as described 1 . (d) Michaelis-Menten kinetics of L-carnitine counterflow in proteoliposomes reconstituted with wild-type CaiT (WT) or CaiT-D288A. Initial rates of L-[methyl- 14 C]carnitine uptake were measured at given substrate concentrations and plotted using the kinetic module of GraphPad Prism. For all experiments, standard deviations were calculated from triplicate determinations of a representative experiment.

    Article Snippet: Relative amounts of CaiT with given amino acid replacements in membranes of E. coli JW0039 harboring plasmid pT7-5/caiT were estimated by Western blot analysis with HRP-linked mouse monoclonal IgG His-probe Antibody (H-3) (Santa Cruz Biotechnology) directed against the His tag at the C terminus of each CaiT variant by the enhances chemiluminescence method according to the manufacturer’s protocol.

    Techniques: Functional Assay, Western Blot, Negative Control, Purification, Concentration Assay, Filtration

    Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific IgG antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using HRP-conjugated anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P

    Journal: International Journal of Nanomedicine

    Article Title: A niosome formulation modulates the Th1/Th2 bias immune response in mice and also provides protection against anthrax spore challenge

    doi: 10.2147/IJN.S153150

    Figure Lengend Snippet: Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific IgG antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using HRP-conjugated anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P

    Article Snippet: HRP-conjugated anti-mouse IgG (Santa Cruz Biotechnology Inc.) was added at a dilution of 1:10,000 and incubated at 37°C for 1 hour.

    Techniques: Mouse Assay, Injection, Enzyme-linked Immunosorbent Assay, Software

    Anti-D4 titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, D4, and NISV + D4. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for D4-specific IgG antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against D4. IgG and its isotypes were determined in the serum of immunized mice by ELISA using HRP-conjugated anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. D4 immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. **** P

    Journal: International Journal of Nanomedicine

    Article Title: A niosome formulation modulates the Th1/Th2 bias immune response in mice and also provides protection against anthrax spore challenge

    doi: 10.2147/IJN.S153150

    Figure Lengend Snippet: Anti-D4 titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, D4, and NISV + D4. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for D4-specific IgG antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against D4. IgG and its isotypes were determined in the serum of immunized mice by ELISA using HRP-conjugated anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. D4 immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. **** P

    Article Snippet: HRP-conjugated anti-mouse IgG (Santa Cruz Biotechnology Inc.) was added at a dilution of 1:10,000 and incubated at 37°C for 1 hour.

    Techniques: Mouse Assay, Injection, Enzyme-linked Immunosorbent Assay, Software

    Specific humoral responses elicited by immunization of mice with microneme proteins. The reactivity of immunoglobulins anti-STAg was determined by ELISA in serum samples collected from both immunized (TgMICs) and control (PBS) mice 15 days after the last antigen injection. Each point/bar represents the average absorbance ± SD of the serum samples from 4 animals. (A) Absorbance provided by the reaction of serum IgG with STAg. (B) Absorbance provided by the reaction of serum IgG1 and IgG2a (diluted 1:25) with STAg. The average absorbance ± SD generated by the reaction of serum IgG1 or IgG2a from each group of immunized mice was significantly higher than the corresponding values provided by control mice, with the exception of the TgMIC6-immunized group, whose results were not significantly different of those of the control group. Three independent experiments were performed, and data from one representative experiment is shown. Asterisks represent statistical significant differences (*p

    Journal: PLoS ONE

    Article Title: Vaccination with Recombinant Microneme Proteins Confers Protection against Experimental Toxoplasmosis in Mice

    doi: 10.1371/journal.pone.0143087

    Figure Lengend Snippet: Specific humoral responses elicited by immunization of mice with microneme proteins. The reactivity of immunoglobulins anti-STAg was determined by ELISA in serum samples collected from both immunized (TgMICs) and control (PBS) mice 15 days after the last antigen injection. Each point/bar represents the average absorbance ± SD of the serum samples from 4 animals. (A) Absorbance provided by the reaction of serum IgG with STAg. (B) Absorbance provided by the reaction of serum IgG1 and IgG2a (diluted 1:25) with STAg. The average absorbance ± SD generated by the reaction of serum IgG1 or IgG2a from each group of immunized mice was significantly higher than the corresponding values provided by control mice, with the exception of the TgMIC6-immunized group, whose results were not significantly different of those of the control group. Three independent experiments were performed, and data from one representative experiment is shown. Asterisks represent statistical significant differences (*p

    Article Snippet: Plates were then incubated at 37°C for 1 h, washed four times, and incubated with horseradish-peroxidase-conjugated goat anti-mouse IgG, IgG1, or IgG2b (Santa Cruz Biotechnology), at 1:5000 dilution in blocking buffer, for 1 h, at 37°C.

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay, Injection, Generated

    SDS-PAGE and western blot analysis of native and recombinant microneme proteins. SDS-PAGE of recombinant proteins (panels A, B, and C, Coomassie Blue stained) or native (panel D, silver-stained) proteins. Heterologous expression was noted in E . coli (DE3) and recombinant proteins were detected in inclusion bodies. Lane 1: protein expression before induction. Lane 2: protein expression after induction. Lane 3: purified and refolded histidine-tagged recombinant proteins, displayed apparent molecular masses of 70-kDa (TgMIC1, panel A), 80-kDa (TgMIC4, panel B), and 30-kDa (TgMIC6, panel C). Panel E shows the electrophoretical profile of the Lac+ fraction, composed of the native proteins TgMIC1 (53-kDa) and TgMIC4 (68-kDa). Lane M: Molecular mass markers. Reactivity of recombinant and native microneme proteins with anti-Lac+ mouse serum was examined by western blot (Panel E), developed with peroxidase-conjugated goat anti-mouse IgG.

    Journal: PLoS ONE

    Article Title: Vaccination with Recombinant Microneme Proteins Confers Protection against Experimental Toxoplasmosis in Mice

    doi: 10.1371/journal.pone.0143087

    Figure Lengend Snippet: SDS-PAGE and western blot analysis of native and recombinant microneme proteins. SDS-PAGE of recombinant proteins (panels A, B, and C, Coomassie Blue stained) or native (panel D, silver-stained) proteins. Heterologous expression was noted in E . coli (DE3) and recombinant proteins were detected in inclusion bodies. Lane 1: protein expression before induction. Lane 2: protein expression after induction. Lane 3: purified and refolded histidine-tagged recombinant proteins, displayed apparent molecular masses of 70-kDa (TgMIC1, panel A), 80-kDa (TgMIC4, panel B), and 30-kDa (TgMIC6, panel C). Panel E shows the electrophoretical profile of the Lac+ fraction, composed of the native proteins TgMIC1 (53-kDa) and TgMIC4 (68-kDa). Lane M: Molecular mass markers. Reactivity of recombinant and native microneme proteins with anti-Lac+ mouse serum was examined by western blot (Panel E), developed with peroxidase-conjugated goat anti-mouse IgG.

    Article Snippet: Plates were then incubated at 37°C for 1 h, washed four times, and incubated with horseradish-peroxidase-conjugated goat anti-mouse IgG, IgG1, or IgG2b (Santa Cruz Biotechnology), at 1:5000 dilution in blocking buffer, for 1 h, at 37°C.

    Techniques: SDS Page, Western Blot, Recombinant, Staining, Expressing, Purification

    Experimental Protocol. (A) In the first experimental procedure mice were subcutaneously (s.c.) vaccinated with microneme proteins emulsified in Freund’s complete adjuvant. Mice were boosted at the same dose and regimen on day 15 and 30 after first injection, now emulsified in Freund’s incomplete adjuvant. Fifteen days after the last injection, blood and spleen samples were collected to assess serum IgG, in vitro T cell proliferation, and cytokine concentrations. (B) One month after the last immunization procedure, the mice were orally infected with 80 cysts of the ME49 strain and the mortality was monitored daily for 1 month. To evaluate the tissue cyst burden, the brain of the mice infected with 40 cysts was removed 1 month after the challenge and the mean number of cysts per brain was determined. Additionally, blood samples from mice challenged with 40 cysts were collected 30 days after T . gondii challenge and the serum cytokine levels were analyzed.

    Journal: PLoS ONE

    Article Title: Vaccination with Recombinant Microneme Proteins Confers Protection against Experimental Toxoplasmosis in Mice

    doi: 10.1371/journal.pone.0143087

    Figure Lengend Snippet: Experimental Protocol. (A) In the first experimental procedure mice were subcutaneously (s.c.) vaccinated with microneme proteins emulsified in Freund’s complete adjuvant. Mice were boosted at the same dose and regimen on day 15 and 30 after first injection, now emulsified in Freund’s incomplete adjuvant. Fifteen days after the last injection, blood and spleen samples were collected to assess serum IgG, in vitro T cell proliferation, and cytokine concentrations. (B) One month after the last immunization procedure, the mice were orally infected with 80 cysts of the ME49 strain and the mortality was monitored daily for 1 month. To evaluate the tissue cyst burden, the brain of the mice infected with 40 cysts was removed 1 month after the challenge and the mean number of cysts per brain was determined. Additionally, blood samples from mice challenged with 40 cysts were collected 30 days after T . gondii challenge and the serum cytokine levels were analyzed.

    Article Snippet: Plates were then incubated at 37°C for 1 h, washed four times, and incubated with horseradish-peroxidase-conjugated goat anti-mouse IgG, IgG1, or IgG2b (Santa Cruz Biotechnology), at 1:5000 dilution in blocking buffer, for 1 h, at 37°C.

    Techniques: Mouse Assay, Injection, In Vitro, Infection

    Specific binding ELISA of hE16 to plant-derived DIII. Serial dilutions of hE16 purified from mammalian or plant cells were incubated in sample wells coated with plant-produced WNV DIII and detected with an HRP-conjugated anti-human gamma antibody. A commercial generic human IgG was used as a negative control. Mean ± SD of samples from three independent experiments is presented.

    Journal: BioMed Research International

    Article Title: A Plant-Produced Antigen Elicits Potent Immune Responses against West Nile Virus in Mice

    doi: 10.1155/2014/952865

    Figure Lengend Snippet: Specific binding ELISA of hE16 to plant-derived DIII. Serial dilutions of hE16 purified from mammalian or plant cells were incubated in sample wells coated with plant-produced WNV DIII and detected with an HRP-conjugated anti-human gamma antibody. A commercial generic human IgG was used as a negative control. Mean ± SD of samples from three independent experiments is presented.

    Article Snippet: After washing with PBST, the plates were incubated with an HRP-conjugated goat anti-mouse IgG1 (Santa Cruz Biotech) or anti-mouse IgG2a (Southern Biotech).

    Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay, Derivative Assay, Purification, Incubation, Produced, Negative Control

    In vivo immunogenicity of K. pneumoniae EVs. ( a ) K. pneumoniae EV-reactive IgG antibody in serum from mice immunized with 10, 100, 100 ng of K. pneumoniae EVs. This antibody was measured by ELISA ( n =5; * P

    Journal: Experimental & Molecular Medicine

    Article Title: Vaccination with Klebsiella pneumoniae-derived extracellular vesicles protects against bacteria-induced lethality via both humoral and cellular immunity

    doi: 10.1038/emm.2015.59

    Figure Lengend Snippet: In vivo immunogenicity of K. pneumoniae EVs. ( a ) K. pneumoniae EV-reactive IgG antibody in serum from mice immunized with 10, 100, 100 ng of K. pneumoniae EVs. This antibody was measured by ELISA ( n =5; * P

    Article Snippet: The serum was diluted with the reagent diluent (1:500) and was then incubated in the EV-coated plate for 2 h. After incubation, the plates were washed and treated with 200 ng ml−1 of horseradish peroxidase-conjugated goat anti-mouse immunoglobulin G (IgG) antibody (Santa Cruz Biotechnology, CA, USA) for 2 h. After washing the plates, the enhanced chemiluminescence substrate (Thermo Scientific, Kenilworth, NJ, USA) was added and analyzed using a Victor Wallac 1420 apparatus (PerkinElmer, Waltham, MA, USA).

    Techniques: In Vivo, Mouse Assay, Enzyme-linked Immunosorbent Assay

    Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific IgG antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using HRP-conjugated anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P

    Journal: International Journal of Nanomedicine

    Article Title: A niosome formulation modulates the Th1/Th2 bias immune response in mice and also provides protection against anthrax spore challenge

    doi: 10.2147/IJN.S153150

    Figure Lengend Snippet: Anti-PA titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, PA, and NISV + PA. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for PA-specific IgG antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against PA. IgG and its isotypes were determined in the serum of immunized mice by ELISA using HRP-conjugated anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. PA immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. ** P

    Article Snippet: Horseradish peroxidase (HRP)-conjugated anti-mouse IgG, IgG1, and IgG2a were purchased from Santa Cruz Biotechnology Inc. (Dallas, TX, USA).

    Techniques: Mouse Assay, Injection, Enzyme-linked Immunosorbent Assay, Software

    Anti-D4 titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, D4, and NISV + D4. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for D4-specific IgG antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against D4. IgG and its isotypes were determined in the serum of immunized mice by ELISA using HRP-conjugated anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. D4 immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. **** P

    Journal: International Journal of Nanomedicine

    Article Title: A niosome formulation modulates the Th1/Th2 bias immune response in mice and also provides protection against anthrax spore challenge

    doi: 10.2147/IJN.S153150

    Figure Lengend Snippet: Anti-D4 titers elicited by Swiss albino mice immunized intraperitoneally with PBS, blank NISV, D4, and NISV + D4. Notes: Subsequent booster doses were injected on the 14th and 28th day. Sera were collected from individual mice and analyzed in triplicates for D4-specific IgG antibodies and its isotypes by ELISA: ( A ) shows the IgG level elicited by the different groups at different time intervals; ( B ) shows the level of IgG1-specific antibodies, while ( C ) shows the levels of IgG2a-specific antibodies against D4. IgG and its isotypes were determined in the serum of immunized mice by ELISA using HRP-conjugated anti-mice IgG, IgG1, and IgG2a antibodies (1:10,000 dilution). For data preparation and statistical analysis, GraphPad Prism Version 6.05 software was used. The results were expressed as mean value with SD from individual mice group. D4 immunized without particle group was compared with particulated groups for calculating P -value using the two-way ANOVA followed by Tukey’s multiple comparisons test. **** P

    Article Snippet: Horseradish peroxidase (HRP)-conjugated anti-mouse IgG, IgG1, and IgG2a were purchased from Santa Cruz Biotechnology Inc. (Dallas, TX, USA).

    Techniques: Mouse Assay, Injection, Enzyme-linked Immunosorbent Assay, Software

    Expression of CD23 in mouse AECs Cell lysates (50 µg) derived from primary nasal ( A ), tracheal ( B ), and lung ( B ) epithelial cells were electrophoresed on a 12 % SDS-PAGE gel under reducing condition. The separated proteins were transferred onto a nitrocellulose membrane, blotted with rat anti-mouse CD23 mAb (B3B4) followed by HRP-conjugated rabbit anti-rat IgG Ab. Proteins were visualized using the ECL method. Lysates (50 µg) from mouse spleen or CHO cells were used as a positive or negative control, respectively. β-tubulin was used as an internal control. Arrows indicate mouse CD23 and β-tubulin. ( C ) Immunohistochemical staining of mouse lung. Normal mouse lung was prepared in OCT medium and cryosectioned at 5 µm. Frozen tissue sections were fixed and permeabilized with ice-cold acetone and blocked with 10% normal goat serum. A spleen section was used as a positive control. Sections were incubated with rabbit anti-CD23 Ab or normal rabbit IgG, followed by staining with Alexa Fluor 555-conjugated goat anti-rabbit Ab. Nuclei were stained with DAPI. Images were captured using a Zeiss LSM510 confocal microscope. Samples were visualized under consisten contrast and brightness settings.

    Journal: Mucosal immunology

    Article Title: Inhibition of CD23-mediated IgE transcytosis suppresses the initiation and development of allergic airway inflammation

    doi: 10.1038/mi.2015.16

    Figure Lengend Snippet: Expression of CD23 in mouse AECs Cell lysates (50 µg) derived from primary nasal ( A ), tracheal ( B ), and lung ( B ) epithelial cells were electrophoresed on a 12 % SDS-PAGE gel under reducing condition. The separated proteins were transferred onto a nitrocellulose membrane, blotted with rat anti-mouse CD23 mAb (B3B4) followed by HRP-conjugated rabbit anti-rat IgG Ab. Proteins were visualized using the ECL method. Lysates (50 µg) from mouse spleen or CHO cells were used as a positive or negative control, respectively. β-tubulin was used as an internal control. Arrows indicate mouse CD23 and β-tubulin. ( C ) Immunohistochemical staining of mouse lung. Normal mouse lung was prepared in OCT medium and cryosectioned at 5 µm. Frozen tissue sections were fixed and permeabilized with ice-cold acetone and blocked with 10% normal goat serum. A spleen section was used as a positive control. Sections were incubated with rabbit anti-CD23 Ab or normal rabbit IgG, followed by staining with Alexa Fluor 555-conjugated goat anti-rabbit Ab. Nuclei were stained with DAPI. Images were captured using a Zeiss LSM510 confocal microscope. Samples were visualized under consisten contrast and brightness settings.

    Article Snippet: HRP-conjugated bovine anti-goat IgG Ab was from Santa Cruz Biotechnology (Santa Cruz, CA).

    Techniques: Expressing, Derivative Assay, SDS Page, Negative Control, Immunohistochemistry, Staining, Positive Control, Incubation, Microscopy

    Epithelial CD23 facilitates airway inflammation after OVA sensitization and challenge in chimeric mice A. Mice were sensitized (i.p.) with OVA or left untreated. Lung and trachea epithelial cells were isolated and cell lysates (50 µg) were subjected to 12 % SDS-PAGE gel electrophoresis under reducing conditions. Proteins were transferred onto nitrocellulose membranes and blotted with rat-anti mouse CD23 mAb (B3B4) followed by HRP-conjugated rabbit anti-rat IgG Ab. Proteins were visualized using ECL. Arrows indicate mouse CD23 and β-tubulin. B–D. WT/WT, CD23KO/WT, WT/CD23KO chimeric mice were i.p. sensitized with 100 µg OVA plus 4 mg alum at day 0 and subsequently injected (i.p.) with 100 µg OVA on day 7 and 14. Mice were challenged on day 21 with nebulized 1 % OVA for 30 min. 5 h after OVA challenge, sera were collected and OVA quantitated by ELISA (B). 24 h after OVA challenge, BAL fluid and sera were collected and OVA-specific IgE (C) and IL-4 (D) concentrations assessed by ELISA. *P

    Journal: Mucosal immunology

    Article Title: Inhibition of CD23-mediated IgE transcytosis suppresses the initiation and development of allergic airway inflammation

    doi: 10.1038/mi.2015.16

    Figure Lengend Snippet: Epithelial CD23 facilitates airway inflammation after OVA sensitization and challenge in chimeric mice A. Mice were sensitized (i.p.) with OVA or left untreated. Lung and trachea epithelial cells were isolated and cell lysates (50 µg) were subjected to 12 % SDS-PAGE gel electrophoresis under reducing conditions. Proteins were transferred onto nitrocellulose membranes and blotted with rat-anti mouse CD23 mAb (B3B4) followed by HRP-conjugated rabbit anti-rat IgG Ab. Proteins were visualized using ECL. Arrows indicate mouse CD23 and β-tubulin. B–D. WT/WT, CD23KO/WT, WT/CD23KO chimeric mice were i.p. sensitized with 100 µg OVA plus 4 mg alum at day 0 and subsequently injected (i.p.) with 100 µg OVA on day 7 and 14. Mice were challenged on day 21 with nebulized 1 % OVA for 30 min. 5 h after OVA challenge, sera were collected and OVA quantitated by ELISA (B). 24 h after OVA challenge, BAL fluid and sera were collected and OVA-specific IgE (C) and IL-4 (D) concentrations assessed by ELISA. *P

    Article Snippet: HRP-conjugated bovine anti-goat IgG Ab was from Santa Cruz Biotechnology (Santa Cruz, CA).

    Techniques: Mouse Assay, Isolation, SDS Page, Nucleic Acid Electrophoresis, Injection, Enzyme-linked Immunosorbent Assay

    Production of autoantibodies to erythrocytes in SOD1-deficient mice ( A ) Detection of IgG bound to erythrocytes from SOD1 −/− mice. Collected erythrocytes were reacted with FITC-labelled anti-mouse IgG followed by FACS analysis. Relative values of bound IgG are shown. n =7 for SOD1 +/+ and n =5 for SOD1 −/− . ( B ) Cytosolic and membrane fractions of RBCs were incubated with blood plasma from SOD1 +/+ and SOD1 −/− mice (1:50 dilution) followed by reaction with an HRP-conjugated anti-mouse IgG. ( C ) Immunoblot analysis using blood plasma from an SOD1 −/− mouse and serum of a rabbit that had been immunized with bovine CAII. Western blot analysis of human CAII and RBCs with blood plasma from a SOD1 −/− mouse at 40 weeks of age (1:50 dilution; upper panel). While mass marker b contains CAII, mass marker a does not. Characteristics of an anti-bovine CAII antibody (1:1000 dilution) raised in rabbit by immunoblotting of the same samples (lower panel); representative of several experiments. ( D ) Immunoprecipitation of bovine CAII and lysate of mouse RBCs with an anti-bovine CAII polyclonal antibody. Immunoblotting was performed with blood plasma from SOD1 −/− mice and an anti-CAII monoclonal antibody (mAb). Representative data from several experiments. ( E ) The titres of the anti-CAII antibody in blood plasma were elevated, as judged by ELISA. The inset shows an enlargement of the results obtained for SOD1 +/+ and SOD1 −/− mice at 14 weeks of age.

    Journal: Biochemical Journal

    Article Title: Elevated oxidative stress in erythrocytes due to a SOD1 deficiency causes anaemia and triggers autoantibody production

    doi: 10.1042/BJ20061386

    Figure Lengend Snippet: Production of autoantibodies to erythrocytes in SOD1-deficient mice ( A ) Detection of IgG bound to erythrocytes from SOD1 −/− mice. Collected erythrocytes were reacted with FITC-labelled anti-mouse IgG followed by FACS analysis. Relative values of bound IgG are shown. n =7 for SOD1 +/+ and n =5 for SOD1 −/− . ( B ) Cytosolic and membrane fractions of RBCs were incubated with blood plasma from SOD1 +/+ and SOD1 −/− mice (1:50 dilution) followed by reaction with an HRP-conjugated anti-mouse IgG. ( C ) Immunoblot analysis using blood plasma from an SOD1 −/− mouse and serum of a rabbit that had been immunized with bovine CAII. Western blot analysis of human CAII and RBCs with blood plasma from a SOD1 −/− mouse at 40 weeks of age (1:50 dilution; upper panel). While mass marker b contains CAII, mass marker a does not. Characteristics of an anti-bovine CAII antibody (1:1000 dilution) raised in rabbit by immunoblotting of the same samples (lower panel); representative of several experiments. ( D ) Immunoprecipitation of bovine CAII and lysate of mouse RBCs with an anti-bovine CAII polyclonal antibody. Immunoblotting was performed with blood plasma from SOD1 −/− mice and an anti-CAII monoclonal antibody (mAb). Representative data from several experiments. ( E ) The titres of the anti-CAII antibody in blood plasma were elevated, as judged by ELISA. The inset shows an enlargement of the results obtained for SOD1 +/+ and SOD1 −/− mice at 14 weeks of age.

    Article Snippet: After washing twice in TBST for 30 min, the blots were incubated with HRP (horseradish peroxidase)-conjugated goat anti-rabbit IgG antibody (Santa Cruz).

    Techniques: Mouse Assay, FACS, Incubation, Western Blot, Marker, Immunoprecipitation, Enzyme-linked Immunosorbent Assay

    effect of drug treatment on Hsc70 and protein disulphide isomerase (PDI) expression in small intestinal villi isolated from ECwt-infected mice. A: flow cytometric analysis was performed on intestinal epithelial cells (IEC) isolated from villi from ECwt-infected mice (n = 2 mice for each experimental group) that had been treated with ibuprofen (IBF) (20 mg/kg/day), diclofenac (DCF) (1 mg/kg/day), pioglitazone (PGZ)(30 mg/kg/day), rosiglitazone (RGZ) (4 mg/kg/day), N-acetylcysteine (NAC) (18 mg/kg/day), ascorbic acid (AA) (20 mg/kg/day) or difenoxilate sodium (7.5 mg/kg/day) during three days post-inoculation (d.p.i.). Goat primary antibodies (Abs) against Hsc70 or PDI and fluorescein isothiocyanate (FITC)-conjugated mouse anti-goat IgG secondary Abs were used. Immunofluorescence analysis was performed on a Cyan (Dako) flow cytometer using a FlowJo software; B: radio immunoprecipitation assay lysates from villi isolated from ECwt-infected mice (n = 3 mice for each experimental group) that had been treated with NAC (18 mg/kg/day) during three days after 24 h post-inoculation. were added to ELISA plates coated with rabbit polyclonal Abs against Hsc70 or PDI. Goat Abs against Hsc70 or PDI were used as detection Abs and the reaction was revealed using horseradish peroxidase (HRP)-conjugated donkey anti-goat Abs and phenylenediamine dihydrochloride substrate before reading at optical density (OD) 492 nm . Lysates from uninfected villi were used as a control. Data are expressed as mean OD 492 nm Hsc70 or PDI antigen. Graph shows significant increase of Hsc70 and PDI expression (p

    Journal: Memórias do Instituto Oswaldo Cruz

    Article Title: Inhibition of rotavirus ECwt infection in ICR suckling mice by N-acetylcysteine, peroxisome proliferator-activated receptor gamma agonists and cyclooxygenase-2 inhibitors

    doi: 10.1590/0074-0276108062013011

    Figure Lengend Snippet: effect of drug treatment on Hsc70 and protein disulphide isomerase (PDI) expression in small intestinal villi isolated from ECwt-infected mice. A: flow cytometric analysis was performed on intestinal epithelial cells (IEC) isolated from villi from ECwt-infected mice (n = 2 mice for each experimental group) that had been treated with ibuprofen (IBF) (20 mg/kg/day), diclofenac (DCF) (1 mg/kg/day), pioglitazone (PGZ)(30 mg/kg/day), rosiglitazone (RGZ) (4 mg/kg/day), N-acetylcysteine (NAC) (18 mg/kg/day), ascorbic acid (AA) (20 mg/kg/day) or difenoxilate sodium (7.5 mg/kg/day) during three days post-inoculation (d.p.i.). Goat primary antibodies (Abs) against Hsc70 or PDI and fluorescein isothiocyanate (FITC)-conjugated mouse anti-goat IgG secondary Abs were used. Immunofluorescence analysis was performed on a Cyan (Dako) flow cytometer using a FlowJo software; B: radio immunoprecipitation assay lysates from villi isolated from ECwt-infected mice (n = 3 mice for each experimental group) that had been treated with NAC (18 mg/kg/day) during three days after 24 h post-inoculation. were added to ELISA plates coated with rabbit polyclonal Abs against Hsc70 or PDI. Goat Abs against Hsc70 or PDI were used as detection Abs and the reaction was revealed using horseradish peroxidase (HRP)-conjugated donkey anti-goat Abs and phenylenediamine dihydrochloride substrate before reading at optical density (OD) 492 nm . Lysates from uninfected villi were used as a control. Data are expressed as mean OD 492 nm Hsc70 or PDI antigen. Graph shows significant increase of Hsc70 and PDI expression (p

    Article Snippet: HRP-conjugated donkey anti-goat IgG Ab (0.4 µg/mL; SC-2020, Santa Cruz Biotechnology Inc) was used as a secondary Ab and the reaction was developed with intensified luminescence (Pierce) or AEC.

    Techniques: Expressing, Isolation, Infection, Mouse Assay, Flow Cytometry, Immunofluorescence, Cytometry, Software, Radio Immunoprecipitation, Enzyme-linked Immunosorbent Assay