alexa fluor 488 conjugated anti rabbit  (Jackson Immuno)

 
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    Name:
    Alexa Fluor 488 AffiniPure F ab ₂ Fragment Donkey Anti Mouse IgG
    Description:
    F ab 2 fragment antibodies are generated by pepsin digestion of whole IgG antibodies to remove most of the Fc region while leaving some of the hinge region F ab 2 fragments have two antigen binding Fab portions linked together by disulfide bonds and therefore they are divalent The average molecular weight is about 110 kDa They are used for specific applications such as to avoid binding of secondary antibodies to live cells with Fc receptors or to Protein A or Protein G Based on immunoelectrophoresis and or ELISA the antibody reacts with whole molecule mouse IgG It also reacts with the light chains of other mouse immunoglobulins No antibody was detected against non immunoglobulin serum proteins The antibody has been tested by ELISA and or solid phase adsorbed to ensure minimal cross reaction with bovine chicken goat guinea pig syrian hamster horse human rabbit and sheep serum proteins but it may cross react with immunoglobulins from other species
    Catalog Number:
    715-546-150
    Price:
    180.0
    Category:
    F ab ₂ Fragment Affinity Purified Antibodies
    Conjugate:
    Alexa Fluor 488
    Size:
    0 3 mg
    Format:
    F(ab')₂ Fragment
    Host:
    Donkey
    Buy from Supplier


    Structured Review

    Jackson Immuno alexa fluor 488 conjugated anti rabbit
    Induction of LDLR expression and activity by triciribine. ( a ) Sterol-fed or sterol-starved HepG2 cells were treated with vehicle or the indicated doses of triciribine for 14 hours and then harvested for examination by Western blotting. Left panel: One representative blot is shown (n = 5). Unprocessed blots are shown in Supplementary Fig. S7 . The graph in the right panel shows quantification of the Western blots, relative to similarly cultured, vehicle-treated cells. ( b ) HepG2 cells were cultured as in a and then treated with or without 1 μM triciribine, harvested at the indicated time points and then analyzed by immunoblotting. Left panel: One representative blot is shown (n = 5). Unprocessed blots are shown in Supplementary Fig. S8 . The graph in the right panel shows quantification of the Western blots, relative to similarly cultured cells harvested at 0 hour. ( c ) Sterol-fed HepG2 cells were treated with 1 µM for 14 hours and then examined by fluorescence microscopy as described in Materials and Methods. Green: LDLR, <t>Alexa</t> fluor 488, blue: nuclei, DAPI. One representative experiment is shown (n = 3). ( d ) Sterol-fed HepG2 cells were exposed to vehicle or 1 μM triciribine for 14 hours before analysis for mean fluorescence intensity (as a measure of cell-surface LDLR levels) by flow cytometry. The graph shows the results plotted relative to vehicle-treated control (n = 4). ( e ) Sterol-fed HepG2 cells were cultured and treated as described in d. After exposure of cells to DiD-LDL (10 μg/ml) during the last 2 hours of treatment, cells were harvested and analyzed by flow cytometry to determine the mean fluorescent intensity of internalized DiD-LDL. The graph shows the results plotted relative to vehicle-treated control (n = 4). ( f ) Sterol-fed immortalized human hepatocytes (IHH) and HeLa cells were treated with vehicle or the indicated concentrations of triciribine for 14 hours and then harvested and analyzed by Western blotting. One representative blot is shown (n = 3). Unprocessed blots are shown in Supplementary Fig. S9 . Error bars represent SD. * p
    F ab 2 fragment antibodies are generated by pepsin digestion of whole IgG antibodies to remove most of the Fc region while leaving some of the hinge region F ab 2 fragments have two antigen binding Fab portions linked together by disulfide bonds and therefore they are divalent The average molecular weight is about 110 kDa They are used for specific applications such as to avoid binding of secondary antibodies to live cells with Fc receptors or to Protein A or Protein G Based on immunoelectrophoresis and or ELISA the antibody reacts with whole molecule mouse IgG It also reacts with the light chains of other mouse immunoglobulins No antibody was detected against non immunoglobulin serum proteins The antibody has been tested by ELISA and or solid phase adsorbed to ensure minimal cross reaction with bovine chicken goat guinea pig syrian hamster horse human rabbit and sheep serum proteins but it may cross react with immunoglobulins from other species
    https://www.bioz.com/result/alexa fluor 488 conjugated anti rabbit/product/Jackson Immuno
    Average 98 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    alexa fluor 488 conjugated anti rabbit - by Bioz Stars, 2021-09
    98/100 stars

    Images

    1) Product Images from "Triciribine increases LDLR expression and LDL uptake through stabilization of LDLR mRNA"

    Article Title: Triciribine increases LDLR expression and LDL uptake through stabilization of LDLR mRNA

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-34237-6

    Induction of LDLR expression and activity by triciribine. ( a ) Sterol-fed or sterol-starved HepG2 cells were treated with vehicle or the indicated doses of triciribine for 14 hours and then harvested for examination by Western blotting. Left panel: One representative blot is shown (n = 5). Unprocessed blots are shown in Supplementary Fig. S7 . The graph in the right panel shows quantification of the Western blots, relative to similarly cultured, vehicle-treated cells. ( b ) HepG2 cells were cultured as in a and then treated with or without 1 μM triciribine, harvested at the indicated time points and then analyzed by immunoblotting. Left panel: One representative blot is shown (n = 5). Unprocessed blots are shown in Supplementary Fig. S8 . The graph in the right panel shows quantification of the Western blots, relative to similarly cultured cells harvested at 0 hour. ( c ) Sterol-fed HepG2 cells were treated with 1 µM for 14 hours and then examined by fluorescence microscopy as described in Materials and Methods. Green: LDLR, Alexa fluor 488, blue: nuclei, DAPI. One representative experiment is shown (n = 3). ( d ) Sterol-fed HepG2 cells were exposed to vehicle or 1 μM triciribine for 14 hours before analysis for mean fluorescence intensity (as a measure of cell-surface LDLR levels) by flow cytometry. The graph shows the results plotted relative to vehicle-treated control (n = 4). ( e ) Sterol-fed HepG2 cells were cultured and treated as described in d. After exposure of cells to DiD-LDL (10 μg/ml) during the last 2 hours of treatment, cells were harvested and analyzed by flow cytometry to determine the mean fluorescent intensity of internalized DiD-LDL. The graph shows the results plotted relative to vehicle-treated control (n = 4). ( f ) Sterol-fed immortalized human hepatocytes (IHH) and HeLa cells were treated with vehicle or the indicated concentrations of triciribine for 14 hours and then harvested and analyzed by Western blotting. One representative blot is shown (n = 3). Unprocessed blots are shown in Supplementary Fig. S9 . Error bars represent SD. * p
    Figure Legend Snippet: Induction of LDLR expression and activity by triciribine. ( a ) Sterol-fed or sterol-starved HepG2 cells were treated with vehicle or the indicated doses of triciribine for 14 hours and then harvested for examination by Western blotting. Left panel: One representative blot is shown (n = 5). Unprocessed blots are shown in Supplementary Fig. S7 . The graph in the right panel shows quantification of the Western blots, relative to similarly cultured, vehicle-treated cells. ( b ) HepG2 cells were cultured as in a and then treated with or without 1 μM triciribine, harvested at the indicated time points and then analyzed by immunoblotting. Left panel: One representative blot is shown (n = 5). Unprocessed blots are shown in Supplementary Fig. S8 . The graph in the right panel shows quantification of the Western blots, relative to similarly cultured cells harvested at 0 hour. ( c ) Sterol-fed HepG2 cells were treated with 1 µM for 14 hours and then examined by fluorescence microscopy as described in Materials and Methods. Green: LDLR, Alexa fluor 488, blue: nuclei, DAPI. One representative experiment is shown (n = 3). ( d ) Sterol-fed HepG2 cells were exposed to vehicle or 1 μM triciribine for 14 hours before analysis for mean fluorescence intensity (as a measure of cell-surface LDLR levels) by flow cytometry. The graph shows the results plotted relative to vehicle-treated control (n = 4). ( e ) Sterol-fed HepG2 cells were cultured and treated as described in d. After exposure of cells to DiD-LDL (10 μg/ml) during the last 2 hours of treatment, cells were harvested and analyzed by flow cytometry to determine the mean fluorescent intensity of internalized DiD-LDL. The graph shows the results plotted relative to vehicle-treated control (n = 4). ( f ) Sterol-fed immortalized human hepatocytes (IHH) and HeLa cells were treated with vehicle or the indicated concentrations of triciribine for 14 hours and then harvested and analyzed by Western blotting. One representative blot is shown (n = 3). Unprocessed blots are shown in Supplementary Fig. S9 . Error bars represent SD. * p

    Techniques Used: Expressing, Activity Assay, Western Blot, Cell Culture, Fluorescence, Microscopy, Flow Cytometry, Cytometry

    Related Articles

    Binding Assay:

    Article Title: Genetic inactivation of SARM1 axon degeneration pathway improves outcome trajectory after experimental traumatic brain injury based on pathological, radiological, and functional measures
    Article Snippet: .. Microglia/macrophages were identified using polyclonal rabbit antibody against ionized calcium binding adaptor molecule 1 (IBA1; 1:500; Wako, Richmond, VA; Cat# 019-19741, RRID:AB_839504) followed by incubation with AlexaFluor-488-conjugated secondary antibody (1:400, Jackson ImmunoResearch Cat# 715-546-151, RRID: AB_2340850). ..

    Incubation:

    Article Title: Genetic inactivation of SARM1 axon degeneration pathway improves outcome trajectory after experimental traumatic brain injury based on pathological, radiological, and functional measures
    Article Snippet: .. Microglia/macrophages were identified using polyclonal rabbit antibody against ionized calcium binding adaptor molecule 1 (IBA1; 1:500; Wako, Richmond, VA; Cat# 019-19741, RRID:AB_839504) followed by incubation with AlexaFluor-488-conjugated secondary antibody (1:400, Jackson ImmunoResearch Cat# 715-546-151, RRID: AB_2340850). ..

    Article Title: Spermatogonia Loss Correlates with LAMA 1 Expression in Human Prepubertal Testes Stored for Fertility Preservation
    Article Snippet: .. After washing the slides in a TBS buffer, the secondary antibody incubation was performed at room temperature for 1 h in a 1:2 TBS diluted blocking buffer with a secondary antibody: Donkey anti rabbit Cy3 (711-166-152, Jackson Immuno Research) and donkey anti mouse AF488 (715-546-150, Jackson Immuno Research). ..

    Staining:

    Article Title: Sirt6 Regulates the Development of Medullary Thymic Epithelial Cells and Contributes to the Establishment of Central Immune Tolerance
    Article Snippet: .. The secondary antibodies were used for staining: Alexa Fluor 594-conjugated donkey anti-rabbit IgG (H + L) (Jackson ImmunoResearch Laboratories, 711-586-152) diluted by 1:400, Alexa Fluor 488-conjugated donkey anti-rat IgG (H + L) (Jackson ImmunoResearch Laboratories, 712-546-150) diluted by 1:400, and Alexa Fluor 488-conjugated donkey anti-mice IgG (H + L) antibodies (Jackson ImmunoResearch Laboratories, 715-546-150) diluted by 1:300. ..

    Blocking Assay:

    Article Title: Spermatogonia Loss Correlates with LAMA 1 Expression in Human Prepubertal Testes Stored for Fertility Preservation
    Article Snippet: .. After washing the slides in a TBS buffer, the secondary antibody incubation was performed at room temperature for 1 h in a 1:2 TBS diluted blocking buffer with a secondary antibody: Donkey anti rabbit Cy3 (711-166-152, Jackson Immuno Research) and donkey anti mouse AF488 (715-546-150, Jackson Immuno Research). ..

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  • 93
    Jackson Immuno alexa fluor conjugated affinipure rabbit anti human igg
    Unmutated common ancestors of MuSK mAbs bind to the MuSK autoantigen. MuSK-specific mAbs and their UCAs were tested for surface binding to MuSK on MuSK-GFP– transfected HEK cells. (A) Representative cell-based assay (CBA) contour plots are shown for the three MuSK-mAbs and their UCA. The x-axis represents GFP fluorescence intensity and, consequently, the fraction of HEK cells transfected with MuSK. The y-axis represents <t>Alexa</t> Fluor 647 fluorescence intensity, which corresponds to secondary anti–human <t>IgG</t> Fc antibody binding and, consequently, primary antibody binding to MuSK. Hence, transfected cells are located in the right quadrants and cells with MuSK antibody binding in the upper quadrants. The plots show testing with a mAb concentration of 1.25 µg/ml. (B) Binding to MuSK was tested over a series of ten two-fold dilutions of each mAb ranging from 10-0.02 µg/ml. Humanized MuSK mAb 4A3 was used as the positive control and AChR-specific mAb 637 as the negative control. The ΔMFI was calculated by subtracting the signal from non-transfected cells from the signal of transfected cells. Each data point represents a separate replicate within the same experiment, which was performed in triplicate. Bars or symbols represent means and error bars SDs. Values greater than the mean + 4SD of the negative control mAb at 1.25 µg/ml (indicated by the horizontal dotted line) were considered positive.
    Alexa Fluor Conjugated Affinipure Rabbit Anti Human Igg, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/alexa fluor conjugated affinipure rabbit anti human igg/product/Jackson Immuno
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
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    95
    Jackson Immuno alexa 488 conjugated anti rabbit igg secondary antibodies
    Secretion of Type I collagen was induced by purified <t>IgG</t> from asbestos-exposed mice. (A) L929 cells were grown to confluence, treated with purified IgG from saline- or tremolite-exposed mice for 3 days, and then collagen production was detected using anti-collagen Type I antibody followed by Alexa 488-conjugated secondary antibody. Fluorescence was quantified by LSC. (B) Collagen production by L929 cells was also measured by Sircol assay, as described in Materials and methods section. Purified IgG = IgG from tremolite-exposed mice, Asb = tremolite asbestos alone at 40 μg/cm 2 , Asb and IgG = combined treatment with both tremolite and purified IgG from tremolite-instilled mice. N = 4 in each group, * P
    Alexa 488 Conjugated Anti Rabbit Igg Secondary Antibodies, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/alexa 488 conjugated anti rabbit igg secondary antibodies/product/Jackson Immuno
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
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    86
    Jackson Immuno alexa fluor 488 conjugated goat anti rabbit polyclonal antibodies
    Flow cytometric analysis of intracellular Ob-R expression in permeabilized MCs stimulated by leptin. Representative flow cytometry histogram showing intracellular Ob-R expression ( a ): shaded tracing, isotype control; open tracings, Ob-R expression in unstimulated cells (green), and in cells stimulated with leptin at concentrations of 0.1 ng/ml (dark blue), 1 ng/ml (red), 10 ng/ml (orange), and 100 ng/ml (light blue). Expression levels of intracellular Ob-R in unstimulated and leptin-stimulated MCs ( b ). Constitutive Ob-R expression served as a control and was referred to as 100%. The results are presented as a percentage of constitutive Ob-R expression. Results are the mean of fluorescent intensity ± SD of three independent experiments performed in duplicate. The signal was visualized with green <t>Alexa</t> Fluor 488. * P
    Alexa Fluor 488 Conjugated Goat Anti Rabbit Polyclonal Antibodies, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/alexa fluor 488 conjugated goat anti rabbit polyclonal antibodies/product/Jackson Immuno
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    alexa fluor 488 conjugated goat anti rabbit polyclonal antibodies - by Bioz Stars, 2021-09
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    99
    Jackson Immuno alexa fluor 488 conjugated goat anti rabbit igg
    Mutation analysis of pV NoLS1. (A) Schematic representation of BAdV-3 pV depicting the amino acid sequence of NoLS1 and NoLS2. The thick line represents the BAdV-3 pV gene. The thin line represents the deleted region. The basic residue rich motifs (m1, m2, and m3) are shown in different font sizes. The mutations are indicated by a thin line with black-filled circles. The numbers above represent the amino acid of BAdV-3 pV. The name of the plasmids is depicted on the left of the panel. The name of the protein is depicted on the right of the panel. (B) Sub cellular localization of pV mutants. Vero cells were co-transfected with plasmid pDsRed.B23 and individual indicated plasmid DNAs. At 48 h post-transfection, cells were fixed with 4% formaldehyde. BAdV-3 wild-type and pV NoLS1 mutant proteins were visualized by indirect immunofluorescence microscopy using anti-pV antiserum and <t>Alexa</t> Fluor 488-conjugated goat anti-rabbit IgG (Jackson Immunoresearch). The DsRed.B23.1 was visualized by direct fluorescence microscopy. Nuclei were stained with DAPI. The name of the plasmids is depicted on the left of the panel. The name of the protein is depicted on the right of the panel. (C) Schematic representation of fusion proteins containing BAdV-3 pV NoLSs. The DsRed is represented by ( ). The pV is represented by ( ). The dotted box represents B23.1. The white box represents BAdV-3 pV nucleolar localization signals amino acids 21–50 or 380–389. The black box represents the EYFP gene. The numbers above represent amino acids of BAdV-3 pV. (D) Sub cellular localization of EYFP-fusion protein. Vero cells were co-transfected with individual indicated plasmids expressing EYFP, cV.EY, or EYFP-NoLS fusion proteins and pDsRed.B23.1 DNAs, and fixed with 4% formaldehyde at 48h post-transfection. The cV.EY (panel f), DsRed.B23.1 (panels c, g, k, and o), EYFP (panel b), NoLS1.EY (panel j), and NoLS2.EY (panel n) were visualized by direct fluorescence microscopy. Nuclei were stained with DAPI (panels a, e, i, and m). Merge (panels d, h, l, and p).
    Alexa Fluor 488 Conjugated Goat Anti Rabbit Igg, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/alexa fluor 488 conjugated goat anti rabbit igg/product/Jackson Immuno
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    Image Search Results


    Unmutated common ancestors of MuSK mAbs bind to the MuSK autoantigen. MuSK-specific mAbs and their UCAs were tested for surface binding to MuSK on MuSK-GFP– transfected HEK cells. (A) Representative cell-based assay (CBA) contour plots are shown for the three MuSK-mAbs and their UCA. The x-axis represents GFP fluorescence intensity and, consequently, the fraction of HEK cells transfected with MuSK. The y-axis represents Alexa Fluor 647 fluorescence intensity, which corresponds to secondary anti–human IgG Fc antibody binding and, consequently, primary antibody binding to MuSK. Hence, transfected cells are located in the right quadrants and cells with MuSK antibody binding in the upper quadrants. The plots show testing with a mAb concentration of 1.25 µg/ml. (B) Binding to MuSK was tested over a series of ten two-fold dilutions of each mAb ranging from 10-0.02 µg/ml. Humanized MuSK mAb 4A3 was used as the positive control and AChR-specific mAb 637 as the negative control. The ΔMFI was calculated by subtracting the signal from non-transfected cells from the signal of transfected cells. Each data point represents a separate replicate within the same experiment, which was performed in triplicate. Bars or symbols represent means and error bars SDs. Values greater than the mean + 4SD of the negative control mAb at 1.25 µg/ml (indicated by the horizontal dotted line) were considered positive.

    Journal: bioRxiv

    Article Title: Self-antigen driven affinity maturation is required for pathogenic monovalent IgG4 autoantibody development

    doi: 10.1101/2020.03.14.988758

    Figure Lengend Snippet: Unmutated common ancestors of MuSK mAbs bind to the MuSK autoantigen. MuSK-specific mAbs and their UCAs were tested for surface binding to MuSK on MuSK-GFP– transfected HEK cells. (A) Representative cell-based assay (CBA) contour plots are shown for the three MuSK-mAbs and their UCA. The x-axis represents GFP fluorescence intensity and, consequently, the fraction of HEK cells transfected with MuSK. The y-axis represents Alexa Fluor 647 fluorescence intensity, which corresponds to secondary anti–human IgG Fc antibody binding and, consequently, primary antibody binding to MuSK. Hence, transfected cells are located in the right quadrants and cells with MuSK antibody binding in the upper quadrants. The plots show testing with a mAb concentration of 1.25 µg/ml. (B) Binding to MuSK was tested over a series of ten two-fold dilutions of each mAb ranging from 10-0.02 µg/ml. Humanized MuSK mAb 4A3 was used as the positive control and AChR-specific mAb 637 as the negative control. The ΔMFI was calculated by subtracting the signal from non-transfected cells from the signal of transfected cells. Each data point represents a separate replicate within the same experiment, which was performed in triplicate. Bars or symbols represent means and error bars SDs. Values greater than the mean + 4SD of the negative control mAb at 1.25 µg/ml (indicated by the horizontal dotted line) were considered positive.

    Article Snippet: The binding of each mAb was detected with Alexa Fluor® -conjugated AffiniPure Rabbit Anti-Human IgG, Fcγ (309-605-008, Jackson Immunoresearch) on a BD LSRFortessa® (BD Biosciences).

    Techniques: Binding Assay, Transfection, Cell Based Assay, Crocin Bleaching Assay, Fluorescence, Concentration Assay, Positive Control, Negative Control

    Secretion of Type I collagen was induced by purified IgG from asbestos-exposed mice. (A) L929 cells were grown to confluence, treated with purified IgG from saline- or tremolite-exposed mice for 3 days, and then collagen production was detected using anti-collagen Type I antibody followed by Alexa 488-conjugated secondary antibody. Fluorescence was quantified by LSC. (B) Collagen production by L929 cells was also measured by Sircol assay, as described in Materials and methods section. Purified IgG = IgG from tremolite-exposed mice, Asb = tremolite asbestos alone at 40 μg/cm 2 , Asb and IgG = combined treatment with both tremolite and purified IgG from tremolite-instilled mice. N = 4 in each group, * P

    Journal: Journal of immunotoxicology

    Article Title: Alteration of fibroblast phenotype by asbestos-induced autoantibodies

    doi: 10.3109/1547691X.2011.562257

    Figure Lengend Snippet: Secretion of Type I collagen was induced by purified IgG from asbestos-exposed mice. (A) L929 cells were grown to confluence, treated with purified IgG from saline- or tremolite-exposed mice for 3 days, and then collagen production was detected using anti-collagen Type I antibody followed by Alexa 488-conjugated secondary antibody. Fluorescence was quantified by LSC. (B) Collagen production by L929 cells was also measured by Sircol assay, as described in Materials and methods section. Purified IgG = IgG from tremolite-exposed mice, Asb = tremolite asbestos alone at 40 μg/cm 2 , Asb and IgG = combined treatment with both tremolite and purified IgG from tremolite-instilled mice. N = 4 in each group, * P

    Article Snippet: To determine whether the autoantibodies had an effect on signal transduction in fibroblasts, L929 cells were plated to 8-well chamber slides and treated with pooled serum from asbestos-treated mice or pooled cleared serum (IgG removed using Protein G) for 2 h. The cells were then washed, fixed with ice cold acetone, blocked as above, and stained with rabbit antibodies to STAT-1 (H95) or SMAD2/3 (FL-425) (both Santa Cruz Biotech) followed by Alexa 488-conjugated anti-rabbit IgG secondary antibodies (Jackson ImmunoResearch Labs, West Grove, PA).

    Techniques: Purification, Mouse Assay, Fluorescence

    Binding of serum antibodies to mouse fibroblasts indicates presence of anti-fibroblast antibodies in serum from asbestos-instilled mice. (A) Mouse primary skin fibroblasts were fixed with paraformaldehyde and stained using serum from (left) saline- or (right) tremolite-exposed mice as the primary antibody, followed by incubation with Alexa-488-conjugated anti-mouse IgG secondary Ab. Nuclei are stained with propidium iodide. Images (at 400×) are representative of multiple experiments. (B) A cell-based ELISA was performed using L929 mouse fibroblasts, to measure the binding of serum antibodies to the cells. As in A, binding of serum antibodies was compared between sera from saline- or tremolite-treated mice. N = 7, * P

    Journal: Journal of immunotoxicology

    Article Title: Alteration of fibroblast phenotype by asbestos-induced autoantibodies

    doi: 10.3109/1547691X.2011.562257

    Figure Lengend Snippet: Binding of serum antibodies to mouse fibroblasts indicates presence of anti-fibroblast antibodies in serum from asbestos-instilled mice. (A) Mouse primary skin fibroblasts were fixed with paraformaldehyde and stained using serum from (left) saline- or (right) tremolite-exposed mice as the primary antibody, followed by incubation with Alexa-488-conjugated anti-mouse IgG secondary Ab. Nuclei are stained with propidium iodide. Images (at 400×) are representative of multiple experiments. (B) A cell-based ELISA was performed using L929 mouse fibroblasts, to measure the binding of serum antibodies to the cells. As in A, binding of serum antibodies was compared between sera from saline- or tremolite-treated mice. N = 7, * P

    Article Snippet: To determine whether the autoantibodies had an effect on signal transduction in fibroblasts, L929 cells were plated to 8-well chamber slides and treated with pooled serum from asbestos-treated mice or pooled cleared serum (IgG removed using Protein G) for 2 h. The cells were then washed, fixed with ice cold acetone, blocked as above, and stained with rabbit antibodies to STAT-1 (H95) or SMAD2/3 (FL-425) (both Santa Cruz Biotech) followed by Alexa 488-conjugated anti-rabbit IgG secondary antibodies (Jackson ImmunoResearch Labs, West Grove, PA).

    Techniques: Binding Assay, Mouse Assay, Staining, Incubation, In-Cell ELISA

    Flow cytometric analysis of intracellular Ob-R expression in permeabilized MCs stimulated by leptin. Representative flow cytometry histogram showing intracellular Ob-R expression ( a ): shaded tracing, isotype control; open tracings, Ob-R expression in unstimulated cells (green), and in cells stimulated with leptin at concentrations of 0.1 ng/ml (dark blue), 1 ng/ml (red), 10 ng/ml (orange), and 100 ng/ml (light blue). Expression levels of intracellular Ob-R in unstimulated and leptin-stimulated MCs ( b ). Constitutive Ob-R expression served as a control and was referred to as 100%. The results are presented as a percentage of constitutive Ob-R expression. Results are the mean of fluorescent intensity ± SD of three independent experiments performed in duplicate. The signal was visualized with green Alexa Fluor 488. * P

    Journal: Immunologic Research

    Article Title: Leptin receptor is expressed by tissue mast cells

    doi: 10.1007/s12026-018-9029-0

    Figure Lengend Snippet: Flow cytometric analysis of intracellular Ob-R expression in permeabilized MCs stimulated by leptin. Representative flow cytometry histogram showing intracellular Ob-R expression ( a ): shaded tracing, isotype control; open tracings, Ob-R expression in unstimulated cells (green), and in cells stimulated with leptin at concentrations of 0.1 ng/ml (dark blue), 1 ng/ml (red), 10 ng/ml (orange), and 100 ng/ml (light blue). Expression levels of intracellular Ob-R in unstimulated and leptin-stimulated MCs ( b ). Constitutive Ob-R expression served as a control and was referred to as 100%. The results are presented as a percentage of constitutive Ob-R expression. Results are the mean of fluorescent intensity ± SD of three independent experiments performed in duplicate. The signal was visualized with green Alexa Fluor 488. * P

    Article Snippet: Alexa Fluor® 488-conjugated goat anti-rabbit polyclonal antibodies were obtained from Jackson ImmunoResearch Laboratories, Inc. (West Grove, USA).

    Techniques: Flow Cytometry, Expressing, Cytometry

    Confocal microscopy images of intracellular Ob-R expression in MCs stimulated by leptin. Cells were incubated with medium alone (unstimulated cells; NS) ( a ), leptin at concentrations of 0.1 ( b ), 1 ( c ), 10 ( d ), or 100 ng/ml ( e ). Single confocal sections (midsection of cells) showing the presence of Ob-R and fluorescence intensity diagrams beside microphotographs showing the distribution of fluorescence in cells. Data shown are from one representative experiment out of three performed in duplicate. The signal was visualized with green Alexa Fluor 488

    Journal: Immunologic Research

    Article Title: Leptin receptor is expressed by tissue mast cells

    doi: 10.1007/s12026-018-9029-0

    Figure Lengend Snippet: Confocal microscopy images of intracellular Ob-R expression in MCs stimulated by leptin. Cells were incubated with medium alone (unstimulated cells; NS) ( a ), leptin at concentrations of 0.1 ( b ), 1 ( c ), 10 ( d ), or 100 ng/ml ( e ). Single confocal sections (midsection of cells) showing the presence of Ob-R and fluorescence intensity diagrams beside microphotographs showing the distribution of fluorescence in cells. Data shown are from one representative experiment out of three performed in duplicate. The signal was visualized with green Alexa Fluor 488

    Article Snippet: Alexa Fluor® 488-conjugated goat anti-rabbit polyclonal antibodies were obtained from Jackson ImmunoResearch Laboratories, Inc. (West Grove, USA).

    Techniques: Confocal Microscopy, Expressing, Incubation, Fluorescence

    Confocal microscopy images of surface Ob-R expression in MCs stimulated by leptin. Cells were incubated with medium alone (unstimulated cells; NS) ( a ), leptin at concentrations of 0.1 ( b ), 1 ( c ), 10 ( d ), or 100 ng/ml ( e ). Single confocal sections (midsection of cells) showing the presence of Ob-R and fluorescence intensity diagrams beside microphotographs showing the distribution of fluorescence in cells. Data shown are from one representative experiment out of three performed in duplicate. The signal was visualized with green Alexa Fluor 488

    Journal: Immunologic Research

    Article Title: Leptin receptor is expressed by tissue mast cells

    doi: 10.1007/s12026-018-9029-0

    Figure Lengend Snippet: Confocal microscopy images of surface Ob-R expression in MCs stimulated by leptin. Cells were incubated with medium alone (unstimulated cells; NS) ( a ), leptin at concentrations of 0.1 ( b ), 1 ( c ), 10 ( d ), or 100 ng/ml ( e ). Single confocal sections (midsection of cells) showing the presence of Ob-R and fluorescence intensity diagrams beside microphotographs showing the distribution of fluorescence in cells. Data shown are from one representative experiment out of three performed in duplicate. The signal was visualized with green Alexa Fluor 488

    Article Snippet: Alexa Fluor® 488-conjugated goat anti-rabbit polyclonal antibodies were obtained from Jackson ImmunoResearch Laboratories, Inc. (West Grove, USA).

    Techniques: Confocal Microscopy, Expressing, Incubation, Fluorescence

    Mutation analysis of pV NoLS1. (A) Schematic representation of BAdV-3 pV depicting the amino acid sequence of NoLS1 and NoLS2. The thick line represents the BAdV-3 pV gene. The thin line represents the deleted region. The basic residue rich motifs (m1, m2, and m3) are shown in different font sizes. The mutations are indicated by a thin line with black-filled circles. The numbers above represent the amino acid of BAdV-3 pV. The name of the plasmids is depicted on the left of the panel. The name of the protein is depicted on the right of the panel. (B) Sub cellular localization of pV mutants. Vero cells were co-transfected with plasmid pDsRed.B23 and individual indicated plasmid DNAs. At 48 h post-transfection, cells were fixed with 4% formaldehyde. BAdV-3 wild-type and pV NoLS1 mutant proteins were visualized by indirect immunofluorescence microscopy using anti-pV antiserum and Alexa Fluor 488-conjugated goat anti-rabbit IgG (Jackson Immunoresearch). The DsRed.B23.1 was visualized by direct fluorescence microscopy. Nuclei were stained with DAPI. The name of the plasmids is depicted on the left of the panel. The name of the protein is depicted on the right of the panel. (C) Schematic representation of fusion proteins containing BAdV-3 pV NoLSs. The DsRed is represented by ( ). The pV is represented by ( ). The dotted box represents B23.1. The white box represents BAdV-3 pV nucleolar localization signals amino acids 21–50 or 380–389. The black box represents the EYFP gene. The numbers above represent amino acids of BAdV-3 pV. (D) Sub cellular localization of EYFP-fusion protein. Vero cells were co-transfected with individual indicated plasmids expressing EYFP, cV.EY, or EYFP-NoLS fusion proteins and pDsRed.B23.1 DNAs, and fixed with 4% formaldehyde at 48h post-transfection. The cV.EY (panel f), DsRed.B23.1 (panels c, g, k, and o), EYFP (panel b), NoLS1.EY (panel j), and NoLS2.EY (panel n) were visualized by direct fluorescence microscopy. Nuclei were stained with DAPI (panels a, e, i, and m). Merge (panels d, h, l, and p).

    Journal: Frontiers in Microbiology

    Article Title: Nuclear and Nucleolar Localization of Bovine Adenovirus-3 Protein V

    doi: 10.3389/fmicb.2020.579593

    Figure Lengend Snippet: Mutation analysis of pV NoLS1. (A) Schematic representation of BAdV-3 pV depicting the amino acid sequence of NoLS1 and NoLS2. The thick line represents the BAdV-3 pV gene. The thin line represents the deleted region. The basic residue rich motifs (m1, m2, and m3) are shown in different font sizes. The mutations are indicated by a thin line with black-filled circles. The numbers above represent the amino acid of BAdV-3 pV. The name of the plasmids is depicted on the left of the panel. The name of the protein is depicted on the right of the panel. (B) Sub cellular localization of pV mutants. Vero cells were co-transfected with plasmid pDsRed.B23 and individual indicated plasmid DNAs. At 48 h post-transfection, cells were fixed with 4% formaldehyde. BAdV-3 wild-type and pV NoLS1 mutant proteins were visualized by indirect immunofluorescence microscopy using anti-pV antiserum and Alexa Fluor 488-conjugated goat anti-rabbit IgG (Jackson Immunoresearch). The DsRed.B23.1 was visualized by direct fluorescence microscopy. Nuclei were stained with DAPI. The name of the plasmids is depicted on the left of the panel. The name of the protein is depicted on the right of the panel. (C) Schematic representation of fusion proteins containing BAdV-3 pV NoLSs. The DsRed is represented by ( ). The pV is represented by ( ). The dotted box represents B23.1. The white box represents BAdV-3 pV nucleolar localization signals amino acids 21–50 or 380–389. The black box represents the EYFP gene. The numbers above represent amino acids of BAdV-3 pV. (D) Sub cellular localization of EYFP-fusion protein. Vero cells were co-transfected with individual indicated plasmids expressing EYFP, cV.EY, or EYFP-NoLS fusion proteins and pDsRed.B23.1 DNAs, and fixed with 4% formaldehyde at 48h post-transfection. The cV.EY (panel f), DsRed.B23.1 (panels c, g, k, and o), EYFP (panel b), NoLS1.EY (panel j), and NoLS2.EY (panel n) were visualized by direct fluorescence microscopy. Nuclei were stained with DAPI (panels a, e, i, and m). Merge (panels d, h, l, and p).

    Article Snippet: Anti-RPA194 antibody (C-1; Santa Cruz Biotechnology),Anti-β-actin monoclonal antibody (Sigma-Aldrich), Alexa Fluor 488-conjugated goat anti-rabbit IgG (Jackson Immunoresearch), TRITC-conjugated goat anti-mouse IgG (Jackson Immunoresearch), TRITC-conjugated goat anti-rabbit IgG (Jackson Immunoresearch), Alexa Fluor 647-conjugated goat anti-rabbit IgG (Invitrogen), Alexa Fluor 680 conjugated goat anti-rabbit antibody (Invitrogen), and IRDye800 conjugated goat anti-mouse antibody (Rockland) were purchased.

    Techniques: Mutagenesis, Sequencing, Transfection, Plasmid Preparation, Immunofluorescence, Microscopy, Fluorescence, Staining, Expressing

    Analysis of BAdV-3 pV nucleolar/nuclear localization signals. (A) Schematic representation of BAdV-3 pV. The thick black line represents BAdV-3 pV. The numbers below represent the amino acids of pV. Potential nuclear localization signal (NLS) sequences are depicted. The name of the plasmid is depicted on the left. The name of the protein is depicted on the right. (B) Schematic diagram represents mutant pV. Thick black lines represent pV gene, thin black lines represent the deleted regions. The name of the plasmid is depicted on the left. The name of the protein is depicted on the right. (C) Sub cellular localization of wild-type pV and pV mutants. Vero cells were transfected with those plasmids expressing wild-type pV and mutant pV genes individually and fixed with 4% formaldehyde at 48 h post-transfection. BAdV-3 pV was visualized by indirect immunofluorescence using anti-pV antiserum and Alexa Fluor 488-conjugated goat anti-rabbit IgG (Jackson Immunoresearch). Nuclei were stained with DAPI and nucleoli were visualized with indirect immunostaining by using RPA194 antibody (C-1; Santa Cruz Biotechnology) and TRITC-conjugated goat anti-mouse IgG (Jackson Immuno-research). The name of the plasmid is depicted on the left. The name of the protein is depicted on the right.

    Journal: Frontiers in Microbiology

    Article Title: Nuclear and Nucleolar Localization of Bovine Adenovirus-3 Protein V

    doi: 10.3389/fmicb.2020.579593

    Figure Lengend Snippet: Analysis of BAdV-3 pV nucleolar/nuclear localization signals. (A) Schematic representation of BAdV-3 pV. The thick black line represents BAdV-3 pV. The numbers below represent the amino acids of pV. Potential nuclear localization signal (NLS) sequences are depicted. The name of the plasmid is depicted on the left. The name of the protein is depicted on the right. (B) Schematic diagram represents mutant pV. Thick black lines represent pV gene, thin black lines represent the deleted regions. The name of the plasmid is depicted on the left. The name of the protein is depicted on the right. (C) Sub cellular localization of wild-type pV and pV mutants. Vero cells were transfected with those plasmids expressing wild-type pV and mutant pV genes individually and fixed with 4% formaldehyde at 48 h post-transfection. BAdV-3 pV was visualized by indirect immunofluorescence using anti-pV antiserum and Alexa Fluor 488-conjugated goat anti-rabbit IgG (Jackson Immunoresearch). Nuclei were stained with DAPI and nucleoli were visualized with indirect immunostaining by using RPA194 antibody (C-1; Santa Cruz Biotechnology) and TRITC-conjugated goat anti-mouse IgG (Jackson Immuno-research). The name of the plasmid is depicted on the left. The name of the protein is depicted on the right.

    Article Snippet: Anti-RPA194 antibody (C-1; Santa Cruz Biotechnology),Anti-β-actin monoclonal antibody (Sigma-Aldrich), Alexa Fluor 488-conjugated goat anti-rabbit IgG (Jackson Immunoresearch), TRITC-conjugated goat anti-mouse IgG (Jackson Immunoresearch), TRITC-conjugated goat anti-rabbit IgG (Jackson Immunoresearch), Alexa Fluor 647-conjugated goat anti-rabbit IgG (Invitrogen), Alexa Fluor 680 conjugated goat anti-rabbit antibody (Invitrogen), and IRDye800 conjugated goat anti-mouse antibody (Rockland) were purchased.

    Techniques: Plasmid Preparation, Mutagenesis, Transfection, Expressing, Immunofluorescence, Staining, Immunostaining

    Expression of pV. Proteins from BAdV-3 infected MDBK cells (A,B) or indicated plasmid DNA transfected cells ( C ; lanes 3 and 4) or mock infected/transfected cells were harvested at different time points, separated by SDS-PAGE, and transferred to nitrocellulose membranes. The separated proteins were probed by Western blot using anti-pV serum. The position of the molecular weight marker (lane M) in kD was used for sizing the protein bands. (D) CRL cells were transfected with plasmid pDsRed.B23 DNA and infected by BAdV-3 (panels a–d) or co-transfected with plasmid pcV and pDsRed.B23.1 DNA (panels e–h) and fixed at 24 h post-infection\transfection. The DsRed.B23.1 was visualized by direct fluorescence microscopy (panels b, f). BAdV-3 pV was visualized by indirect immunofluorescence microscopy (panels c, g) using anti-pV antiserum and Alexa Fluor 488-conjugated goat anti-rabbit IgG. The nuclei were stained with DAPI.

    Journal: Frontiers in Microbiology

    Article Title: Nuclear and Nucleolar Localization of Bovine Adenovirus-3 Protein V

    doi: 10.3389/fmicb.2020.579593

    Figure Lengend Snippet: Expression of pV. Proteins from BAdV-3 infected MDBK cells (A,B) or indicated plasmid DNA transfected cells ( C ; lanes 3 and 4) or mock infected/transfected cells were harvested at different time points, separated by SDS-PAGE, and transferred to nitrocellulose membranes. The separated proteins were probed by Western blot using anti-pV serum. The position of the molecular weight marker (lane M) in kD was used for sizing the protein bands. (D) CRL cells were transfected with plasmid pDsRed.B23 DNA and infected by BAdV-3 (panels a–d) or co-transfected with plasmid pcV and pDsRed.B23.1 DNA (panels e–h) and fixed at 24 h post-infection\transfection. The DsRed.B23.1 was visualized by direct fluorescence microscopy (panels b, f). BAdV-3 pV was visualized by indirect immunofluorescence microscopy (panels c, g) using anti-pV antiserum and Alexa Fluor 488-conjugated goat anti-rabbit IgG. The nuclei were stained with DAPI.

    Article Snippet: Anti-RPA194 antibody (C-1; Santa Cruz Biotechnology),Anti-β-actin monoclonal antibody (Sigma-Aldrich), Alexa Fluor 488-conjugated goat anti-rabbit IgG (Jackson Immunoresearch), TRITC-conjugated goat anti-mouse IgG (Jackson Immunoresearch), TRITC-conjugated goat anti-rabbit IgG (Jackson Immunoresearch), Alexa Fluor 647-conjugated goat anti-rabbit IgG (Invitrogen), Alexa Fluor 680 conjugated goat anti-rabbit antibody (Invitrogen), and IRDye800 conjugated goat anti-mouse antibody (Rockland) were purchased.

    Techniques: Expressing, Infection, Plasmid Preparation, Transfection, SDS Page, Western Blot, Molecular Weight, Marker, Fluorescence, Microscopy, Immunofluorescence, Staining