Structured Review

GE Healthcare hrp conjugated anti mouse
Bsg accumulation at the NMJ. (A and B) NMJs of w (A) or protein-trap/+ (B) larvae double stained with anti-Bsg (A) or <t>anti-GFP</t> (B) antibodies and anti-Dlg antibodies (A′ and B′). (C) elav -Gal4/+; UAS-GFP- bsg /+ larvae stained with anti-GFP (C) and <t>anti-HRP</t> (C′) antibodies. (D and E) NMJs of w (D) and protein-trap/+ (E) larvae double stained with anti-Bsg (D) or anti-GFP (E) and anti-HRP (D′ and E′) antibodies. A–E correspond to z projections of serial confocal sections. Bar, 10 μm. (F) Tangential confocal section of a protein-trap/+ synaptic bouton stained with anti-GFP and NC82 anti-BRP antibodies and plot of the intensity profiles of GFP (green) and NC82 (pink) stainings across the bouton (section is indicated by white marks on the overlay). A–E are from muscle 6/7 NMJs, and F is from muscle 4 NMJ. Bar, 5 μm.
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Images

1) Product Images from "The Ig cell adhesion molecule Basigin controls compartmentalization and vesicle release at Drosophila melanogaster synapses"

Article Title: The Ig cell adhesion molecule Basigin controls compartmentalization and vesicle release at Drosophila melanogaster synapses

Journal: The Journal of Cell Biology

doi: 10.1083/jcb.200701111

Bsg accumulation at the NMJ. (A and B) NMJs of w (A) or protein-trap/+ (B) larvae double stained with anti-Bsg (A) or anti-GFP (B) antibodies and anti-Dlg antibodies (A′ and B′). (C) elav -Gal4/+; UAS-GFP- bsg /+ larvae stained with anti-GFP (C) and anti-HRP (C′) antibodies. (D and E) NMJs of w (D) and protein-trap/+ (E) larvae double stained with anti-Bsg (D) or anti-GFP (E) and anti-HRP (D′ and E′) antibodies. A–E correspond to z projections of serial confocal sections. Bar, 10 μm. (F) Tangential confocal section of a protein-trap/+ synaptic bouton stained with anti-GFP and NC82 anti-BRP antibodies and plot of the intensity profiles of GFP (green) and NC82 (pink) stainings across the bouton (section is indicated by white marks on the overlay). A–E are from muscle 6/7 NMJs, and F is from muscle 4 NMJ. Bar, 5 μm.
Figure Legend Snippet: Bsg accumulation at the NMJ. (A and B) NMJs of w (A) or protein-trap/+ (B) larvae double stained with anti-Bsg (A) or anti-GFP (B) antibodies and anti-Dlg antibodies (A′ and B′). (C) elav -Gal4/+; UAS-GFP- bsg /+ larvae stained with anti-GFP (C) and anti-HRP (C′) antibodies. (D and E) NMJs of w (D) and protein-trap/+ (E) larvae double stained with anti-Bsg (D) or anti-GFP (E) and anti-HRP (D′ and E′) antibodies. A–E correspond to z projections of serial confocal sections. Bar, 10 μm. (F) Tangential confocal section of a protein-trap/+ synaptic bouton stained with anti-GFP and NC82 anti-BRP antibodies and plot of the intensity profiles of GFP (green) and NC82 (pink) stainings across the bouton (section is indicated by white marks on the overlay). A–E are from muscle 6/7 NMJs, and F is from muscle 4 NMJ. Bar, 5 μm.

Techniques Used: Staining

Distribution of actin cytoskeleton markers is altered in bsg larvae. (A and B) Wild-type (A) and Df/bsg 6293 (B) muscle 4 NMJs stained with anti–α-Spec antibodies. Bar, 10 μm. (C–E) Heterozygous control (C) and Df/bsg 6293 (D and E) boutons stained with anti–α-Spec (C1, D1, and E1), anti-HRP (C2, D2, and E2), and anti-Dlg (C4, D4, and E4) antibodies. Bar, 5 μm. (F and G) w (F) and Df/bsg 1217 (G) bouton stained with anti-Wasp (F and G) and anti–α-Spec (F′ and G′, red) antibodies. (H and I) w (H) and Df /bsg 6293 (I) boutons stained with anti–α-Spec (H and I) and anti-Fusch (H′ and I′) antibodies. Bar, 5 μm. (J and K) Synaptic boutons of wild-type (J) and Df/bsg 6293 (K) larvae expressing a fusion of GFP with the F-actin binding domain of Moesin (GMA), under the control of elav -Gal4. GFP-GMA expression is shown in J and K, and is in green in J′ and K′. HRP staining is shown in red in J′ and K′. Bar, 5 μm. Images A and B correspond to z projections of serial confocal sections throughout entire boutons (step size: 0.3 μm), and images C–K correspond to single optical slices taken through bouton centers. (L) Graph showing the percentage of NMJ 6/7 branches containing presynaptic Spec + or GMA + aggregates. **, P
Figure Legend Snippet: Distribution of actin cytoskeleton markers is altered in bsg larvae. (A and B) Wild-type (A) and Df/bsg 6293 (B) muscle 4 NMJs stained with anti–α-Spec antibodies. Bar, 10 μm. (C–E) Heterozygous control (C) and Df/bsg 6293 (D and E) boutons stained with anti–α-Spec (C1, D1, and E1), anti-HRP (C2, D2, and E2), and anti-Dlg (C4, D4, and E4) antibodies. Bar, 5 μm. (F and G) w (F) and Df/bsg 1217 (G) bouton stained with anti-Wasp (F and G) and anti–α-Spec (F′ and G′, red) antibodies. (H and I) w (H) and Df /bsg 6293 (I) boutons stained with anti–α-Spec (H and I) and anti-Fusch (H′ and I′) antibodies. Bar, 5 μm. (J and K) Synaptic boutons of wild-type (J) and Df/bsg 6293 (K) larvae expressing a fusion of GFP with the F-actin binding domain of Moesin (GMA), under the control of elav -Gal4. GFP-GMA expression is shown in J and K, and is in green in J′ and K′. HRP staining is shown in red in J′ and K′. Bar, 5 μm. Images A and B correspond to z projections of serial confocal sections throughout entire boutons (step size: 0.3 μm), and images C–K correspond to single optical slices taken through bouton centers. (L) Graph showing the percentage of NMJ 6/7 branches containing presynaptic Spec + or GMA + aggregates. **, P

Techniques Used: Staining, Expressing, Binding Assay

GFP expression pattern in bsg protein-trap lines. (A and A′) Third instar larva heterozygous for the protein-trap insertion, stained with anti-GFP antibodies (A and A′, green), anti-HRP antibodies (A′, red), and phalloidin (A′, blue). (B) GFP-Bsg accumulation in heterozygous larval brain. (C) Western blot of total extracts from w larvae (left) or larvae heterozygous (protein-trap/+; middle) or homozygous (protein-trap; right) for a protein-trap insertion, probed with anti-Bsg and anti-GFP antibodies. (D) Alignment of D. melanogaster and human Bsg proteins. The green triangle indicates the location of GFP insertion.
Figure Legend Snippet: GFP expression pattern in bsg protein-trap lines. (A and A′) Third instar larva heterozygous for the protein-trap insertion, stained with anti-GFP antibodies (A and A′, green), anti-HRP antibodies (A′, red), and phalloidin (A′, blue). (B) GFP-Bsg accumulation in heterozygous larval brain. (C) Western blot of total extracts from w larvae (left) or larvae heterozygous (protein-trap/+; middle) or homozygous (protein-trap; right) for a protein-trap insertion, probed with anti-Bsg and anti-GFP antibodies. (D) Alignment of D. melanogaster and human Bsg proteins. The green triangle indicates the location of GFP insertion.

Techniques Used: Expressing, Staining, Western Blot

Related Articles

Luciferase:

Article Title: Investigation of interactions between TLR2, MyD88 and TIRAP by bioluminescence resonance energy transfer is hampered by artefacts of protein overexpression
Article Snippet: .. Membranes were probed with antibodies against TLR1 (Cell Signalling Technology), TLR2 (D7G9Z, Cell Signalling Technology), MyD88 (D80F5, Cell Signalling Technology), TIRAP (EPR3509, Abcam), GFP (cross-reacts with Venus, 13.1/7.1, Roche), or Renilla luciferase (EPR17791, Abcam), followed by secondary probing with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) as previously described [ ]. .. HRP-conjugated antibodies against β-actin (AC-15, Sigma) and GAPDH (Proteintech) were used to demonstrate even lane loading.

Expressing:

Article Title: Highly Dynamic Host Actin Reorganization around Developing Plasmodium Inside Hepatocytes
Article Snippet: Cells were infected and used in time lapse experiments after 48 hours of selection, GSN expression was quantified by qRT-PCR and Western Blot as described before . .. Antibodies used in the Western blot include anti-GSN (BD Transduction Laboratories), anti-α tubulin (Sigma) and HRP conjugated anti-mouse (Amersham).

Article Title: iNOS polymorphism modulates iNOS/NO expression via impaired antioxidant and ROS content in P. vivax and P. falciparum infection
Article Snippet: 2.8 Evaluation of NOS protein in malaria patients by Western blot analysis For evaluating the induction of iNOS protein expression from P. vivax , P. falciparum and healthy subjects; PBMCs were thoroughly washed twice with ice-cold PBS and resuspended in ice cold RIPA buffer for lysis with 1% Triton X100% and 0.1% SDS following the methods as previously described and visualised by the procedure of staining using Colloidal Coomassie Blue G-250. .. Equal amounts of proteins (30 micrograms) were separated by SDS-PAGE, transferred to PVDF membrane sheets (Millipore, Billerica, MA, USA) and probed with the anti-iNOS (BD Biosciences; rabbit polyclonal, diluted 1:1000) and HRP-conjugated anti-mouse for iNOS secondary antibody (at 1:2000 in 1% skimmed milk TBST), and detection was done by enhanced chemiluminescence (ECL; Amersham Biosciences, Piscataway, NJ) Western blotting detection reagents to visualize immunoreactive bands.

Synthesized:

Article Title: TRPC5 Channel Is the Mediator of Neurotrophin-3 in Regulating Dendritic Growth via CaMKIIα in Rat Hippocampal Neurons
Article Snippet: The 7-OH-3-NC was synthesized according to the previous report and confirmed by mass spectrometry and nuclear magnetic resonance spectroscopy ( ; ). .. We purchased anti-TRPC4, anti-TRPC5, and anti-TRPC6 antibodies from Alomone Labs; anti-HSP60 and anti-TrkC from R & D Systems (for immunostaining) and Santa Cruz Biotechnology (for immunoprecipitation); anti-TrkA, anti-PLCγ1, anti-CaMKIV, and anti-pan-CaMKII from Cell Signaling Technology; anti-TrkB from Becton Dickinson; anti-α-tubulin and anti-CaMKIIα from Sigma-Aldrich; anti-IP3 R, anti-pCaMKI (Thr177), anti-pCaMKIIα (Thr286), and anti-pCaMKIV (Thr196), anti-rabbit IgG, HRP-conjugated anti-goat IgG from Santa Cruz Biotechnology; anti-CaMKIγ from Abcam; HRP-conjugated anti-mouse, rabbit IgG from GE Healthcare; and Alexa fluor 488, 546 conjugated anti-mouse, rabbit IgG from Invitrogen.

Immunostaining:

Article Title: TRPC5 Channel Is the Mediator of Neurotrophin-3 in Regulating Dendritic Growth via CaMKIIα in Rat Hippocampal Neurons
Article Snippet: .. We purchased anti-TRPC4, anti-TRPC5, and anti-TRPC6 antibodies from Alomone Labs; anti-HSP60 and anti-TrkC from R & D Systems (for immunostaining) and Santa Cruz Biotechnology (for immunoprecipitation); anti-TrkA, anti-PLCγ1, anti-CaMKIV, and anti-pan-CaMKII from Cell Signaling Technology; anti-TrkB from Becton Dickinson; anti-α-tubulin and anti-CaMKIIα from Sigma-Aldrich; anti-IP3 R, anti-pCaMKI (Thr177), anti-pCaMKIIα (Thr286), and anti-pCaMKIV (Thr196), anti-rabbit IgG, HRP-conjugated anti-goat IgG from Santa Cruz Biotechnology; anti-CaMKIγ from Abcam; HRP-conjugated anti-mouse, rabbit IgG from GE Healthcare; and Alexa fluor 488, 546 conjugated anti-mouse, rabbit IgG from Invitrogen. .. Anti-TRPC3 was produced in our laboratory ( ).

Electrophoresis:

Article Title: Preferential Secretion of Thymic Stromal Lymphopoietin (TSLP) by Terminally Differentiated Esophageal Epithelial Cells: Relevance to Eosinophilic Esophagitis (EoE)
Article Snippet: Proteins were separated by electrophoresis using NuPage 4–12% Bis-Tris gels (Invitrogen) and transferred to nitrocellulose membranes. .. HRP-conjugated anti-mouse (1:5000) (GE Healthcare) was used as a secondary antibody.

Modification:

Article Title: Human renal angiomyolipoma cells of male and female origin can migrate and are influenced by microenvironmental factors
Article Snippet: HRP-conjugated anti-mouse (Amersham-GE Healthcare, Buckinghamshire, UK) was diluted 1:6000 (ERK1/2 and pERK1/2) and 1:8000 (vinculin) in PBS Tween 0.1%. .. The ratio pERK1/2/ ERK1/2 was expressed as percentage optical density modification relative to control conditions.

Incubation:

Article Title: TKS5-positive invadopodia-like structures in human tumor surgical specimens
Article Snippet: .. Membranes were washed and incubated with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) and imaged using enhanced chemiluminescence (Supersignal West Pico PLUS, ThermoFisher Scientific) and a film developer. .. Our initial goal was to analyze TKS5 protein expression by immunofluorescence in pancreatic surgical tumor samples of various degrees of malignancy (from benign to highly malignant) to evaluate whether TKS5 expression levels correlated with the invasive/malignant potential of these lesions.

Article Title: Preferential Secretion of Thymic Stromal Lymphopoietin (TSLP) by Terminally Differentiated Esophageal Epithelial Cells: Relevance to Eosinophilic Esophagitis (EoE)
Article Snippet: Membranes were blocked with 2.5% nonfat dry milk and 2.5% bovine serum albumin overnight at 4°C, incubated with primary antibody, washed in TBST, incubated with secondary antibody, and washed in TBST. .. HRP-conjugated anti-mouse (1:5000) (GE Healthcare) was used as a secondary antibody.

Article Title: Effects of Serine 129 Phosphorylation on α-Synuclein Aggregation, Membrane Association, and Internalization *
Article Snippet: Synaptosomal membranes prepared from α-syn-deficient mice were incubated for 10 min at 37 °C with 1.5 mg/ml α-syn-deficient cytosol supplemented with 3 μg of recombinant non-phosphorylated or phosphorylated WT, A30P, or A53T α-syn as described ( ). .. Bound HRP-conjugated anti-mouse or anti-rabbit IgG was revealed by chemiluminescence using ECL Plus (GE Healthcare) and quantified with a Li-Cor Odyssey Fc imager and Image Studio software (Li-Cor).

Article Title: Human renal angiomyolipoma cells of male and female origin can migrate and are influenced by microenvironmental factors
Article Snippet: After 5 minutes and 4 hours incubation, cells were collected and treated as detailed by Mognetti et al. [ ]. .. HRP-conjugated anti-mouse (Amersham-GE Healthcare, Buckinghamshire, UK) was diluted 1:6000 (ERK1/2 and pERK1/2) and 1:8000 (vinculin) in PBS Tween 0.1%.

Cell Culture:

Article Title: Human renal angiomyolipoma cells of male and female origin can migrate and are influenced by microenvironmental factors
Article Snippet: Cells were seeded in 10 cm diameter Petri dishes, cultured until sub-confluence, then 17-β-estradiol (1 nM) was added. .. HRP-conjugated anti-mouse (Amersham-GE Healthcare, Buckinghamshire, UK) was diluted 1:6000 (ERK1/2 and pERK1/2) and 1:8000 (vinculin) in PBS Tween 0.1%.

Mass Spectrometry:

Article Title: TRPC5 Channel Is the Mediator of Neurotrophin-3 in Regulating Dendritic Growth via CaMKIIα in Rat Hippocampal Neurons
Article Snippet: The 7-OH-3-NC was synthesized according to the previous report and confirmed by mass spectrometry and nuclear magnetic resonance spectroscopy ( ; ). .. We purchased anti-TRPC4, anti-TRPC5, and anti-TRPC6 antibodies from Alomone Labs; anti-HSP60 and anti-TrkC from R & D Systems (for immunostaining) and Santa Cruz Biotechnology (for immunoprecipitation); anti-TrkA, anti-PLCγ1, anti-CaMKIV, and anti-pan-CaMKII from Cell Signaling Technology; anti-TrkB from Becton Dickinson; anti-α-tubulin and anti-CaMKIIα from Sigma-Aldrich; anti-IP3 R, anti-pCaMKI (Thr177), anti-pCaMKIIα (Thr286), and anti-pCaMKIV (Thr196), anti-rabbit IgG, HRP-conjugated anti-goat IgG from Santa Cruz Biotechnology; anti-CaMKIγ from Abcam; HRP-conjugated anti-mouse, rabbit IgG from GE Healthcare; and Alexa fluor 488, 546 conjugated anti-mouse, rabbit IgG from Invitrogen.

BIA-KA:

Article Title: Preferential Secretion of Thymic Stromal Lymphopoietin (TSLP) by Terminally Differentiated Esophageal Epithelial Cells: Relevance to Eosinophilic Esophagitis (EoE)
Article Snippet: HRP-conjugated anti-mouse (1:5000) (GE Healthcare) was used as a secondary antibody. .. HRP-conjugated anti-mouse (1:5000) (GE Healthcare) was used as a secondary antibody.

Article Title: Investigation of interactions between TLR2, MyD88 and TIRAP by bioluminescence resonance energy transfer is hampered by artefacts of protein overexpression
Article Snippet: Protein concentrations were determined using the Pierce BCA Protein Assay Kit (Thermo Fisher) and equalized between samples. .. Membranes were probed with antibodies against TLR1 (Cell Signalling Technology), TLR2 (D7G9Z, Cell Signalling Technology), MyD88 (D80F5, Cell Signalling Technology), TIRAP (EPR3509, Abcam), GFP (cross-reacts with Venus, 13.1/7.1, Roche), or Renilla luciferase (EPR17791, Abcam), followed by secondary probing with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) as previously described [ ].

Acrylamide Gel Assay:

Article Title: TKS5-positive invadopodia-like structures in human tumor surgical specimens
Article Snippet: Cells were lysed in NP40-containing lysis buffer and proteins separated in an 8% acrylamide gel. .. Membranes were washed and incubated with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) and imaged using enhanced chemiluminescence (Supersignal West Pico PLUS, ThermoFisher Scientific) and a film developer.

Western Blot:

Article Title: The Ig cell adhesion molecule Basigin controls compartmentalization and vesicle release at Drosophila melanogaster synapses
Article Snippet: .. The following antibodies were used for Western blot analysis: rabbit anti-GFP (1:500 [Santa Cruz Biotechnology, Inc.] or 1:2,500 [Torrey Pines]) and HRP-conjugated anti–mouse, –rat, or –rabbit (1:2,000; GE Healthcare) antibodies. .. Cell aggregation assay 3 ml of a 106 cells/ml culture of met-Gal4–expressing S2 cells were plated and transfected the next day with 3.5 μg of either UAS-Golgi-GFP or UAS-GFP-Bsg.

Article Title: TGF-?/Smad2/3 Signaling Directly Regulates Several miRNAs in Mouse ES Cells and Early Embryos
Article Snippet: Paragraph title: Western Blot ... Secondary antibodies used: HRP-conjugated anti-mouse (1∶10,000) (GE Healthcare, UK) and HRP conjugated anti-rabbit (1∶5,000) (Calbiochem, UK).

Article Title: Highly Dynamic Host Actin Reorganization around Developing Plasmodium Inside Hepatocytes
Article Snippet: .. Antibodies used in the Western blot include anti-GSN (BD Transduction Laboratories), anti-α tubulin (Sigma) and HRP conjugated anti-mouse (Amersham). ..

Article Title: Preferential Secretion of Thymic Stromal Lymphopoietin (TSLP) by Terminally Differentiated Esophageal Epithelial Cells: Relevance to Eosinophilic Esophagitis (EoE)
Article Snippet: Paragraph title: Western Blot ... HRP-conjugated anti-mouse (1:5000) (GE Healthcare) was used as a secondary antibody.

Article Title: Investigation of interactions between TLR2, MyD88 and TIRAP by bioluminescence resonance energy transfer is hampered by artefacts of protein overexpression
Article Snippet: Paragraph title: Western blotting and immunoprecipitation ... Membranes were probed with antibodies against TLR1 (Cell Signalling Technology), TLR2 (D7G9Z, Cell Signalling Technology), MyD88 (D80F5, Cell Signalling Technology), TIRAP (EPR3509, Abcam), GFP (cross-reacts with Venus, 13.1/7.1, Roche), or Renilla luciferase (EPR17791, Abcam), followed by secondary probing with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) as previously described [ ].

Article Title: Inhibition of glycolytic enzyme hexokinase II (HK2) suppresses lung tumor growth
Article Snippet: Paragraph title: Western blotting ... Then, primary antibody was detected with HRP-conjugated anti-mouse or anti-rabbit secondary antibody (GE Healthcare).

Article Title: Effects of Serine 129 Phosphorylation on α-Synuclein Aggregation, Membrane Association, and Internalization *
Article Snippet: Membranes were then centrifuged at 24,000 × g for 10 min, washed twice with buffer to remove excess α-syn (25 m m HEPES, 125 m m KOAc, and 2.5 m m MgCl2 ), and resuspended in 1% SDS buffer for Western blotting. .. Bound HRP-conjugated anti-mouse or anti-rabbit IgG was revealed by chemiluminescence using ECL Plus (GE Healthcare) and quantified with a Li-Cor Odyssey Fc imager and Image Studio software (Li-Cor).

Article Title: iNOS polymorphism modulates iNOS/NO expression via impaired antioxidant and ROS content in P. vivax and P. falciparum infection
Article Snippet: .. Equal amounts of proteins (30 micrograms) were separated by SDS-PAGE, transferred to PVDF membrane sheets (Millipore, Billerica, MA, USA) and probed with the anti-iNOS (BD Biosciences; rabbit polyclonal, diluted 1:1000) and HRP-conjugated anti-mouse for iNOS secondary antibody (at 1:2000 in 1% skimmed milk TBST), and detection was done by enhanced chemiluminescence (ECL; Amersham Biosciences, Piscataway, NJ) Western blotting detection reagents to visualize immunoreactive bands. .. Immunoreactive bands were digitally scanned and analyzed with NIH ImageJ for quantification by densitometry as protein expression relative to that of healthy subjects.

Article Title: Human renal angiomyolipoma cells of male and female origin can migrate and are influenced by microenvironmental factors
Article Snippet: Paragraph title: Western blotting ... HRP-conjugated anti-mouse (Amersham-GE Healthcare, Buckinghamshire, UK) was diluted 1:6000 (ERK1/2 and pERK1/2) and 1:8000 (vinculin) in PBS Tween 0.1%.

Article Title: ALOX12 is required for p53-mediated tumor suppression through a distinct ferroptosis pathway
Article Snippet: Paragraph title: Western Blotting and Antibodies. ... HRP-conjugated anti-mouse and anti-rabbit secondary antibody (GE Healthcare) and anti-rat (Southern Biotech) were used.

Electrotransfer:

Article Title: TKS5-positive invadopodia-like structures in human tumor surgical specimens
Article Snippet: After electrotransfer, membranes were blocked in 5% milk in PBS and incubated overnight at 4 °C in primary antibody diluted in PBS containing 0.1% Tween 20 and 0.3% BSA. .. Membranes were washed and incubated with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) and imaged using enhanced chemiluminescence (Supersignal West Pico PLUS, ThermoFisher Scientific) and a film developer.

Concentration Assay:

Article Title: Preferential Secretion of Thymic Stromal Lymphopoietin (TSLP) by Terminally Differentiated Esophageal Epithelial Cells: Relevance to Eosinophilic Esophagitis (EoE)
Article Snippet: Mouse anti-involucrin (Sigma, St. Louis MO) was used at a concentration of (1:10,000) and mouse anti-actin (Santa Cruz) was used at a concentration of 1:200. .. HRP-conjugated anti-mouse (1:5000) (GE Healthcare) was used as a secondary antibody.

Infection:

Article Title: Highly Dynamic Host Actin Reorganization around Developing Plasmodium Inside Hepatocytes
Article Snippet: Cells were infected and used in time lapse experiments after 48 hours of selection, GSN expression was quantified by qRT-PCR and Western Blot as described before . .. Antibodies used in the Western blot include anti-GSN (BD Transduction Laboratories), anti-α tubulin (Sigma) and HRP conjugated anti-mouse (Amersham).

Nuclear Magnetic Resonance:

Article Title: TRPC5 Channel Is the Mediator of Neurotrophin-3 in Regulating Dendritic Growth via CaMKIIα in Rat Hippocampal Neurons
Article Snippet: The 7-OH-3-NC was synthesized according to the previous report and confirmed by mass spectrometry and nuclear magnetic resonance spectroscopy ( ; ). .. We purchased anti-TRPC4, anti-TRPC5, and anti-TRPC6 antibodies from Alomone Labs; anti-HSP60 and anti-TrkC from R & D Systems (for immunostaining) and Santa Cruz Biotechnology (for immunoprecipitation); anti-TrkA, anti-PLCγ1, anti-CaMKIV, and anti-pan-CaMKII from Cell Signaling Technology; anti-TrkB from Becton Dickinson; anti-α-tubulin and anti-CaMKIIα from Sigma-Aldrich; anti-IP3 R, anti-pCaMKI (Thr177), anti-pCaMKIIα (Thr286), and anti-pCaMKIV (Thr196), anti-rabbit IgG, HRP-conjugated anti-goat IgG from Santa Cruz Biotechnology; anti-CaMKIγ from Abcam; HRP-conjugated anti-mouse, rabbit IgG from GE Healthcare; and Alexa fluor 488, 546 conjugated anti-mouse, rabbit IgG from Invitrogen.

Protein Concentration:

Article Title: iNOS polymorphism modulates iNOS/NO expression via impaired antioxidant and ROS content in P. vivax and P. falciparum infection
Article Snippet: Protein concentration was determined using a protein assay kit (Bio-Rad, Hercules, CA, USA). .. Equal amounts of proteins (30 micrograms) were separated by SDS-PAGE, transferred to PVDF membrane sheets (Millipore, Billerica, MA, USA) and probed with the anti-iNOS (BD Biosciences; rabbit polyclonal, diluted 1:1000) and HRP-conjugated anti-mouse for iNOS secondary antibody (at 1:2000 in 1% skimmed milk TBST), and detection was done by enhanced chemiluminescence (ECL; Amersham Biosciences, Piscataway, NJ) Western blotting detection reagents to visualize immunoreactive bands.

Quantitation Assay:

Article Title: TGF-?/Smad2/3 Signaling Directly Regulates Several miRNAs in Mouse ES Cells and Early Embryos
Article Snippet: Secondary antibodies used: HRP-conjugated anti-mouse (1∶10,000) (GE Healthcare, UK) and HRP conjugated anti-rabbit (1∶5,000) (Calbiochem, UK). .. Quantitation of protein bands was performed on scans of films using ImageJ ( http://rsbweb.nih.gov/ij/ ).

Binding Assay:

Article Title: Effects of Serine 129 Phosphorylation on α-Synuclein Aggregation, Membrane Association, and Internalization *
Article Snippet: Paragraph title: α-Syn Membrane Binding ... Bound HRP-conjugated anti-mouse or anti-rabbit IgG was revealed by chemiluminescence using ECL Plus (GE Healthcare) and quantified with a Li-Cor Odyssey Fc imager and Image Studio software (Li-Cor).

Molecular Weight:

Article Title: The Ig cell adhesion molecule Basigin controls compartmentalization and vesicle release at Drosophila melanogaster synapses
Article Snippet: The highest molecular weight isoform detected in total larval extracts is not detected in body wall extracts ( ) and might correspond to a glycosylated form, as described for mammalian Bsg. .. The following antibodies were used for Western blot analysis: rabbit anti-GFP (1:500 [Santa Cruz Biotechnology, Inc.] or 1:2,500 [Torrey Pines]) and HRP-conjugated anti–mouse, –rat, or –rabbit (1:2,000; GE Healthcare) antibodies.

shRNA:

Article Title: Highly Dynamic Host Actin Reorganization around Developing Plasmodium Inside Hepatocytes
Article Snippet: Paragraph title: Lentiviral shRNA knockdown of Gelsolin ... Antibodies used in the Western blot include anti-GSN (BD Transduction Laboratories), anti-α tubulin (Sigma) and HRP conjugated anti-mouse (Amersham).

Transfection:

Article Title: Investigation of interactions between TLR2, MyD88 and TIRAP by bioluminescence resonance energy transfer is hampered by artefacts of protein overexpression
Article Snippet: Western blotting and immunoprecipitation HEK293FT cells transfected as for BRET assay, THP1 cells, or human primary monocyte-derived dendritic cells were lysed as described previously [ ]. .. Membranes were probed with antibodies against TLR1 (Cell Signalling Technology), TLR2 (D7G9Z, Cell Signalling Technology), MyD88 (D80F5, Cell Signalling Technology), TIRAP (EPR3509, Abcam), GFP (cross-reacts with Venus, 13.1/7.1, Roche), or Renilla luciferase (EPR17791, Abcam), followed by secondary probing with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) as previously described [ ].

Mouse Assay:

Article Title: Effects of Serine 129 Phosphorylation on α-Synuclein Aggregation, Membrane Association, and Internalization *
Article Snippet: Synaptosomal membranes prepared from α-syn-deficient mice were incubated for 10 min at 37 °C with 1.5 mg/ml α-syn-deficient cytosol supplemented with 3 μg of recombinant non-phosphorylated or phosphorylated WT, A30P, or A53T α-syn as described ( ). .. Bound HRP-conjugated anti-mouse or anti-rabbit IgG was revealed by chemiluminescence using ECL Plus (GE Healthcare) and quantified with a Li-Cor Odyssey Fc imager and Image Studio software (Li-Cor).

Selection:

Article Title: Highly Dynamic Host Actin Reorganization around Developing Plasmodium Inside Hepatocytes
Article Snippet: Cells were infected and used in time lapse experiments after 48 hours of selection, GSN expression was quantified by qRT-PCR and Western Blot as described before . .. Antibodies used in the Western blot include anti-GSN (BD Transduction Laboratories), anti-α tubulin (Sigma) and HRP conjugated anti-mouse (Amersham).

Quantitative RT-PCR:

Article Title: Highly Dynamic Host Actin Reorganization around Developing Plasmodium Inside Hepatocytes
Article Snippet: Cells were infected and used in time lapse experiments after 48 hours of selection, GSN expression was quantified by qRT-PCR and Western Blot as described before . .. Antibodies used in the Western blot include anti-GSN (BD Transduction Laboratories), anti-α tubulin (Sigma) and HRP conjugated anti-mouse (Amersham).

Spectroscopy:

Article Title: TRPC5 Channel Is the Mediator of Neurotrophin-3 in Regulating Dendritic Growth via CaMKIIα in Rat Hippocampal Neurons
Article Snippet: The 7-OH-3-NC was synthesized according to the previous report and confirmed by mass spectrometry and nuclear magnetic resonance spectroscopy ( ; ). .. We purchased anti-TRPC4, anti-TRPC5, and anti-TRPC6 antibodies from Alomone Labs; anti-HSP60 and anti-TrkC from R & D Systems (for immunostaining) and Santa Cruz Biotechnology (for immunoprecipitation); anti-TrkA, anti-PLCγ1, anti-CaMKIV, and anti-pan-CaMKII from Cell Signaling Technology; anti-TrkB from Becton Dickinson; anti-α-tubulin and anti-CaMKIIα from Sigma-Aldrich; anti-IP3 R, anti-pCaMKI (Thr177), anti-pCaMKIIα (Thr286), and anti-pCaMKIV (Thr196), anti-rabbit IgG, HRP-conjugated anti-goat IgG from Santa Cruz Biotechnology; anti-CaMKIγ from Abcam; HRP-conjugated anti-mouse, rabbit IgG from GE Healthcare; and Alexa fluor 488, 546 conjugated anti-mouse, rabbit IgG from Invitrogen.

Lysis:

Article Title: TKS5-positive invadopodia-like structures in human tumor surgical specimens
Article Snippet: Cells were lysed in NP40-containing lysis buffer and proteins separated in an 8% acrylamide gel. .. Membranes were washed and incubated with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) and imaged using enhanced chemiluminescence (Supersignal West Pico PLUS, ThermoFisher Scientific) and a film developer.

Article Title: Inhibition of glycolytic enzyme hexokinase II (HK2) suppresses lung tumor growth
Article Snippet: Western blotting Washed the cells twice with ice-cold PBS and then lysed with ice-cold lysis buffer (50 mM HEPES pH 7.4, 150 mM NaCl, 2 mM MgCl2, 5 mM EGTA pH 8.0, 1 mM dithiothreitol, 0.5 % Triton X–100, 10 % glycerol, 1 mM Na3VO4, 1 μM microcystin–LR and protease/phosphatase inhibitor cocktail) for 30 min on ice. .. Then, primary antibody was detected with HRP-conjugated anti-mouse or anti-rabbit secondary antibody (GE Healthcare).

Article Title: iNOS polymorphism modulates iNOS/NO expression via impaired antioxidant and ROS content in P. vivax and P. falciparum infection
Article Snippet: 2.8 Evaluation of NOS protein in malaria patients by Western blot analysis For evaluating the induction of iNOS protein expression from P. vivax , P. falciparum and healthy subjects; PBMCs were thoroughly washed twice with ice-cold PBS and resuspended in ice cold RIPA buffer for lysis with 1% Triton X100% and 0.1% SDS following the methods as previously described and visualised by the procedure of staining using Colloidal Coomassie Blue G-250. .. Equal amounts of proteins (30 micrograms) were separated by SDS-PAGE, transferred to PVDF membrane sheets (Millipore, Billerica, MA, USA) and probed with the anti-iNOS (BD Biosciences; rabbit polyclonal, diluted 1:1000) and HRP-conjugated anti-mouse for iNOS secondary antibody (at 1:2000 in 1% skimmed milk TBST), and detection was done by enhanced chemiluminescence (ECL; Amersham Biosciences, Piscataway, NJ) Western blotting detection reagents to visualize immunoreactive bands.

Purification:

Article Title: Effects of Serine 129 Phosphorylation on α-Synuclein Aggregation, Membrane Association, and Internalization *
Article Snippet: Bound HRP-conjugated anti-mouse or anti-rabbit IgG was revealed by chemiluminescence using ECL Plus (GE Healthcare) and quantified with a Li-Cor Odyssey Fc imager and Image Studio software (Li-Cor). .. All measurements were done in the linear range using standardized dilutions of mouse brain lysates or purified α-syn.

SDS Page:

Article Title: Investigation of interactions between TLR2, MyD88 and TIRAP by bioluminescence resonance energy transfer is hampered by artefacts of protein overexpression
Article Snippet: Samples and Novex Sharp protein size marker (Thermo Fisher) were separated by SDS-PAGE using the NuPAGE system with 4–12% Bis-Tris gels (Thermo Fisher) and transferred onto nitrocellulose membranes using the Criterion transfer system (Bio-Rad). .. Membranes were probed with antibodies against TLR1 (Cell Signalling Technology), TLR2 (D7G9Z, Cell Signalling Technology), MyD88 (D80F5, Cell Signalling Technology), TIRAP (EPR3509, Abcam), GFP (cross-reacts with Venus, 13.1/7.1, Roche), or Renilla luciferase (EPR17791, Abcam), followed by secondary probing with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) as previously described [ ].

Article Title: Inhibition of glycolytic enzyme hexokinase II (HK2) suppresses lung tumor growth
Article Snippet: Equivalent samples were resolved by SDS-PAGE and transferred onto PVDF membranes. .. Then, primary antibody was detected with HRP-conjugated anti-mouse or anti-rabbit secondary antibody (GE Healthcare).

Article Title: iNOS polymorphism modulates iNOS/NO expression via impaired antioxidant and ROS content in P. vivax and P. falciparum infection
Article Snippet: .. Equal amounts of proteins (30 micrograms) were separated by SDS-PAGE, transferred to PVDF membrane sheets (Millipore, Billerica, MA, USA) and probed with the anti-iNOS (BD Biosciences; rabbit polyclonal, diluted 1:1000) and HRP-conjugated anti-mouse for iNOS secondary antibody (at 1:2000 in 1% skimmed milk TBST), and detection was done by enhanced chemiluminescence (ECL; Amersham Biosciences, Piscataway, NJ) Western blotting detection reagents to visualize immunoreactive bands. .. Immunoreactive bands were digitally scanned and analyzed with NIH ImageJ for quantification by densitometry as protein expression relative to that of healthy subjects.

Software:

Article Title: Effects of Serine 129 Phosphorylation on α-Synuclein Aggregation, Membrane Association, and Internalization *
Article Snippet: .. Bound HRP-conjugated anti-mouse or anti-rabbit IgG was revealed by chemiluminescence using ECL Plus (GE Healthcare) and quantified with a Li-Cor Odyssey Fc imager and Image Studio software (Li-Cor). .. All measurements were done in the linear range using standardized dilutions of mouse brain lysates or purified α-syn.

Article Title: Human renal angiomyolipoma cells of male and female origin can migrate and are influenced by microenvironmental factors
Article Snippet: HRP-conjugated anti-mouse (Amersham-GE Healthcare, Buckinghamshire, UK) was diluted 1:6000 (ERK1/2 and pERK1/2) and 1:8000 (vinculin) in PBS Tween 0.1%. .. Densitometric analysis was performed by ImageJ software.

Recombinant:

Article Title: Effects of Serine 129 Phosphorylation on α-Synuclein Aggregation, Membrane Association, and Internalization *
Article Snippet: Synaptosomal membranes prepared from α-syn-deficient mice were incubated for 10 min at 37 °C with 1.5 mg/ml α-syn-deficient cytosol supplemented with 3 μg of recombinant non-phosphorylated or phosphorylated WT, A30P, or A53T α-syn as described ( ). .. Bound HRP-conjugated anti-mouse or anti-rabbit IgG was revealed by chemiluminescence using ECL Plus (GE Healthcare) and quantified with a Li-Cor Odyssey Fc imager and Image Studio software (Li-Cor).

Bioluminescence Resonance Energy Transfer:

Article Title: Investigation of interactions between TLR2, MyD88 and TIRAP by bioluminescence resonance energy transfer is hampered by artefacts of protein overexpression
Article Snippet: Western blotting and immunoprecipitation HEK293FT cells transfected as for BRET assay, THP1 cells, or human primary monocyte-derived dendritic cells were lysed as described previously [ ]. .. Membranes were probed with antibodies against TLR1 (Cell Signalling Technology), TLR2 (D7G9Z, Cell Signalling Technology), MyD88 (D80F5, Cell Signalling Technology), TIRAP (EPR3509, Abcam), GFP (cross-reacts with Venus, 13.1/7.1, Roche), or Renilla luciferase (EPR17791, Abcam), followed by secondary probing with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) as previously described [ ].

Produced:

Article Title: TRPC5 Channel Is the Mediator of Neurotrophin-3 in Regulating Dendritic Growth via CaMKIIα in Rat Hippocampal Neurons
Article Snippet: We purchased anti-TRPC4, anti-TRPC5, and anti-TRPC6 antibodies from Alomone Labs; anti-HSP60 and anti-TrkC from R & D Systems (for immunostaining) and Santa Cruz Biotechnology (for immunoprecipitation); anti-TrkA, anti-PLCγ1, anti-CaMKIV, and anti-pan-CaMKII from Cell Signaling Technology; anti-TrkB from Becton Dickinson; anti-α-tubulin and anti-CaMKIIα from Sigma-Aldrich; anti-IP3 R, anti-pCaMKI (Thr177), anti-pCaMKIIα (Thr286), and anti-pCaMKIV (Thr196), anti-rabbit IgG, HRP-conjugated anti-goat IgG from Santa Cruz Biotechnology; anti-CaMKIγ from Abcam; HRP-conjugated anti-mouse, rabbit IgG from GE Healthcare; and Alexa fluor 488, 546 conjugated anti-mouse, rabbit IgG from Invitrogen. .. Anti-TRPC3 was produced in our laboratory ( ).

Immunoprecipitation:

Article Title: TRPC5 Channel Is the Mediator of Neurotrophin-3 in Regulating Dendritic Growth via CaMKIIα in Rat Hippocampal Neurons
Article Snippet: .. We purchased anti-TRPC4, anti-TRPC5, and anti-TRPC6 antibodies from Alomone Labs; anti-HSP60 and anti-TrkC from R & D Systems (for immunostaining) and Santa Cruz Biotechnology (for immunoprecipitation); anti-TrkA, anti-PLCγ1, anti-CaMKIV, and anti-pan-CaMKII from Cell Signaling Technology; anti-TrkB from Becton Dickinson; anti-α-tubulin and anti-CaMKIIα from Sigma-Aldrich; anti-IP3 R, anti-pCaMKI (Thr177), anti-pCaMKIIα (Thr286), and anti-pCaMKIV (Thr196), anti-rabbit IgG, HRP-conjugated anti-goat IgG from Santa Cruz Biotechnology; anti-CaMKIγ from Abcam; HRP-conjugated anti-mouse, rabbit IgG from GE Healthcare; and Alexa fluor 488, 546 conjugated anti-mouse, rabbit IgG from Invitrogen. .. Anti-TRPC3 was produced in our laboratory ( ).

Article Title: Investigation of interactions between TLR2, MyD88 and TIRAP by bioluminescence resonance energy transfer is hampered by artefacts of protein overexpression
Article Snippet: Paragraph title: Western blotting and immunoprecipitation ... Membranes were probed with antibodies against TLR1 (Cell Signalling Technology), TLR2 (D7G9Z, Cell Signalling Technology), MyD88 (D80F5, Cell Signalling Technology), TIRAP (EPR3509, Abcam), GFP (cross-reacts with Venus, 13.1/7.1, Roche), or Renilla luciferase (EPR17791, Abcam), followed by secondary probing with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) as previously described [ ].

Marker:

Article Title: Investigation of interactions between TLR2, MyD88 and TIRAP by bioluminescence resonance energy transfer is hampered by artefacts of protein overexpression
Article Snippet: Samples and Novex Sharp protein size marker (Thermo Fisher) were separated by SDS-PAGE using the NuPAGE system with 4–12% Bis-Tris gels (Thermo Fisher) and transferred onto nitrocellulose membranes using the Criterion transfer system (Bio-Rad). .. Membranes were probed with antibodies against TLR1 (Cell Signalling Technology), TLR2 (D7G9Z, Cell Signalling Technology), MyD88 (D80F5, Cell Signalling Technology), TIRAP (EPR3509, Abcam), GFP (cross-reacts with Venus, 13.1/7.1, Roche), or Renilla luciferase (EPR17791, Abcam), followed by secondary probing with HRP-conjugated anti-mouse or anti-rabbit antibodies (GE Healthcare) as previously described [ ].

Staining:

Article Title: iNOS polymorphism modulates iNOS/NO expression via impaired antioxidant and ROS content in P. vivax and P. falciparum infection
Article Snippet: 2.8 Evaluation of NOS protein in malaria patients by Western blot analysis For evaluating the induction of iNOS protein expression from P. vivax , P. falciparum and healthy subjects; PBMCs were thoroughly washed twice with ice-cold PBS and resuspended in ice cold RIPA buffer for lysis with 1% Triton X100% and 0.1% SDS following the methods as previously described and visualised by the procedure of staining using Colloidal Coomassie Blue G-250. .. Equal amounts of proteins (30 micrograms) were separated by SDS-PAGE, transferred to PVDF membrane sheets (Millipore, Billerica, MA, USA) and probed with the anti-iNOS (BD Biosciences; rabbit polyclonal, diluted 1:1000) and HRP-conjugated anti-mouse for iNOS secondary antibody (at 1:2000 in 1% skimmed milk TBST), and detection was done by enhanced chemiluminescence (ECL; Amersham Biosciences, Piscataway, NJ) Western blotting detection reagents to visualize immunoreactive bands.

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    GE Healthcare hrp conjugated vwf antibody
    ADAMTS13 –/– mice have ULVWF in plasma. <t>vWF</t> multimers in plasma from either wild-type or ADAMTS13 –/– C57BL/6 mice were separated by electrophoresis on SDS 1% agarose gels and then detected by Western blotting with an <t>HRP-conjugated</t> vWF antibody. Numbers in the center indicate the number of vWF protomers contained in the indicated band. ADAMTS13 –/– mice displayed a series of high–molecular weight multimers that were larger than the largest seen in wild-type mice.
    Hrp Conjugated Vwf Antibody, supplied by GE Healthcare, 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/hrp conjugated vwf antibody/product/GE Healthcare
    Average 86 stars, based on 1 article reviews
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    hrp conjugated vwf antibody - by Bioz Stars, 2020-04
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    86
    GE Healthcare ecl hrp conjugated anti mouse antibody
    Effect of SMT C1100 on in vivo utrophin levels in the mdx mouse. (A) Two-fold increase in utrophin mRNA following daily oral administration of mdx mice with SMT C1100 (50 mg/kg/day) or vehicle only (PBS-Tween-20 (0.1%)/5% DMSO) from three weeks of age for four weeks. Results represent the mean ± S.E from six mice per treatment group and are corrected for β-actin. *p = 0.019; (B) Utrophin protein levels in heart and diaphragm following treatment of mdx mice as described in (A) above. Blots were stained with anti-utrophin (MANCHO3; 1∶100) and <t>ECL</t> <t>HRP-conjugated</t> anti-mouse antibody (GE Healthcare). Bands were quantified using Image J and arbitrary units represent utrophin levels corrected for equal loading by α-actinin immunostaining. Results represent a mean ±S.E from eight mice per treatment group except for heart vehicle only which is based on n = 7. *p
    Ecl Hrp Conjugated Anti Mouse Antibody, supplied by GE Healthcare, 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/ecl hrp conjugated anti mouse antibody/product/GE Healthcare
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    ecl hrp conjugated anti mouse antibody - by Bioz Stars, 2020-04
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    85
    GE Healthcare mouse anti e tag hrp conjugated antibody
    ELISA (A) and Western blot (B) showing binding of anti-SEB ScFv with SEB antigen. ELISA was performed with different dilutions of soluble anti-SEB ScFv antibody. Periplasmic extract of untransformed HB2151 cells was used as a negative control (OD, 0.0945). (B) Lane 1, prestained molecular mass markers (70, 55, 35, 27, and 15 kDa); lane 2, r-SEB band at 30.5 kDa showing binding with anti-SEB ScFv. The bound anti-SEB ScFv antibody was detected using <t>anti-E-tag</t> <t>HRP-conjugated</t> antibody.
    Mouse Anti E Tag Hrp Conjugated Antibody, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 85 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    mouse anti e tag hrp conjugated antibody - by Bioz Stars, 2020-04
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    ADAMTS13 –/– mice have ULVWF in plasma. vWF multimers in plasma from either wild-type or ADAMTS13 –/– C57BL/6 mice were separated by electrophoresis on SDS 1% agarose gels and then detected by Western blotting with an HRP-conjugated vWF antibody. Numbers in the center indicate the number of vWF protomers contained in the indicated band. ADAMTS13 –/– mice displayed a series of high–molecular weight multimers that were larger than the largest seen in wild-type mice.

    Journal: The Journal of Clinical Investigation

    Article Title: N-acetylcysteine reduces the size and activity of von Willebrand factor in human plasma and mice

    doi: 10.1172/JCI41062

    Figure Lengend Snippet: ADAMTS13 –/– mice have ULVWF in plasma. vWF multimers in plasma from either wild-type or ADAMTS13 –/– C57BL/6 mice were separated by electrophoresis on SDS 1% agarose gels and then detected by Western blotting with an HRP-conjugated vWF antibody. Numbers in the center indicate the number of vWF protomers contained in the indicated band. ADAMTS13 –/– mice displayed a series of high–molecular weight multimers that were larger than the largest seen in wild-type mice.

    Article Snippet: Plasma vWF from either control or NAC-treated mice was separated on a 1.5% agarose gel by electrophoresis. vWF was detected by HRP-conjugated vWF antibody in Western blots, and the chemiluminescent signals were captured by CCD camera (ImageQuant 350; GE Healthcare Life Sciences).

    Techniques: Mouse Assay, Electrophoresis, Western Blot, Molecular Weight

    NAC inhibits the ability of vWF to bind platelets and collagen. ( A ) Pooled plasma was incubated with NAC at concentrations of 0, 4, 8, 16, or 32 mM at 37°C for 30 minutes. Residual NAC was then alkylated with NEM. The reaction mixture was used as the source of vWF to agglutinate fixed platelets in the presence of 1 mg/ml of ristocetin. Data represent mean ± SD from 4 independent experiments. ( B ) Purified vWF was incubated with NAC at concentrations of 0, 2, 4, 8, and 16 mM at 37°C for 30 minutes. Residual NAC was then alkylated with NEM. The reaction mixture was added to an ELISA plate coated with collagen, and bound vWF was detected with an HRP-conjugated vWF antibody. Data represent mean ± SD from 3 independent experiments.

    Journal: The Journal of Clinical Investigation

    Article Title: N-acetylcysteine reduces the size and activity of von Willebrand factor in human plasma and mice

    doi: 10.1172/JCI41062

    Figure Lengend Snippet: NAC inhibits the ability of vWF to bind platelets and collagen. ( A ) Pooled plasma was incubated with NAC at concentrations of 0, 4, 8, 16, or 32 mM at 37°C for 30 minutes. Residual NAC was then alkylated with NEM. The reaction mixture was used as the source of vWF to agglutinate fixed platelets in the presence of 1 mg/ml of ristocetin. Data represent mean ± SD from 4 independent experiments. ( B ) Purified vWF was incubated with NAC at concentrations of 0, 2, 4, 8, and 16 mM at 37°C for 30 minutes. Residual NAC was then alkylated with NEM. The reaction mixture was added to an ELISA plate coated with collagen, and bound vWF was detected with an HRP-conjugated vWF antibody. Data represent mean ± SD from 3 independent experiments.

    Article Snippet: Plasma vWF from either control or NAC-treated mice was separated on a 1.5% agarose gel by electrophoresis. vWF was detected by HRP-conjugated vWF antibody in Western blots, and the chemiluminescent signals were captured by CCD camera (ImageQuant 350; GE Healthcare Life Sciences).

    Techniques: Incubation, Purification, Enzyme-linked Immunosorbent Assay

    Effect of SMT C1100 on in vivo utrophin levels in the mdx mouse. (A) Two-fold increase in utrophin mRNA following daily oral administration of mdx mice with SMT C1100 (50 mg/kg/day) or vehicle only (PBS-Tween-20 (0.1%)/5% DMSO) from three weeks of age for four weeks. Results represent the mean ± S.E from six mice per treatment group and are corrected for β-actin. *p = 0.019; (B) Utrophin protein levels in heart and diaphragm following treatment of mdx mice as described in (A) above. Blots were stained with anti-utrophin (MANCHO3; 1∶100) and ECL HRP-conjugated anti-mouse antibody (GE Healthcare). Bands were quantified using Image J and arbitrary units represent utrophin levels corrected for equal loading by α-actinin immunostaining. Results represent a mean ±S.E from eight mice per treatment group except for heart vehicle only which is based on n = 7. *p

    Journal: PLoS ONE

    Article Title: Daily Treatment with SMTC1100, a Novel Small Molecule Utrophin Upregulator, Dramatically Reduces the Dystrophic Symptoms in themdx Mouse

    doi: 10.1371/journal.pone.0019189

    Figure Lengend Snippet: Effect of SMT C1100 on in vivo utrophin levels in the mdx mouse. (A) Two-fold increase in utrophin mRNA following daily oral administration of mdx mice with SMT C1100 (50 mg/kg/day) or vehicle only (PBS-Tween-20 (0.1%)/5% DMSO) from three weeks of age for four weeks. Results represent the mean ± S.E from six mice per treatment group and are corrected for β-actin. *p = 0.019; (B) Utrophin protein levels in heart and diaphragm following treatment of mdx mice as described in (A) above. Blots were stained with anti-utrophin (MANCHO3; 1∶100) and ECL HRP-conjugated anti-mouse antibody (GE Healthcare). Bands were quantified using Image J and arbitrary units represent utrophin levels corrected for equal loading by α-actinin immunostaining. Results represent a mean ±S.E from eight mice per treatment group except for heart vehicle only which is based on n = 7. *p

    Article Snippet: Morris, Oswestry, UK) and ECL HRP-conjugated anti-mouse antibody (GE Healthcare).

    Techniques: In Vivo, Mouse Assay, Staining, Immunostaining

    In vitro activity of SMT C1100. (A) SMT C1100 dose response in murine H2k-mdx utrnA-luc cells expressing the human utrophin promoter linked to a luciferase reporter gene. Cells were treated with compound for 48 h in standard growth medium containing 0.3% DMSO. The chart shows relative luminescence (RLU) in relation to five different doses of SMT C1100. A Four Parameter Logistic Model was used to generate an EC 50 . Points represent a mean ±S.E. of three experiments and are typical of the results for all batches of SMTC1100. The structure for SMTC1100 is shown; (B) SMT C1100 significantly increased mRNA copy number of the utrophin transcript in SkMC cells. In this assay Gene Expression Assay 4326315 was used for β-actin detection and Gene Expression Assay Hu01125984_m1 was used for utrophin transcript detection (both Applied Biosystems). Cells were exposed to SMT C1100 in standard media with 1% DMSO (vehicle) for 72 hours with six biological replicates. *p = 0.01 relative to vehicle only; (C) Utrophin protein levels in human DMD cell line treated with SMT C1100 (1 µM) or vehicle (0.1% DMSO). Blots were stained with anti-utrophin (MANCHO3; 1∶100) and ECL HRP-conjugated anti-mouse antibody (GE Healthcare). Bands were quantified using Image J and arbitrary units represent utrophin levels corrected for equal loading by α-actinin immunostaining. Results represent a mean ± S.E based on n = 3. †p = 0.00683; §p

    Journal: PLoS ONE

    Article Title: Daily Treatment with SMTC1100, a Novel Small Molecule Utrophin Upregulator, Dramatically Reduces the Dystrophic Symptoms in themdx Mouse

    doi: 10.1371/journal.pone.0019189

    Figure Lengend Snippet: In vitro activity of SMT C1100. (A) SMT C1100 dose response in murine H2k-mdx utrnA-luc cells expressing the human utrophin promoter linked to a luciferase reporter gene. Cells were treated with compound for 48 h in standard growth medium containing 0.3% DMSO. The chart shows relative luminescence (RLU) in relation to five different doses of SMT C1100. A Four Parameter Logistic Model was used to generate an EC 50 . Points represent a mean ±S.E. of three experiments and are typical of the results for all batches of SMTC1100. The structure for SMTC1100 is shown; (B) SMT C1100 significantly increased mRNA copy number of the utrophin transcript in SkMC cells. In this assay Gene Expression Assay 4326315 was used for β-actin detection and Gene Expression Assay Hu01125984_m1 was used for utrophin transcript detection (both Applied Biosystems). Cells were exposed to SMT C1100 in standard media with 1% DMSO (vehicle) for 72 hours with six biological replicates. *p = 0.01 relative to vehicle only; (C) Utrophin protein levels in human DMD cell line treated with SMT C1100 (1 µM) or vehicle (0.1% DMSO). Blots were stained with anti-utrophin (MANCHO3; 1∶100) and ECL HRP-conjugated anti-mouse antibody (GE Healthcare). Bands were quantified using Image J and arbitrary units represent utrophin levels corrected for equal loading by α-actinin immunostaining. Results represent a mean ± S.E based on n = 3. †p = 0.00683; §p

    Article Snippet: Morris, Oswestry, UK) and ECL HRP-conjugated anti-mouse antibody (GE Healthcare).

    Techniques: In Vitro, Activity Assay, Expressing, Luciferase, Staining, Immunostaining

    ELISA (A) and Western blot (B) showing binding of anti-SEB ScFv with SEB antigen. ELISA was performed with different dilutions of soluble anti-SEB ScFv antibody. Periplasmic extract of untransformed HB2151 cells was used as a negative control (OD, 0.0945). (B) Lane 1, prestained molecular mass markers (70, 55, 35, 27, and 15 kDa); lane 2, r-SEB band at 30.5 kDa showing binding with anti-SEB ScFv. The bound anti-SEB ScFv antibody was detected using anti-E-tag HRP-conjugated antibody.

    Journal: Applied and Environmental Microbiology

    Article Title: Construction of a Single-Chain Variable-Fragment Antibody against the Superantigen Staphylococcal Enterotoxin B ▿

    doi: 10.1128/AEM.01441-10

    Figure Lengend Snippet: ELISA (A) and Western blot (B) showing binding of anti-SEB ScFv with SEB antigen. ELISA was performed with different dilutions of soluble anti-SEB ScFv antibody. Periplasmic extract of untransformed HB2151 cells was used as a negative control (OD, 0.0945). (B) Lane 1, prestained molecular mass markers (70, 55, 35, 27, and 15 kDa); lane 2, r-SEB band at 30.5 kDa showing binding with anti-SEB ScFv. The bound anti-SEB ScFv antibody was detected using anti-E-tag HRP-conjugated antibody.

    Article Snippet: Plasmid pCANTAB 5E, Escherichia coli strains TG1 and HB2151, M13K07 helper phage, mouse anti-M13 horseradish peroxidase (HRP)-conjugated antibody, mouse anti-E tag HRP-conjugated antibody, and restriction enzymes SfiI and NotI were procured from GE Healthcare UK Limited (Buckinghamshire, United Kingdom).

    Techniques: Enzyme-linked Immunosorbent Assay, Western Blot, Binding Assay, Negative Control

    LGG induces ROS that oxidizes cysteines. (A) Intestinal epithelial cells (SKCO15) were contacted with 10 8 CFU/mL of E. coli or LGG for 15 min, and then with 15 µM HydroCy3 for 30 min before confocal microscopic analysis at 555 nm, (scale bars 20 µm). (B) SKCO15 were contacted with 10 8 CFU/mL of E. coli or LGG for 15 min, labeled for 30 min with a thiol-reactive, Thiol Tracker™ fluorescence probe, and then analyzed by fluorescence microscopy at 405 nm (scale bars 200 µm). Mean image intensity is shown at bottom left for A and B. (C) Biotinylated-iodoacetamide (BIAM) labeling of cysteine residues in lysates of LGG or E. coli contacted SKCO15 cells, followed by pull-down of labeled residues with streptavidin conjugated agarose and detected by Western blot using HRP conjugated streptavidin as a probe. The relative intensity of each lane of the blot is shown in arbitrary units to the left. Each value was normalized to calnexin that served as a loading control to give the relative oxidation amounts. (D) Dual labeling of LGG or E. coli contacted (15 mins) SKCO15 cultured cells with HydroCy3 (red) and Lysotracker (green). Note co-localization of lysotracker and hydro-Cy3 in LGG contacted cells (bars 10 µm).

    Journal: Redox Biology

    Article Title: Proteomic analysis of microbial induced redox-dependent intestinal signaling

    doi: 10.1016/j.redox.2018.11.011

    Figure Lengend Snippet: LGG induces ROS that oxidizes cysteines. (A) Intestinal epithelial cells (SKCO15) were contacted with 10 8 CFU/mL of E. coli or LGG for 15 min, and then with 15 µM HydroCy3 for 30 min before confocal microscopic analysis at 555 nm, (scale bars 20 µm). (B) SKCO15 were contacted with 10 8 CFU/mL of E. coli or LGG for 15 min, labeled for 30 min with a thiol-reactive, Thiol Tracker™ fluorescence probe, and then analyzed by fluorescence microscopy at 405 nm (scale bars 200 µm). Mean image intensity is shown at bottom left for A and B. (C) Biotinylated-iodoacetamide (BIAM) labeling of cysteine residues in lysates of LGG or E. coli contacted SKCO15 cells, followed by pull-down of labeled residues with streptavidin conjugated agarose and detected by Western blot using HRP conjugated streptavidin as a probe. The relative intensity of each lane of the blot is shown in arbitrary units to the left. Each value was normalized to calnexin that served as a loading control to give the relative oxidation amounts. (D) Dual labeling of LGG or E. coli contacted (15 mins) SKCO15 cultured cells with HydroCy3 (red) and Lysotracker (green). Note co-localization of lysotracker and hydro-Cy3 in LGG contacted cells (bars 10 µm).

    Article Snippet: Anti-mouse secondary antibody conjugated to HRP was obtained from GE, while the streptavidin conjugated HRP was obtained from Abcam.

    Techniques: Labeling, Fluorescence, Microscopy, Western Blot, Cell Culture