goat anti anxa1 antibody  (R&D Systems)

Journal: Frontiers in Immunology

Article Title: GPCRs overexpression and impaired fMLP-induced functions in neutrophils from chronic kidney disease patients

doi: 10.3389/fimmu.2024.1387566

Figure Lengend Snippet: CKD neutrophils overexpress intracellular AnxA1. Increased FPR2 expression in CKD neutrophils (A) . AnxA1 expression in neutrophils at intracellular (B) and membrane compartments (C) . AnxA1 secretion from neutrophils (D) and apoptotic neutrophils (E) cultured for 18 hours under serum starvation conditions. (Control n= 3–11 and CKD n= 3–36) **p<0.01; ***p<0.001 CKD vs Control. ns, Not significant.

Article Snippet: Patients’ samples were incubated with anti-AnxA1 goat antibody (1:100) (AF3770; R&D SYSTEM) and anti-myeloperoxidase rabbit antibody (1:50; Biodesign International, Inc., Maine, USA) overnight at 4°C, followed by incubation with anti-goat Alexa Fluor 467 and anti-rabbit Alexa Fluor 488 secondary antibodies (1:200; Invitrogen).

Techniques: Expressing, Membrane, Cell Culture, Control

Journal: Frontiers in Immunology

Article Title: GPCRs overexpression and impaired fMLP-induced functions in neutrophils from chronic kidney disease patients

doi: 10.3389/fimmu.2024.1387566

Figure Lengend Snippet: Infiltrate neutrophils in CKD kidney biopsies are AnxA + . H&E (A–E) and PAS staining (F–J) . Myeloperoxidase (MPO) and AnxA1 labeling (K–O) . Five CKD biopsies were selected and stained for morphological, fibrosis visualization, and immunofluorescence. Figure 5D – Insert: * immune cells infiltrated; MPO and AnxA1 labeling. The image is representative of at least three fields analyzed on each biopsy.

Article Snippet: Patients’ samples were incubated with anti-AnxA1 goat antibody (1:100) (AF3770; R&D SYSTEM) and anti-myeloperoxidase rabbit antibody (1:50; Biodesign International, Inc., Maine, USA) overnight at 4°C, followed by incubation with anti-goat Alexa Fluor 467 and anti-rabbit Alexa Fluor 488 secondary antibodies (1:200; Invitrogen).

Techniques: Staining, Labeling, Immunofluorescence

Journal: International Journal of Molecular Sciences

Article Title: Identification of 56 Proteins Involved in Embryo–Maternal Interactions in the Bovine Oviduct

doi: 10.3390/ijms21020466

Figure Lengend Snippet: Proteins identified as interacting with both the 4–6 cell and morula stages.

Article Snippet: Goat polyclonal antibody raised against ANXA1 (AP22515PU-N) was obtained from Origene (Rockville, MD, USA).

Techniques: Molecular Weight, Control

Journal: International Journal of Molecular Sciences

Article Title: Identification of 56 Proteins Involved in Embryo–Maternal Interactions in the Bovine Oviduct

doi: 10.3390/ijms21020466

Figure Lengend Snippet: Immunolocalization of embryo-interacting proteins in OF-treated embryos ( a , c , e ) and controls ( b , d , f ). Proteins of interest are in red and nuclei are in blue. Embryos were incubated with anti-ANXA1 ( a , b ), anti-OVGP1 ( c , d ) and anti-PYGL ( d , e ). Inserts: controls with isotypes instead of primary antibodies.

Article Snippet: Goat polyclonal antibody raised against ANXA1 (AP22515PU-N) was obtained from Origene (Rockville, MD, USA).

Techniques: Incubation

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: Annexin-A1 reduces survival and enhances morbidity and virus replication after influenza virus A/PR/8/34 infection. Wild-type and Annexin-A1 (ANXA1) −/− mice were infected intra-tracheally with 25 pfu/g influenza A/PR/8/34 and survival was monitored up to 12 days post infection. ( a ) Weight loss was significantly reduced in ANXA1 −/− ( n =10) mice compared with wild-type mice ( n =10). ( b ) ANXA1 −/− mice ( n =10) exhibited significantly reduced mortality compared with the wild type mice ( n =10) ( p =0.0084, log rank). ( c ) Lungs were extracted at day 5 post-infection and assessed for viral titers using plaque assays on MDCK cells. ( d ) M viral gene expression or ( e ) NS1 gene expression was assessed at indicated days post-infection using qPCR. CT values were converted to viral titers using a standard curve for M gene or NS1 expression. ( f ) Mice were infected with a sub-lethal dose of IAV (12.5 pfu/g). Weight loss was significantly reduced in ANXA1 −/− ( n =4) mice compared with wild-type mice ( n =4) at day 3 post-infection. ( g ) Lungs were extracted at day 3 post-infection and assessed for viral titers using plaque assays on MDCK cells. ( h ) NS1 viral gene expression was assessed at day 3 post-infection using qPCR. CT values were converted to viral titers using a standard curve for M gene or NS1 expression. * P <0.05, ** P <0.01

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

Techniques: Virus, Infection, Gene Expression, Expressing

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: ANXA1 deficiency does not affect cytokine production in the lungs, but enhances leukocyte infiltration wild-type and Annexin-A1 (ANXA1) −/− mice were infected intra-tracheally with influenza A/PR/8/34. ( a – c ) Cell free bronchoalveolar lavage fluids were analyzed at indicated days post-infection for the indicated cytokines. ** P <0.01 versus WT on the same day. ( d ) Total cells were counted using trypan blue staining. ( e – i ) Differential leukocyte quantification was performed using multicolor flow cytometry with the indicated antibodies. * P <0.05; ** P <0.01 versus UI control. ξ P <0.05; ξξ P <0.01 versus WT on the same day. ( j ) Lungs were harvested 5 days and 10 days post-infection from WT and ANXA1 −/− mice, respectively, and stained with haematoxylin and Eosin. ( k ) Histological scoring of representative lung sections. Results shown are of n =4–5 mice per group, repeated at least twice

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

Techniques: Infection, Staining, Flow Cytometry, Control

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: ANXA1 is induced and cleaved upon IAV infection. ( a and b ) Wild-type and Annexin-A1 (ANXA1) −/− deficient mice were infected intra-tracheally with influenza A/PR/8/34. Lungs were extracted at indicated days post-infection and assessed for ANXA1 expression using qPCR (mean±S.E. of 3 mice) or Western blotting. ( c and d ) A549 lung epithelial cells were infected with 1 MOI of A/PR8. ANXA1 expression was measured at indicated time points using qPCR (mean±S.E. of at least 3 independent experiments,* P <0.05, ** P <0.01 versus UI) or Western blotting. ( e ) ANXA1 expression in nasal swabs from healthy controls or influenza A infected patients was measured using ELISA. * P <0.05 in 12 controls and 11 patients

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

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

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: ANXA1 promotes virus replication in vitro . ( a and b ) Scramble or ANXA1-shRNA transfected A549 lung epithelial cells were infected with 1 MOI of influenza A/PR8 for the indicated time points. Virus was quantified in supernatants using qPCR for M gene expression or NS1 expression. CT values were converted to viral particles using a standard curve for M1 or NS1 expression (mean±S.D. of duplicates with at least three independent repeats). ( c ) Viral titers were measured in the supernatants using plaque assays (mean±S.D. of duplicates with at least three independent repeats). ( d ) A549 cells transfected with empty vector (EV) or ANXA1 pcDNA3.1-V5 (ANXA1-V5) were infected with 1 MOI of influenza A/PR8. Virus was quantified in supernatants using qPCR for M1 gene expression. CT values were converted to viral particles using a standard curve for M1 expression (mean±S.E. of three independent experiments. * P <0.05; ** P <0.01 versus Scr-sh or EV at the same time points

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

Techniques: Virus, In Vitro, shRNA, Transfection, Infection, Gene Expression, Expressing, Plasmid Preparation

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: ANXA1 enhances IAV binding and nuclear infection. ( a and b ) Scramble or ANXA1-shRNA transfected A549 lung epithelial cells were infected with 1 MOI of influenza A/PR8 for the indicated time points on ice. ( c and d ). A549 cells were washed with PBS or PBS+3 mM EDTA for 30 min prior to infection with 1 MOI of influenza A/PR8 for 15 min on ice. ( e – h ) Scramble or ANXA1-shRNA transfected A549 lung epithelial cells were infected with 1 MOI of influenza A/PR8 for the indicated time points at 37 °C. Viral NP (red) and ANXA1 (green) were visualized using confocal microscopy. For a – f , the number of viral particles were quantified per cell. For g and h , the per cent of cells with infected nuclei were counted per field

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

Techniques: Binding Assay, Infection, shRNA, Transfection, Confocal Microscopy

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: ANXA1 enhances early and late endosome trafficking of IAV. Scramble or ANXA1-shRNA transfected A549 lung epithelial cells were infected with 1 MOI of influenza A/PR8 for the indicated time points. ( a ) Percentage of virus colocalized in early endosomes was quantified. ( b ) Number of early endosomes per cell was quantified. ( c and d ) Early endosomes were stained with Rab5a (green), viral NP (red) and DAPI (blue). ( e ) Percentage of virus colocalized in late endosomes was quantified. ( f ) Number of late endosomes per cell was quantified. ( g and h ) Late endosomes were stained with Rab7a (green), viral NP (red) and DAPI (blue). ( i ) Colocalization of ANXA (red) and early (Rab5a)/late (Rab7a) endosomes (green)

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

Techniques: shRNA, Transfection, Infection, Virus, Staining

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: ANXA1 enhances IAV-induced apoptosis. Wild-type and Annexin-A1 (ANXA1) −/− deficient mice were infected intra-tracheally with influenza A/PR/8/34. Lungs were extracted at 3 days post-infection and apoptosis was assessed using ( a ) caspase 3 or 7 cleavage or ( b ) Annexin-V/PI staining. Tubulin was used as a loading control. ( c and d ) A549 lung epithelial cells (control or shRNA silenced for ANXA1 (ANXA1-sh) were infected with 1 MOI of influenza A/PR8. Cell viability was assessed after 24 h using crystal violet staining. ( e ) Caspase-3 activity was measured in control or ANXA1-siRNA A549 cells uninfected or infected with 1 MOI of influenza A/PR/8/34 for indicated times. * P <0.05; ** P <0.01 versus 0. ( f ) A549 cells were infected with media (M) or indicated MOI of influenza A/PR8 for 24 h. Cell lysates were subjected to immunoblot analysis for the indicated antibodies. GADPH was used as a loading control. ( g and h ) Scramble and ANXA1-shRNA transfected A549 cells were infected with 1 MOI of influenza A/PR8 and cells were lysed at the indicated time points and subjected to immunoblot analysis for the indicated antibodies. ( i ) Scramble and ANXA1-shRNA transfected A549 cells were infected with 5 MOI of influenza A/PR8 and cells were fractionated into mitochondrial and cytosolic fractions at the indicated time points and subjected to immunoblot analysis for the indicated antibodies. AIF was used as a mitochondrial loading control and GAPDH was used as a cytosolic loading control

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

Techniques: Infection, Staining, Control, shRNA, Activity Assay, Western Blot, Transfection

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: ANXA1 inhibits IAV-induced NF- κ B activation and enhances AKT activation. ( a and b ) Scramble and ANXA1-shRNA transfected A549 cells, or EV and ANXA1-V5 cells were transfected with pNF- κ B-luc reporter and renilla-plasmids for 48 h and infected with 1 MOI of influenza A/PR/8/34 for 4 h. Luciferase activity was measured after 4 h of infection. Luciferase activity was calculated by normalizing with the pGL-luc reporter controls (mean±S.E. of at least 3 independent experiments,** P <0.01 versus Scr-sh infected cells). ( c and d ) Scramble and ANXA1-shRNA transfected A549 cells were infected with 1 MOI of influenza A/PR/8/34 and cells were lysed at the indicated time points and subjected to immunoblot analysis for the indicated antibodies

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

Techniques: Activation Assay, shRNA, Transfection, Infection, Luciferase, Activity Assay, Western Blot

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: ANXA1 associates with NS1 viral protein. ( a ) A549 lung epithelial cells were infected with 1 MOI of A/PR8 and immunoprecipitation was performed with mouse IgG or mouse anti-ANXA1 antibody and probed with the antibodies to the viral proteins. ( b ) 293T cells were transfected with the indicated plasmids for 36 h and co-immunoprecipitation was performed by pulling down with Flag-beads and immunostained with the indicated antibodies. Results are representative of two independent experiments

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

Techniques: Infection, Immunoprecipitation, Transfection

Journal: Cell Death and Differentiation

Article Title: Influenza A virus enhances its propagation through the modulation of Annexin-A1 dependent endosomal trafficking and apoptosis

doi: 10.1038/cdd.2016.19

Figure Lengend Snippet: Summary figure. During the IAV lifecycle, IAV binds to the host cell and is endocytosed into early and late endosomes. Uncoating occurs and the viral membrane fuses to the endosomal membrane, which allows viral RNP to enter the cytoplasm and nucleus. Transcription of viral RNA occurs, and translation of viral protein occurs through host machinery. Nuclear export of viral RNA and budding then occur. ( a ) ANXA1 is shown in this study to enhance binding of the virus to the host cell, and increases endocytosis of the virus. ( b ) ANXA1 is found in early and late endosomes near the nucleus, and silencing of ANXA1 results in lower nuclear infection. ( c ) More virus replication is observed in ANXA1 expressing cells. ( d ) In addition, IAV induces AKT phosphorylation to activate NF- k B and IKB phosphorylation and degradation. cIAP2 is enhanced by IAV, which is AKT dependent. ( e ) ANXA1 is shown to inhibit AKT activation and NF- k B activity. This may result in increased apoptosis. ( f ) ANXA1 promotes the release of cytochrome C from the mitochondria, enhances caspase 3/7 activation and PARP cleavage. ( g ) This leads to apoptosis and enhanced virus replication

Article Snippet: Briefly, Nunc Maxisorb (NUNC) 96 well plates were coated with 50 μ l of 1 μ g/ml goat anti-ANXA1 antibody (Santa Cruz) in PBS overnight at 4 °C.

Techniques: Membrane, Binding Assay, Virus, Infection, Expressing, Phospho-proteomics, Activation Assay, Activity Assay