I) For CD4+ T cell depletion, intraperitoneal injection of Ultra-LEAF™ Purified anti-mouse CD4 Antibody (10 μg/g, clone GK1.5, BioLegend, San Diego, CA, USA) or corresponding isotype antibody (Ultra-LEAF Purified Rat IgG2b, κ Isotype Ctrl Antibody, clone RTK4530, 10 μg/g, BioLegend) was performed in mice on day 15, 25, and 40 PMI. ii) For CXCR3 blockade, intraperitoneal injection of Ultra-LEAF™ Purified anti-mouse CD183 (CXCR3) Antibody (10 μg/g, clone CXCR3-173, BioLegend) or corresponding isotype antibody (Armenian Hamster IgG, clone: HTK888, BioLegend) was performed in mice every 7 days since day 15 PMI. iii) For systemic IFN-g blocking, mice were injected i.p. with 200 mg of anti-IFN-g antibody (InVivoMAb anti-mouse IFNγ, BioXcell, clone XMG1.2) every 3 days since day 15 PMI. An IgG1 isotype antibody (HRPN, BioXcell) was used to serve as control. iv) To block VCAM-1, a InVivoMAb anti-mouse VCAM-1 (10 mg/kg, BioXcell; corresponding isotype antibody: InVivoMAb rat IgG1 isotype control, clone HRPN, BioXcell) was administrated intravenously in MB-injected mice every 3 days since day 15 PMI. For retinal evaluation in abovementioned in vivo antibody intervention experiments, one retina from each recipient mouse was randomly selected and used for RGC density, while the other was for Iba1 and GFAP staining.
Armenian hamster igg
Manufactured by BioLegend
Sourced in United States
Armenian Hamster IgG is an immunoglobulin G (IgG) antibody purified from Armenian hamster serum. IgG is the most abundant antibody isotype in the serum and plays a crucial role in the adaptive immune response.
Automatically generated - may contain errors
Lab products found in correlation
Invivomab anti mouse ifnγ, by BioXCell (1 mentions)
Cxcr3 173, by BioLegend (1 mentions)
Invivomab, by BioXCell (1 mentions)
Tnf α, by Miltenyi Biotec (1 mentions)
Pe cy7 anti ifnγ, by BD (1 mentions)
Anti cd4 pacific blue, by BD (1 mentions)
Anti il 10 pe, by BD (1 mentions)
Anti mouse cd16 cd32 2.4g2, by BD (1 mentions)
Cytofix cytoperm, by BD (1 mentions)
Lsrii flow cytometer, by BD (1 mentions)
Percp cy5.5 anti il 17a, by BD (1 mentions)
Rat igg2b, by BioLegend (1 mentions)
Mouse igg1, by BioLegend (1 mentions)
Mouse igg2a, by BioLegend (1 mentions)
Accuri c6 flow cytometer and software, by BD (1 mentions)
Rat igg2a, by BioLegend (1 mentions)
Cx3cr1 sa011f11, by BioLegend (1 mentions)
Rat igg2a, by Thermo Fisher Scientific (1 mentions)
Facscanto 2, by BD (1 mentions)
Software version 7, by FlowJo (1 mentions)
7 protocols using armenian hamster igg
1
Immune Modulation for Retinal Protection
2
Modulation of Cardiomyocyte Signaling Pathways
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In some experiments, cardiomyocytes were preincubated for 30 min with different pharmacological inhibitors: L-NAME (100 µM, NOS inhibitor), 1H-[1.2.4]oxadiazolo[4.3-a]quinoxalin-1-one [ODQ, 25 µM, soluble guanylate cyclase (sGC) inhibitor], 6,7-dimethyl-3-[(methyl{2-[methyl({1-[3-(trifluoromethyl)phenyl]-1H-indol-3-yl}methyl)amino]ethyl}amino)methyl]-4H-chromen-4-one,-diHCl (SPD 304, 10 µM, TNF-inhibitor, Calbiochem, Darmstadt, Germany). A specific antibody to TNFR1 (purified anti-mouse TNFR type 1/p55, 20 µg/mL, BioLegend, San Diego, CA), a leaf purified Armenian hamster IgG (20 µg/mL, BioLegend) or TNF-α (50 ng/mL, Miltenyi Biotec, Bisley, UK) were also used. All drugs were prepared as a stock solution in appropriate solvents. All the drugs were used at already reported maximally active concentrations. On the day of the experiment, stock solutions were diluted to the desired concentration and the final concentration of DMSO was <0.1%. For the treatment with MPs (4 h) of cardiomyocytes further used in western blot, M199 culture medium was supplemented with creatine (5 mM), carnitine (5 mM), taurine (5 mM), HEPES (25 mM) and penicillin/streptomycin (1%). Unless stated, all chemicals were obtained from Sigma-Aldrich (St. Louis, MO).
3
Flow Cytometry Analysis of T Cell Subsets
4
Exosome Surface Marker Characterization
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Antibodies to CD9 (HI9a), CD63 (NVG-2), CD63 (H5C6), CD81 (Eat-2), CCR9 (9B1), CXCR4 (L276F12), and CX3CR1 (SA011F11) were purchased from BioLegend (San Diego, CA). Isotype controls including Rat IgG2a, Rat IgG2b, Armenian Hamster IgG, Mouse IgG2a, and Mouse IgG1 were also from BioLegend. The antibody to CD9 (KMC8) was obtained from BD Biosciences. The antibodies to CCR7 (4B12), CCR10 (248918) were acquired from R&D Systems. The cells or microbead-conjugated exosomes were stained with the fluorescently labeled antibodies, washed twice with PBS containing 2% FBS and 2 mM ethylenediaminetetraacetic acid (EDTA) (Wako, Osaka, Japan), and analyzed by using BD Accuri C6 flow cytometer and software (BD Biosciences). For this method of microbead conjugation of exosomes, only positive events can be detected and fluorescently quantified in the flow cytometry. In some experiments, total (intracellular plus surface) staining experiment was done by using FIX and PERM Kit (Thermo Fisher Scientific) according to manufacturer's instructions.
5
Multiparameter Flow Cytometry Analysis
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Cell-surface and intracellular staining procedures were performed as previously described (31 (link)). Surfaces of 100 μl of cells (1×106) were stained using 10 μl fluorescein isothiocyanate-conjugated anti-CD25 and peridinin chlorophyll-conjugated anti-CD4 (eBiosciences, San Jose, CA, USA). Isotype control, mouse IgG1, was included in all the experiments. For intracellular staining, cells were saponized, washed in cold flow cytometry staining buffer and stained with phycoerythrin-conjugated anti-human Foxp3, its isotype control rat IgG2a (eBiosciences), Adenomatous polyposis coli anti-mouse/human Helios, and its isotype control Armenian Hamster IgG (BioLegend, Inc., San Diego, CA, USA). Flow cytometry was performed using FACSCanto II (BD Biosciences). Acquisition and analysis gates were restricted to the lymphocyte gate, as determined by their characteristic forward and side scatter properties. Flow data were analyzed using FlowJo software, version 7.6.5 (FlowJo, LLC, Ashland, OR, USA).
6
T cell activation by intestinal epithelial cells
7
Apoptosis Assay of Cell Lines
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B16, MB49, and EL4 cells were seeded in a six-well plate until reaching ~90% of confluency in supplemented RPMI. Cells were treated with 1 and 10 ng/ml of anti-CD29 for 4 h. Armenian hamster IgG (Biolegend, San Diego, California, USA) was used as control. After 4 h, cells were collected and washed twice with cold PBS and then resuspended in 1× binding buffer (Biolegend, USA) at 106 cells/ml. One hundred microliters of the cell suspension was transferred onto a 5-ml culture tube. Five microliters of Annexin-v APC and 7-AAD was added per sample. The cells were gently vortexed and incubated for 20 min at room temperature. One hundred microliters of the binding buffer was added to each tube, and the samples were analyzed by fluorescence-activated cell sorting (FACS).
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