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Our Abpromise guarantee covers the use of ab17147 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|IHC-R||Use at an assay dependent concentration.|
|WB||Use at an assay dependent concentration.|
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|IHC-P||1/25 - 1/50. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.|
Human peripheral blood lymphocytes stained with ab17147 (red line). Human whole blood was processed using a modified protocol based on Chow et al, 2005 (PMID: 16080188). In brief, human whole blood was fixed in 4% formaldehyde (methanol-free) for 10 min at 22°C. Red blood cells were then lyzed by the addition of Triton X-100 (final concentration - 0.1%) for 15 min at 37°C. For experimentation, cells were treated with 50% methanol (-20°C) for 15 min at 4°C. Cells were then incubated with the antibody (ab17147, 1/100 dilution) for 30 min at 4°C. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 4°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >30,000 total events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. Gating strategy - peripheral blood lymphocytes.
ab17147 staining CD8 in human nasal polyp and tonsil tissue sections by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with 10% formalin for 24 hours and a heat mediated antigen retrieval step was performed using EDTA pH 8.0. Samples were then blocked with 20% DMEM and 1% BSA in culture medium for 20 minutes at 21°C followed by incubation with the primary antibody at a 1/150 dilution, for 18 hours at 4°C. A biotinylated rabbit anti-mouse (Fab2) polyclonal was used as secondary antibody at a 1/400 dilution.An Avidin Biotin blocking step was used in addition to those already mentioned.
This image is courtesy of an anonymous Abreview
Blocked with 4% BSA for 2 hours at 20°C.
Incubated with the primary antibody for 30 minutes.
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"