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Full length protein corresponding to Rat CD90/ Thy1.
The affinity of the Fab' of MRC OX-7 for rat Thy-1 is 3 x 109m-1 and for mouse Thy-1.1 is 3 x 108m-1.
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Our Abpromise guarantee covers the use of ab225 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Flow Cyt||Use 0.1µg for 106 cells.|
|IP||Use at an assay dependent concentration.|
|WB||Use a concentration of 5 µg/ml. Detects a band of approximately 35-37 kDa (predicted molecular weight: 17 kDa).|
|IHC-P||Use at an assay dependent concentration. PubMed: 23616767|
|IHC-Fr||Use at an assay dependent concentration. PubMed: 16723538|
Overlay histogram showing PC12 cells stained with ab225 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab225, 0.1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1](ab91353, 1µg/1x106 cells) used under the same conditions. Unlabelled sample (blue line). Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive result in 80% methanol (5 min) fixed PC12 cells used under the same conditions.
Please note that Abcam do not have any data for use of this antibody on non-fixed cells. We welcome any customer feedback.
Immunocytochemistry/ Immunofluorescence analysis of horse adipose and bone marrow stem cells labeling CD90 / Thy1 with ab225 at 1/1000 dilution. Cells were fixed in paraformaldehyde and permeabilized with Triton x100 0.1%. The cells were blocked with 10% BSA for 30 minutes at 37°C, followed by staining with ab225 at 1/1000 for 12 hours in PBS+IGEPAL+BSA+10%NGS at 4°C. Goat F(ab')2 Anti-Mouse IgG - (Fab')2 (Biotin), pre-adsorbed (ab5886) was used as the secondary antibody at 1/400 dilution.
Immunohistochemistry (Frozen sections) analysis of rat spleen tissue labeling CD90 / Thy1 with ab225 at 1/100 dilution. The tissue was fixed in paraformaldehyde followed by blocking in 10% serum for 20 minutes. The tissue was then stained with ab225 at 1/100 in PBS for 6 hours at 4°C. A polyclonal donkey anti-mouse Alexa Fluor® 488 secondary antibody was used at 1/1000 dilution.
Immunocytochemistry/ Immunofluorescence analysis of rat sciatic nerve schwann cells and fibroblasts labeling CD90 / Thy1 with ab225 at 1/400 dilution. Cells were fixed in paraformaldehyde and permeabilized with 0.5% Triton X-100. The cells were blocked with 5% serum for 1 hour at 21°C, followed by staining with ab225 at 1/400 in 0.2% BSA for 1 hour at 37°C. A polyclonal goat anti-mouse Alexa Fluor® 546 secondary antibody was used at 1/500 dilution.
Rat CD90/Thy1 is N-glycosylated at three sites, giving rise to molecules with a range of molecular masses (25-37 kDa).
ab225 stained PC12 cells. The cells were 100% methanol fixed for 5 minutes at -20°C and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1hour at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab225 at 5µg/ml) overnight at +4°C. The secondary antibody (pseudo-colored green) was Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed (ab150117) used at a 1/1000 dilution for 1hour at room temperature. Alexa Fluor® 594 WGA was used to label plasma membranes (pseudo-colored red) at a 1/200 dilution for 1hour at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43µM for 1hour at room temperature.
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