Anti-CD90 / Thy1 抗体 [MRC OX-7] - Hematopoietic Stem Cell Marker

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Immunocytochemistry - Anti-CD90 / Thy1 antibody [MRC OX-7] - Hematopoietic Stem Cell Marker (AB225)

This image was kindly supplied as part of the review submitted by Nick Voilley. Rat retinal ganglion cell labelled with ab225 as a primary antibody and an anti-mouse F(ab)' 2 coupled to Alexa488 as a secondary antibody. The cell is approximately 15 micrometers in diameter. Only the cells labelled in green in the culture bear action potentials when stimulated. These three elements (reactivity to ab225, size and elecrophysiological parameters) clearly indicate the cell is a ganglion cell.

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AbReview12172****

Immunocytochemistry - Anti-CD90 / Thy1 antibody [MRC OX-7] - Hematopoietic Stem Cell Marker (AB225)

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.

This image is courtesy of an anonymous abreview.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-CD90 / Thy1 antibody [MRC OX-7] - Hematopoietic Stem Cell Marker (AB225)

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 22°C.

The secondary antibody used was DyLight^® 488 goat anti-mouse IgG (H+L) ab96879 at 1/500 dilution for 30 min at 22°C.

Isotype control antibody (black line, mouse IgG1 [B11/6], 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.

ab225 gave a positive result in 80% methanol (5 min) fixed PC12 cells used under the same conditions.

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Lab

Immunocytochemistry - Anti-CD90 / Thy1 antibody [MRC OX-7] - Hematopoietic Stem Cell Marker (AB225)

ab225 staining CD90 / Thy1 in NGF-differentiated PC-12 cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Tween for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab225 at 1µg/ml and ab6046, Rabbit polyclonal to beta Tubulin - Loading Control. Cells were then incubated with ab150121, Goat polyclonal Secondary Antibody to Mouse IgM - mu chain (Alexa Fluor^® 488) at 1/1000 dilution (shown in green) and ab150080, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor^® 594) at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).

Image was acquired with a confocal microscope (Leica-Microsystems TCS SP8) and a single confocal section is shown.

• WB

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Western blot - Anti-CD90 / Thy1 antibody [MRC OX-7] - Hematopoietic Stem Cell Marker (AB225)

Rat CD90/Thy1 is N-glycosylated at three sites, giving rise to molecules with a range of molecular masses (25-37 kDa).

All lanes:

Western blot - Anti-CD90 / Thy1 antibody [MRC OX-7] - Hematopoietic Stem Cell Marker (ab225) at 5 µg/mL

Lane 1:

Brain (Rat) Tissue Lysate at 20 µg

Lane 2:

PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Mouse IgG H&L (HRP) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-hrp-preadsorbed-ab97040'>ab97040</a>) at 1/10000 dilution

Predicted band size: 17 kDa

Observed band size: 35-37 kDa

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Exposure time: 20min

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AbReview28841****

Immunocytochemistry - Anti-CD90 / Thy1 antibody [MRC OX-7] - Hematopoietic Stem Cell Marker (AB225)

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.

This image is courtesy of an abreview submitted by Vega Villar, University of Leon.