Anti-CD133 antibody - Stem Cell Marker

• WB

Lab

Western blot - Anti-CD133 antibody - Stem Cell Marker (AB19898)

Lanes 1 - 2 : Merged signal (red and green). Green - ab19898 observed at 97 kDa. Red - loading control, ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab19898 was shown to react with CD133 in western blot. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab19898 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at 1 μg/ml and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-CD133 antibody - Stem Cell Marker (ab19898) at 1 µg/mL

Lane 1:

Caco 2 cell lysate at 20 µg

Lane 2:

HeLa cell lysate at 20 µg

Predicted band size: 97 kDa

Observed band size: 97 kDa

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• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-CD133 antibody - Stem Cell Marker (AB19898)

ab19898 staining CD133 in Caco-2 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 ab19898 at 1µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour magenta). Nuclear DNA was labelled with DAPI (shown in blue). Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

• IP

Lab

Immunoprecipitation - Anti-CD133 antibody - Stem Cell Marker (AB19898)

Immunoprecipitation side-by-side comparison with the recombinant multiclonal antibody ab278053
This immunoprecipitation image is a comparison between ab19898 and the alternative recombinant multiclonal antibody ab278053.
Left side - Recombinant multiclonal to CD133 - ab278053
CD133 was immunoprecipitated from 0.35 mg Caco-2 (Human colorectal adenocarcinoma epithelial cell) whole cell lysate with ab278053 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab278053 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.
Lane 1 : Caco-2 whole cell lysate 10 μg.
Lane 2 : ab278053 IP in Caco-2 whole cell lysate.
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab278053 in Caco-2 whole cell lysate.
Blocking and dilution buffer and concentration : 5% NFDM/TBST.
Exposure time : 3 seconds.
Right side - Polyclonal antibody to [CD133] - ab19898
Same testing conditions as ab278053.
Why choose a recombinant antibody?
Research with confidence - consistent and reproducible results with every batch
Long-term and scalable supply - powered by recombinant technology for fast production
Success from the first experiment - confirmed specificity through extensive validation
Ethical standards compliant - production is animal-free

All lanes:

Immunoprecipitation - Anti-CD133 antibody - Stem Cell Marker (ab19898)

Predicted band size: 97 kDa

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• WB

Project

Western blot - Anti-CD133 antibody - Stem Cell Marker (AB19898)

ab19898 contains a number of potential glycosylation sites (SwissProt) which may explain its migration at a higher molecular weight than predicted.

All lanes:

Western blot - Anti-CD133 antibody - Stem Cell Marker (ab19898) at 1 µg/mL

Lane 1:

MEL-1 (Human embryonic stem cell, male cell line) Whole Cell Lysate (<a href='/en-us/products/unavailable/mel-1-human-embryonic-stem-cell-male-cell-line-whole-cell-lysate-ab27198'>ab27198</a>) at 10 µg

Lane 2:

Western blot - HT-29 whole cell lysate (<a href='/en-us/products/cell-lysates/ht-29-whole-cell-lysate-ab3952'>ab3952</a>) at 10 µg

Lane 3:

Caco-2 whole cell lysate (<a href='/en-us/products/unavailable/caco-2-whole-cell-lysate-ab3950'>ab3950</a>) at 10 µg

Secondary

All lanes:

Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

Predicted band size: 97 kDa

Observed band size: 110 kDa

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• WB

Project

Western blot - Anti-CD133 antibody - Stem Cell Marker (AB19898)

Anti-CD133 antibody ab19898 detects a band corresponding to the expected size of CD133 in Human Embryonic Stem Cell Lysate using Western Blotting. It is likely that the band runs higher than the predicted MW of CD133 due to glycosylation of CD133. In mouse neural stem cell and mouse embryonic stem cell lysate this ~110 kDa band was not detected but a reproducible band of ~30 kDa was seen. This could represent a cleavage product but it may also be that the ~30 kDa band represents non-specific binding by ab19898. However, ab19898 has been shown to recognise mouse neural stem cells using ICC. Blocking using the immunising peptide removed both of these bands.

All lanes:

Western blot - Anti-CD133 antibody - Stem Cell Marker (ab19898) at 1 µg/mL

Lane 1:

Human Embryonic Stem Cell Lysate at 20 µg

Lane 2:

Human Embryonic Stem Cell Lysate at 20 µg with CD133 peptide (<a href='/en-us/products/unavailable/cd133-peptide-ab20651'>ab20651</a>)

Predicted band size: 97 kDa

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• WB

Lab

Western blot - Anti-CD133 antibody - Stem Cell Marker (AB19898)

Western blot side-by-side comparison with the recombinant multiclonal antibody ab278053
This Western blot image is a comparison between ab19898 and the alternative recombinant multiclonal antibody ab278053.
Left side - Recombinant multiclonal to CD133 - ab278053
All lanes : Anti-CD133 antibody [RM1002] (ab278053) at 1/1000 dilution
Lane 1 : Caco-2 (Human colorectal adenocarcinoma epithelial cell) whole cell lysate
Lane 2 : HEK-293T (Human embryonic kidney epithelial cell) whole cell lysate
Lane 3 : NCCIT (Human pluripotent embryonic carcinoma epithelial cell) whole cell lysate
Lane 4 : HT-29 (Human colorectal adenocarcinoma epithelial cell) whole cell lysate
Lysates/proteins at 20 μg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size : 97 kDa
Observed band size : 120 kDa
Blocking and dilution buffer and concentration : 5% NFDM/TBST.
Exposure time : 26 seconds.
Right side - Polyclonal antibody to [CD133] - ab19898
Same testing conditions as ab278053.
Why choose a recombinant antibody?
Research with confidence - consistent and reproducible results with every batch
Long-term and scalable supply - powered by recombinant technology for fast production
Success from the first experiment - confirmed specificity through extensive validation
Ethical standards compliant - production is animal-free

All lanes:

Western blot - Anti-CD133 antibody - Stem Cell Marker (ab19898)

Predicted band size: 97 kDa

false