Human CD40 knockout U-2 OS cell lysate (ab263923)
Overview
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Product name
Human CD40 knockout U-2 OS cell lysate
See all CD40 kits -
Product overview
Knockout cell lysate achieved by CRISPR/Cas9.
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Parental Cell Line
U-2 OS -
Organism
Human -
Mutation description
Knockout achieved by CRISPR/Cas9; X = 1 bp insertion, 2 bp insertion; Frameshift: 99.09% -
Passage number
<20 -
Knockout validation
Next Generation Sequencing (NGS), Western Blot (WB) -
Reconstitution notes
To use as WB control, resuspend the lyophilizate in 50 µL of LDS* Sample Buffer to have a final concentration of 2 mg/ml. For reducing conditions, we recommend a final concentration of 0.1 M DTT.*Usage of SDS sample buffer is not recommended with these lyophilized lysates.
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Notes
Lysate preparation: Our lysates are made using RIPA buffer to which we add a protease inhibitor cocktail and phosphatase inhibitor cocktail (ratio: 300:100:10). This means that the protein of interest is denatured. If you require a native form of the protein please use the live cell version - found here. Please refer to our lysis protocol for further details on how our lysates are prepared.
User storage instructions: Lyophilizate may be stored at 4°C. After reconstitution, store at -20°C for short-term storage or -80°C for long-term storage.
Access thousands of knockout cell lysates, generated from commonly used cancer cell lines.
See here for more information on knockout cell lysates.Abcam has not and does not intend to apply for the REACH Authorisation of customers’ uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.
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Tested applications
Suitable for: WBmore details
Properties
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Storage instructions
Store at -80°C. Please refer to protocols. -
Components 1 kit ab280476 - Human CD40 knockout U2OS cell lysate 1 x 100µg ab263974 - Human wild-type U-2 OS cell lysate 1 x 100µg -
Research areas
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Cell type
epithelial -
Disease
Osteosarcoma -
Gender
Female
Target
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Function
Receptor for TNFSF5/CD40LG. -
Tissue specificity
B-cells and in primary carcinomas. -
Involvement in disease
Defects in CD40 are the cause of hyper-IgM immunodeficiency syndrome type 3 (HIGM3) [MIM:606843]; also known as hyper-IgM syndrome 3. HIGM3 is an autosomal recessive disorder which includes an inability of B cells to undergo isotype switching, one of the final differentiation steps in the humoral immune system, an inability to mount an antibody-specific immune response, and a lack of germinal center formation. -
Sequence similarities
Contains 4 TNFR-Cys repeats. -
Cellular localization
Secreted and Cell membrane. - Information by UniProt
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Alternative names
- AI326936
- B cell associated molecule CD40
- B cell surface antigen CD40
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Associated products
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KO cell lines
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab263923 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Abreviews | Notes |
|---|---|---|
| WB |
Use at an assay dependent concentration. Predicted molecular weight: 30 kDa.
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| Notes |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 30 kDa. |
Images
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Western blot -Human CD40 knockout U-2 OS cell lysate
Lane 1: Wild-type U-2 OS Vehicle Control LPS (0μg/mL, 6h) cell lysate 20 μg
Lane 2: Wild-type U-2 OS Treated LPS (1μg/mL, 6h) cell lysate 20 μg
Lane 3: CD40 knockout U-2 OS Vehicle Control LPS (0μg/mL, 6h) cell lysate 20 μg
Lane 4: CD40 knockout U-2 OS Treated LPS (1μg/mL, 6h) cell lysate 20 μg
Lanes 1 - 4:Merged signal (red and green). Green - ab280207 observed at 45 kDa. Red - loading control ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) observed at 55kDa.
ab280207 was shown to react with CD40 in wild-type U-2 OS cells in Western blot with loss of signal observed in CD40 knockout cell line ab262486 (CD40 knockout cell lysate ab263923). Wild-type U-2 OS and CD40 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab280207 and ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 h t room temperature before imaging.
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Next Generation Sequencing - Human CD40 knockout U-2 OS cell lysate (ab263923)Knockout achieved by CRISPR/Cas9; X = 1 bp insertion, 2 bp insertion; Frameshift: 99.09% -
Western blot - Human CD40 knockout U-2 OS cell lysate (ab263923)Lane 1: Wild-type U-2 OS cell lysate
Lane 2: CD40 knockout U-2 OS cell lysate
Lane 3: Raji (Human Burkitt's lymphoma cell line) whole cell lysate
Lanes 1 - 3: Merged signal (red and green). Green - ab113701 observed at 45 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A] observed at 55kDa.
ab113701 was shown to react with CD40 in wild-type U-2 OS cells in Western blot Loss of signal was observed when CD40 knockout cell line ab262486 (knockout cell lysate ab263923) was used. Wild-type and CD40 knockout U-2 OS cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab113701 and ab7291 (Mouse anti-Alpha Tubulin [DM1A] 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. -
Western blot - Human CD40 knockout U-2 OS cell lysate (ab263923)Lane 1: Wild-type U-2 OS whole cell lysate
Lane 2: CD40 knockout U-2 OS whole cell lysate
Lane 3: Raji (Human Burkitt's lymphoma cell line) whole cell lysate
Lanes 1 - 3: Merged signal (red and green). Green - ab213205 observed at 42 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A] observed at 55kDa.
ab213205 was shown to react with CD40 in U-2 OS wild-type cells in Western blot Loss of signal was observed when CD40 knockout cell line ab262486 (knockout cell lysate ab263923) was used. Wild-type U-2 OS and CD40 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab213205 and ab7291 (Mouse anti-Alpha Tubulin [DM1A] overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. Blots were developed 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.
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab263923 has not yet been referenced specifically in any publications.