Human L1CAM knockout HeLa cell line
- Advanced Validation
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- ICC/IF
Supplier Data
Immunocytochemistry/ Immunofluorescence - Human L1CAM knockout HeLa cell line (AB255401)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized L1CAM KO HeLa (L1CAM knockout human cervical adenocarcinoma epithelial cell) compared to wild-type HeLa cells labeling L1CAM with ab307859 at 1/50 dilution (10.0 ug/ml). Confocal image showing strong membranous and weak cytoplasmic staining in wild-type HeLa cells, while no staining in L1CAM knockout HeLa cells (ab255401). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). ab195887 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 488) was used to counterstain tubulin at 1/200 dilution (2.5 ug/ml) (Red). The nuclear counterstain was DAPI (Blue).
- WB
Unknown
Western blot - Human L1CAM knockout HeLa cell line (AB255401)
Lanes 1-2 : Merged signal (red and green). Green - ab182407 observed at 220 kDa. Red - loading control ab130007 observed at 125 kDa.
ab182407 was shown to react with L1CAM in wild-type HeLa cells in Western blot. Loss of signal was observed when knockout sample ab263786 was used. Wild-type and L1CAM knockout samples were subjected to SDS-PAGE. ab182407 and Anti-Vinculin antibody [VIN-54] (ab130007) were incubated overnight at 4° at 1 in 5000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-L1CAM antibody [EPR18998] (<a href='/en-us/products/primary-antibodies/l1cam-antibody-epr18998-ab182407'>ab182407</a>) at 1/5000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
L1CAM knockout HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human L1CAM knockout HeLa cell line (ab255401)
Secondary
All lanes:
Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution
Predicted band size: 140 kDa
Observed band size: 124 kDa,220 kDa
false
- WB
Unknown
Western blot - Human L1CAM knockout HeLa cell line (AB255401)
Lanes 1-2 : Merged signal (red and green). Green - ab208155 observed at 220 kDa. Red - loading control ab130007 observed at 125 kDa.
ab208155 was shown to react with L1CAM in wild-type HeLa cells in Western blot. Loss of signal was observed when knockout sample ab263786 was used. Wild-type and L1CAM knockout samples were subjected to SDS-PAGE. ab208155 and Anti-Vinculin antibody [VIN-54] (ab130007) were incubated overnight at 4° at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-L1CAM antibody [EPR18750] (<a href='/en-us/products/primary-antibodies/l1cam-antibody-epr18750-ab208155'>ab208155</a>) at 1/1000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
L1CAM knockout HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human L1CAM knockout HeLa cell line (ab255401)
Secondary
All lanes:
Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution
Predicted band size: 140 kDa
Observed band size: 124 kDa,220 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human L1CAM knockout HeLa cell line (AB255401)
Homozygous : 154 bp insertion in exon1
- ICC/IF
Supplier Data
Immunocytochemistry/ Immunofluorescence - Human L1CAM knockout HeLa cell line (AB255401)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized L1CAM KO HeLa (L1CAM knockout human cervical adenocarcinoma epithelial cell) cells labelling L1CAM with ab315360 at 1/50 (10.0 ug/ml) dilution (Red).
Confocal image showing strong membranous and weak cytoplasmic staining in wild-type HeLa cells, while no staining in L1CAM knockout HeLa cells (ab255401). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
ab195887 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 488) was used to counterstain tubulin at 1/200 (2.5ug/ml) dilution (Green). The Nuclear counterstain was DAPI (Blue).
- ICC/IF
Supplier Data
Immunocytochemistry/ Immunofluorescence - Human L1CAM knockout HeLa cell line (AB255401)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized L1CAM KO HeLa (L1CAM knockout human cervical adenocarcinoma epithelial cell) cells labelling L1CAM with ab315361 at 1/50 (10.0 ug/ml) dilution (Red).
Confocal image showing strong membranous and weak cytoplasmic staining in wild-type HeLa cells, while no staining in L1CAM knockout HeLa cells (ab255401). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
ab195887 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 488) was used to counterstain tubulin at 1/200 (2.5ug/ml) dilution (Green). The Nuclear counterstain was DAPI (Blue).
- WB
Lab
Western blot - Human L1CAM knockout HeLa cell line (AB255401)
Blocking/Diluting buffer and concentration : 5% NFDM/TBST.
ab270455 was shown to specifically react with L1CAM in wild-type HeLa cells. Loss of signal was observed when knockout sample from Human L1CAM knockout HeLa cell line (ab255401) was used. Wild-type and L1CAM knockout samples were subjected to SDS-PAGE. ab270455 and ab181602 (Rabbit anti-GAPDH loading control) were incubated 1 hour at room temperature at 1/1000 dilution and 1/200,000 dilution respectively. Blots were developed with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) secondary antibody at 1/100,000 dilution for 1 hour at room temperature before imaging. The blot was developed on a BIO-RAD® ChemiDoc™ MP instrument using the ECL technique.
A549 is negative or very low expression cell line for L1CAM (PMID : 23511563).
All lanes:
Western blot - Anti-L1CAM antibody [EPR23241-224] (<a href='/en-us/products/primary-antibodies/l1cam-antibody-epr23241-224-ab270455'>ab270455</a>) at 1/1000 dilution
Lane 1:
Wild-type HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg
Lane 2:
Western blot - Human L1CAM knockout HeLa cell line (ab255401) at 20 µg
Lane 3:
A375 (human malignant melanoma epithelial cell), whole cell lysate at 20 µg
Lane 4:
A549 (human lung carcinoma epithelial cell), whole cell lysate at 20 µg
Secondary
All lanes:
Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution
Predicted band size: 140 kDa
Observed band size: 250 kDa
false
Exposure time: 3min
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Advanced Validation
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Reactivity data
Product details
We will provide viable cells that proliferate on revival.
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.
What's included?
Properties and storage information
Gene name
Gene editing type
Gene editing method
Knockout validation
Zygosity
Shipped at conditions
Appropriate short-term storage conditions
Appropriate long-term storage conditions
Handling procedures
Initial handling guidelines
Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.
1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method seed all remaining cells into a T25.
4. Incubate the culture at 37°C incubator with 5% CO2. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.
5. Once confluent passage into an appropriate flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.
Subculture guidelines
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x104 cells/cm2 is recommended.
- Cells should be passaged when they have achieved 80-90% confluence.
Culture medium
DMEM (High Glucose) + 10% FBS
Cryopreservation medium
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
L1CAM facilitates neuronal migration axonal growth and synaptic plasticity. This protein participates in the formation and maintenance of neural networks. It interacts with other cell adhesion molecules and components of the extracellular matrix. L1CAM acts as an important modulator within neuronal signaling complexes which impacts the development and regeneration of the nervous system. Insights into its structural and cell interaction properties have made it a focus for understanding nervous system functions.
Pathways
L1CAM participates extensively in the MAPK and PI3K/AKT pathways. These pathways contribute to cellular growth survival and programmed cell death. The protein interacts with other molecules such as integrins and FGFRs to propagate signaling cascades important for cellular response mechanisms. These connections help modulate cytoskeletal dynamics influencing cellular adhesion and motility needed during brain development and response to injury.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
Target data
Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
For licensing inquiries, please contact partnerships@abcam.com