Human IL13RA2 knockout A375 cell line
- Advanced Validation
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IL13RA2 KO cell line available to order. KO validated by Immunocytochemistry, Next Generation Sequencing, Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp deletion in exon 3. To order both knockout and wild-type control cells: select 2 x 1000000Cells/vial. To order only knockout cells: select 1000000Cells/vial.
View Alternative Names
CD213a2, Cancer/testis antigen 19, I13R2_HUMAN, IL-13 receptor subunit alpha-2, IL-13R subunit alpha-2, IL-13R-alpha-2, IL-13RA2, IL13 receptor, IL13BP, IL13R, Interleukin 13 receptor alpha 2 chain, Interleukin-13 receptor subunit alpha-2, Interleukin-13-binding protein
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - Human IL13RA2 knockout A375 cell line (AB273381)
ab260044 staining IL-13 receptor alpha 2 in wild-type A375 cells (top panel) and IL13RA2 knockout A375 cells (bottom panel) (ab273381). The cells were fixed with 100% methanol (5 min) then permeabilized with 0.1% Triton X-100 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 with ab260044 at 1/500 dilution and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).
- WB
Lab
Western blot - Human IL13RA2 knockout A375 cell line (AB273381)
Lanes 1 - 4 : Merged signal (red and green). Green - ab260044 observed at 49 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55kDa.
ab260044 was shown to react with IL-13 receptor alpha 2 in wild-type A375 cells in western blot with loss of signal observed in IL13RA2 knockout cell line ab273371 (knockout cell lysate ab275532). Wild-type and IL13RA2 knockout A375 cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab260044 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4° at a 1 in 1000 dilution 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-IL-13 receptor alpha 2 antibody [EPR22978-163] (<a href='/en-us/products/primary-antibodies/il-13-receptor-alpha-2-antibody-epr22978-163-ab260044'>ab260044</a>) at 1/1000 dilution
Lane 1:
Wild-type A375 cell lysate at 30 µg
Lane 2:
IL13RA2 knockout A375 cell lysate at 30 µg
Lane 2:
Western blot - Human IL13RA2 knockout A375 cell line (ab273381)
Lane 3:
U-87-MG cell lysate at 30 µg
Lane 4:
Daudi cell lysate at 30 µg
Predicted band size: 44 kDa
Observed band size: 49 kDa
false
- NGS
Supplier Data
Next Generation Sequencing - Human IL13RA2 knockout A375 cell line (AB273381)
Homozygous : 1 bp deletion in exon 3
- ICC/IF
Supplier Data
Immunocytochemistry/ Immunofluorescence - Human IL13RA2 knockout A375 cell line (AB273381)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized IL13RA2 KO A375(IL13RA2 knockout human malignant melanoma epithelial cell), ab273381 cells labelling IL13RA2 with ab324598 at 1/1000 (0.505 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2ug/ml) dilution (Green).
Confocal image showing membranous and cytoplasmic staining in wiltype A375 cells (shown in green), showing no staining in IL13RA2 knockout A375 cells. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/1000 (1ug/ml) dilution (Magenta). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2ug/ml) dilution.
- IHC-P
Supplier Data
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Human IL13RA2 knockout A375 cell line (AB273381)
Immunohistochemical analysis of paraffin-embedded (A) Wild-type A375 (human malignant melanoma epithelial cell) cell pellet (B) IL13RA2 knockout A375 cell pellet tissue labeling IL13RA2 with ab324598 at 1/100 (5.05 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
Positive staining on (A) Wild-type A375 cell pellet, no staining on (B) IL13RA2 knockout A375 cell pellet. The primary antibody was incubated for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument
Counterstained with Hematoxylin.
Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins
- Flow Cyt
Supplier Data
Flow Cytometry - Human IL13RA2 knockout A375 cell line (AB273381)
Flow cytometric analysis of IL13RA2 KO A375 (IL13RA2 knockout human malignant melanoma epithelial cell) (Magenta) / A375 (Green) cells labelling IL13RA2 with ab324598 at 1/50 dilution (1ug) / Green and magenta compared with a Rabbit monoclonal IgG (ab172730) (Black) and Grey isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).
Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.
Gated on viable cells.
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 and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant 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.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Female
Target data
Complementary Products
Primary Antibodies
AB274448
Anti-IL-13 receptor alpha 2 antibody [EPR22978-127] - BSA and Azide free (Capture)
RabMAb|Recombinant
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Product promise
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