Human FHL2 knockout U-2 OS cell line
- Advanced Validation
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FHL2 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 CRISPR/Cas9 X = 1 bp insertion Frameshift = 96.21%. To order both knockout and wild-type control cells: select '2 x 1000000 Cells/vial'. To order only knockout cells: select '1000000 Cells/vial'.
View Alternative Names
AAG 11, Aging associated gene 11, DRAL, Down regulated in rhabdomyosarcoma LIM protein, FHL2 protein, FHL2_HUMAN, Four and a half LIM domain protein 2, Four and a half LIM domains 2, Four and a half LIM domains protein 2, KIAA0990, LIM domain protein DRAL, SLIM-3, Skeletal muscle LIM-protein 3
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - Human FHL2 knockout U-2 OS cell line (AB262496)
ab202584 staining FHL2 in wild-type U-2 OS cells (top panel) and FHL2 knockout U-2 OS cells (ab262496) (bottom panel). The cells were fixed with 100% methanol (5 min), 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 ab202584 at 1/200 dilution and ab7291 (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 pseudo color red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - Human FHL2 knockout U-2 OS cell line (AB262496)
ab202584 staining FHL2 in wild-type U-2 OS cells (top panel) and FHL2 knockout U-2 OS cells (ab262496) (bottom panel). The cells were fixed with 4% PFA (10 min), 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 ab202584 at 1/200 dilution and ab7291 (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 pseudo color 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 FHL2 knockout U-2 OS cell line (AB262496)
Lanes 1 - 3 : Merged signal (red and green). Green - ab202584 observed at 32 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A] observed at 55kDa.
ab202584 was shown to react with FHL2 in wild-type U-2 OS cells in western blot with loss of signal observed in FHL2 knockout sample.Wild-type and FHL2 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 ab202584 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-FHL2 antibody [EPR17860-20] (<a href='/en-us/products/primary-antibodies/fhl2-antibody-epr17860-20-ab202584'>ab202584</a>) at 1/1000 dilution
Lane 1:
Wild-type U-2 OS cell lysate at 40 µg
Lane 2:
FHL2 knockout U-2 OS cell lysate at 40 µg
Lane 2:
Western blot - Human FHL2 knockout U-2 OS cell line (ab262496)
Lane 3:
HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 40 µg
Predicted band size: 32 kDa
Observed band size: 32 kDa
false
- WB
Lab
Western blot - Human FHL2 knockout U-2 OS cell line (AB262496)
Lanes 1 - 3 : Merged signal (red and green). Green - ab202586 observed at 32 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A] observed at 55kDa.
ab202586 was shown to react with FHL2 in wild-type U-2 OS cells in western blot with loss of signal observed in FHL2 knockout sample. Wild-type and FHL2 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 ab202586 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-FHL2 antibody [EPR17860-23] (<a href='/en-us/products/primary-antibodies/fhl2-antibody-epr17860-23-ab202586'>ab202586</a>) at 1/1000 dilution
Lane 1:
Wild-type U-2 OS cell lysate at 40 µg
Lane 2:
FHL2 knockout U-2 OS cell lysate at 40 µg
Lane 2:
Western blot - Human FHL2 knockout U-2 OS cell line (ab262496)
Lane 3:
HeLa cell lysate at 40 µg
Predicted band size: 32 kDa
Observed band size: 32 kDa
false
- NGS
Supplier Data
Next Generation Sequencing - Human FHL2 knockout U-2 OS cell line (AB262496)
Knockout achieved by CRISPR/Cas9; X = 1 bp insertion; Frameshift = 96.21%
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
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
McCoY5a + 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
The FHL2 protein serves critical functions in muscle development and differentiation. It operates as part of a multiprotein complex that interacts with cytoskeletal components contributing importantly to muscle cell adhesion and communication. Moreover FHL2 engages in transcriptional regulation and acts as a coactivator or corepressor of specific transcription factors influencing gene expression patterns relevant to muscle function and maintenance.
Pathways
The FHL2 protein plays significant roles in Wnt and MAPK signaling pathways. In the Wnt signaling pathway FHL2 regulates the transcriptional activity of beta-catenin an essential component of the pathway involved in cell growth and differentiation. Additionally FHL2 contributes to the MAPK signaling pathway by influencing interactions between upstream regulators and downstream targets modulating cellular responses to external stimuli. Through these pathways FHL2 interacts with various proteins including GATA4 and TEF-1 which are vital for its regulatory functions.
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
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Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
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