FHL2 KO cell line available to order. KO validated by Immunocytochemistry, Next Generation Sequencing, Western blot. Free of charge wild type control provided. Knockout achieved by CRISPR/Cas9 X = 1 bp insertion Frameshift = 96.21%.
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
FHL2 KO cell line available to order. KO validated by Immunocytochemistry, Next Generation Sequencing, Western blot. Free of charge wild type control provided. Knockout achieved by CRISPR/Cas9 X = 1 bp insertion Frameshift = 96.21%.
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.
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.
The FHL2 protein also known as Four and a Half LIM Domains Protein 2 plays an important role in cellular mechanics. Its molecular mass is approximately 32 kDa. FHL2 is expressed in various tissues with significant presence in cardiac and skeletal muscles as well as the liver. It is involved in the regulation of gene expression by binding to other proteins and DNA through its LIM domains. This ability allows FHL2 to influence cellular structures and functions integrally.
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.
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.
FHL2 has associations with certain cancers like prostate cancer and cardiomyopathy. In prostate cancer FHL2 overexpression leads to increased tumor progression and is often linked to androgen receptors promoting malignant transformation. In cardiomyopathy alterations in FHL2 expression affect heart muscle integrity and function sometimes in conjunction with interactions involving desmin an important structural protein. Understanding FHL2's connections to these diseases offers insights into potential therapeutic targets.
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Anti-FHL2 antibody [EPR17860-23] 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 Anti-FHL2 antibody [EPR17860-23] ab202586 and Anti-alpha Tubulin antibody [DM1A] - Loading Control 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 (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (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-FHL2 antibody [EPR17860-23] (Anti-FHL2 antibody [EPR17860-23] ab202586) 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
Performed under reducing conditions.
Predicted band size: 32 kDa
Observed band size: 32 kDa
Knockout achieved by CRISPR/Cas9; X = 1 bp insertion; Frameshift = 96.21%
Anti-FHL2 antibody [EPR17860-20] 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 Anti-FHL2 antibody [EPR17860-20] ab202584 at 1/200 dilution and Anti-alpha Tubulin antibody [DM1A] - Loading Control 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) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed 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).
Anti-FHL2 antibody [EPR17860-20] 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 Anti-FHL2 antibody [EPR17860-20] ab202584 at 1/200 dilution and Anti-alpha Tubulin antibody [DM1A] - Loading Control 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) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed 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).
Anti-FHL2 antibody [EPR17860-20] 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 Anti-FHL2 antibody [EPR17860-20] ab202584 and Anti-alpha Tubulin antibody [DM1A] - Loading Control 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 (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (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-FHL2 antibody [EPR17860-20] (Anti-FHL2 antibody [EPR17860-20] ab202584) 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
Performed under reducing conditions.
Predicted band size: 32 kDa
Observed band size: 32 kDa
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