Human ACTA2 knockout HeLa cell line
- Advanced Validation
- What is this?
Be the first to review this product! Submit a review
|
(1 Publication)
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - Human ACTA2 knockout HeLa cell line (AB264014)
ab8211 staining alpha smooth muscle Actin in wild-type HeLa cells (top panel) and ACTA2 knockout HeLa cells (ab264014) (bottom panel). 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 ab8211 at 5 ug/ml concentration and ab195884 (Rat monoclonal to Tubulin - Alexa Fluor® 647) at 1/100 dilution overnight at 4oC. 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 ACTA2 knockout HeLa cell line (AB264014)
ab202586 staining FHL2 in wild-type U-2 OS cells (top panel) and FHL2 knockout U-2 OS cells (ab264014) (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 ab202586 at 1/100 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
Unknown
Immunocytochemistry/ Immunofluorescence - Human ACTA2 knockout HeLa cell line (AB264014)
ab150301 staining alpha smooth muscle Actin in wild-type HeLa cells (top panel) and ACTA2 knockout HeLa cells (ab264014) (bottom panel). 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 ab150301 at 5 ug/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4oC followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 ug/ml (shown in green) and a goat secondary antibody to mouse IgG (AAlexa Fluor® 594) (ab150120) at 2 ug/ml (shown in 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 ACTA2 knockout HeLa cell line (AB264014)
ab202586 staining FHL2 in wild-type U-2 OS cells (top panel) and FHL2 knockout U-2 OS cells (ab264014) (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 ab202586 at 1/100 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
Unknown
Immunocytochemistry/ Immunofluorescence - Human ACTA2 knockout HeLa cell line (AB264014)
ab124964 staining alpha smooth muscle Actin in wild-type HeLa cells (top panel) and ACTA2 knockout HeLa cells (ab264014) (bottom panel). 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 ab124964 at 1/500 dilution and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4oC followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 ug/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 ug/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 ACTA2 knockout HeLa cell line (AB264014)
Lanes 1 - 2 : Merged signal (red and green). Green - ab124964 observed at 42 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55kDa.
ab124964 was shown to react with alpha smooth muscle Actin in wild-type HeLa cells in western blot. Loss of signal was observed when ACTA2 knockout sample was used. Wild-type HeLa and ACTA2 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab124964 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4° at a 1 in 10000 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-alpha smooth muscle Actin antibody [EPR5368] (<a href='/en-us/products/primary-antibodies/alpha-smooth-muscle-actin-antibody-epr5368-ab124964'>ab124964</a>) at 1/10000 dilution
Lane 1:
Wild-type HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 2:
ACTA2 knockout HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 2:
Western blot - Human ACTA2 knockout HeLa cell line (ab264014)
Predicted band size: 42 kDa
Observed band size: 42 kDa
false
- WB
Lab
Western blot - Human ACTA2 knockout HeLa cell line (AB264014)
ab203696 was shown to react with alpha smooth muscle Actin (HRP) in wild-type HeLa cells in western blot. Loss of signal was observed when ACTA2 knockout cell line ab264014 (knockout cell lysate ab264499) was used. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab203696 overnight at 4° at a 1 in 5000 dilution and ab184095 (Mouse Anti-GAPDH antibody [mabcam 9484] - Alexa Fluor® 680) at a 1 in 1000 dilution. Blots were developed with Optiblot ECL reagent (ab133456) and imaged.
All lanes:
Western blot - HRP Anti-alpha smooth muscle Actin antibody [1A4] (<a href='/en-us/products/primary-antibodies/hrp-alpha-smooth-muscle-actin-antibody-1a4-ab203696'>ab203696</a>) at 1/5000 dilution
Lane 1:
Wild-type HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 2:
ACTA2 knockout HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Predicted band size: 42 kDa
Observed band size: 42 kDa
false
Exposure time: 20s
Complementary Products
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-alpha smooth muscle Actin antibody [1A4] (AB7817)](https://content.abcam.com/products/images/alpha-smooth-muscle-actin-antibody-1a4-ab7817--immunohistochemistry-formalin-pfa-fixed-paraffin-embedded-sections-img436417.jpg)
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
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
Alpha smooth muscle actin aids in maintaining the structural integrity of tissues by forming part of the actin cytoskeleton an important element in cellular support. It exists in high concentration in stress fibers contributing to cellular movements and shape maintenance. These actions are essential in various dynamic cellular processes such as cell migration and adhesion. Alpha smooth muscle actin does not typically form complexes but it associates with other components of the actin cytoskeleton to ensure cell stability and function.
Pathways
Alpha smooth muscle actin functions significantly within the TGF-beta signaling pathway which influences cell proliferation differentiation and apoptosis. It interacts closely with proteins such as myosin to facilitate cellular contractility and motility. Additionally α-SMA plays a part in the RhoA/Rho kinase (ROCK) pathway connecting with regulators of actin filament organization. These pathways are essential for modulation of smooth muscle contraction and actin filament assembly contributing to vascular development and wound healing.
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
Publications (1)
Recent publications for all applications. Explore the full list and refine your search
Frontiers in immunology 13:1046121 PubMed36483553
2022
Applications
Unspecified application
Species
Unspecified reactive species
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