Human RARA (Retinoic Acid Receptor alpha) knockout HeLa cell line
- Advanced Validation
- What is this?
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- WB
Lab
Western blot - Human RARA (Retinoic Acid Receptor alpha) knockout HeLa cell line (AB265176)
Lanes 1 - 4 : Merged signal (red and green). Green - ab275745 observed at 40 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.
ab275745 was shown to react with Retinoic Acid Receptor alpha in wild-type HeLa cells in western blot. The bands observed in RARA knockout cell line ab265176 (RARA knockout cell lysate ab257629) below 40 kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Wild-type and RARA CRISPR/Cas9 HeLa edited cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab275745 and ab8245 (Mouse anti-GAPDH antibody [6C5]) 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 h at room temperature before imaging.
All lanes:
Western blot - Anti-Retinoic Acid Receptor alpha antibody [EPR23871-271] (<a href='/en-us/products/primary-antibodies/retinoic-acid-receptor-alpha-antibody-epr23871-271-ab275745'>ab275745</a>) at 1/1000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
RARA CRISPR-Cas9 edited HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human RARA (Retinoic Acid Receptor alpha) knockout HeLa cell line (ab265176)
Lane 3:
MCF7 cell lysate at 20 µg
Lane 4:
HepG2 cell lysate at 20 µg
Predicted band size: 51 kDa
Observed band size: 50-55 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human RARA (Retinoic Acid Receptor alpha) knockout HeLa cell line (AB265176)
Homozygous : 1 bp insertion in exon 7.
Complementary Products
Primary Antibodies
AB275745
Anti-Retinoic Acid Receptor alpha antibody [EPR23871-271]
BOND RX™ Validated|RabMAb|Recombinant|KO Validated|20ul selling size
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Retinoic Acid Receptor alpha antibody [EPR23871-271] (AB275745)](https://content.abcam.com/products/images/retinoic-acid-receptor-alpha-antibody-epr23871-271-ab275745--immunohistochemistry-formalin-pfa-fixed-paraffin-embedded-sections-img73455.jpg)
- 4
- 1
Primary Antibodies
AB125001
Anti-Retinoid X Receptor alpha/RXRA antibody [EPR7106]
RabMAb|Recombinant|KO Validated
![Immunocytochemistry/ Immunofluorescence - Anti-Retinoid X Receptor alpha/RXRA antibody [EPR7106] (AB125001)](https://content.abcam.com/products/images/retinoid-x-receptor-alpha-rxra-antibody-epr7106-ab125001--immunocytochemistry-immunofluorescence-img125288.jpg)
- 5
- 5
Reactivity data
Product details
We will provide viable cells that proliferate on revival.
Western blot data indicates that the CRISPR gene edit may have resulted in a truncation of the protein of interest. Please see data images.
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
RARA plays an essential role in mediating the effects of retinoic acid in the body. It is part of a larger receptor complex that interacts with co-regulators to modulate gene expression. This process is significant for embryonic development and the maintenance of normal physiological functions. Through its action RARA contributes to the proper development of organs and is critical for maintaining immune homeostasis and enabling the cellular response to environmental changes.
Pathways
RARA's activity impacts important signaling routes such as the retinoic acid signaling pathway and the Wnt signaling pathway. It collaborates with proteins like retinoid X receptors (RXRs) and other nuclear receptors to influence gene expression processes. These pathways maintain cellular differentiation and tissue homeostasis demonstrating RARA's integrative role in cellular signaling and communication.
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'.
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