ATOX1 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 11 bp deletion in exon 1 and 5 bp deletion in exon 1.
ATOX1_HUMAN, ATX1 antioxidant protein 1 homolog, ATX1 antioxidant protein 1 homolog (yeast), Copper transport protein, Copper transport protein ATOX1, HAH1, MGC138453, MGC138455, Metal transport protein, Metal transport protein ATX1
ATOX1 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 11 bp deletion in exon 1 and 5 bp deletion in exon 1.
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.
ATOX1 also known as Antioxidant 1 Copper Chaperone is a small copper-binding protein with a molecular mass of approximately 7 kDa. It plays an essential mechanical role in the transport of copper ions within the cell. ATOX1 helps deliver copper to key enzymes involved in cellular processes. It is expressed in various tissue types including the liver and the brain facilitating copper homeostasis at a cellular level.
ATOX1 serves as a chaperone that transports copper ions to ATPase proteins like ATP7A and ATP7B in the secretory pathway. ATOX1 works individually rather than as part of a larger protein complex handling copper metabolism effectively. By facilitating the delivery of copper to essential enzymes ATOX1 assists in maintaining critical cellular functions such as oxidative stress response and energy production.
Copper ion transport involving ATOX1 plays an important role in the regulation of metal ion homeostasis pathways. ATOX1 interacts closely with ATP7A and ATP7B transporting copper ions necessary for redox reactions and enzyme activities in these pathways. ATOX1's role in copper regulation also aligns with antioxidant defense processes helping protect cells from damage due to reactive oxygen species.
Disruption in ATOX1 function can be linked to disorders such as Wilson's disease and Menkes disease both of which involve copper metabolism impairment. Wilson's disease involves the protein ATP7B where the absence of proper copper transport leads to toxicity. Menkes disease involves ATP7A mutations leading to a shortage of copper delivery to necessary enzymes. ATOX1's activity is important for delivering copper to these associated ATPases highlighting its importance in related pathologies.
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Lanes 1-4: Merged signal (red and green). Green - Anti-ATOX1 antibody [EPR10352] ab154179 observed at 7 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa.
Anti-ATOX1 antibody [EPR10352] ab154179 Anti-ATOX1 antibody [EPR10352] was shown to specifically react with ATOX1 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266651 (knockout cell lysate Human ATOX1 knockout HEK-293T cell lysate ab257849) was used. Wild-type and ATOX1 knockout samples were subjected to SDS-PAGE. Anti-ATOX1 antibody [EPR10352] ab154179 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4°C at 1 in 500 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (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-ATOX1 antibody [EPR10352] (Anti-ATOX1 antibody [EPR10352] ab154179) at 1/500 dilution
Lane 1: Wild-type HEK293T cell lysate at 20 µg
Lane 2: ATOX1 knockout HEK293T cell lysate at 20 µg
Lane 2: Western blot - Human ATOX1 knockout HEK-293T cell line (ab266651)
Lane 3: HeLa cell lysate at 20 µg
All lanes: Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) at 1/10000 dilution
Predicted band size: 36 kDa, 45 kDa, 62 kDa, 7 kDa
Observed band size: 48 kDa, 50 kDa, 7 kDa, 70 kDa
Representative images of ATOX1 knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using an EVOS M5000 microscope.
Allele-1: 11 bp deletion in exon 1
Allele-2: 5 bp deletion in exon 1.
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