CANX KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 19 bp deletion in exon 2.
CALX_HUMAN, CANX, CNX, Calnexin, FLJ26570, Histocompatibility complex class I antigen binding protein p88, IP90, Major histocompatibility complex class I antigen-binding protein p88, p90
CANX KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 19 bp deletion in exon 2.
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.
Calnexin also known as Canx is a type I integral membrane protein of the endoplasmic reticulum (ER) involved in the process of protein folding. This chaperone protein has an approximate molecular weight of 90 kDa and is known for its role in the quality control of glycoproteins. Calnexin is expressed in the ER of cells where it interacts with nascent polypeptides to ensure proper folding and assembly contributing to cellular homeostasis. It exhibits its function through its lectin-like domain that binds to sugar moieties on glycoproteins.
Calnexin facilitates the proper folding of newly synthesized proteins by forming a complex with another chaperone protein called ERp57. This interaction helps in creating the correct disulfide bonds in glycoproteins which is essential for their stability and functionality. The complex often referred to as the calnexin cycle is critical in preventing the aggregation and misfolding of proteins within the ER. This process ensures that only correctly folded proteins proceed to the Golgi apparatus for further processing and transport.
Calnexin plays an important role in the ER-associated degradation (ERAD) pathway and the unfolded protein response (UPR). In these pathways calnexin ensures that misfolded proteins are retained in the ER or targeted for degradation preventing cellular stress. Calnexin is associated with proteins such as calreticulin another chaperone protein with a similar function in the ER. Together they maintain proteostasis within cells and protect against the accumulation of improperly folded proteins.
Calnexin is linked to several conditions including cystic fibrosis and certain neurodegenerative diseases. In cystic fibrosis the misfolding and subsequent degradation of the CFTR protein are associated with calnexin's role in the ERAD pathway. Similarly in neurodegenerative diseases such as Alzheimer's disrupted protein folding and aggregation are linked to ER stress where calnexin and other chaperone proteins like BiP play a pivotal role in managing protein misfolding. Understanding calnexin's role in these disorders can contribute to developing strategies to mitigate faulty protein folding and its pathological consequences.
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Lanes 1- 2: Merged signal (red and green). Green - Anti-Calnexin antibody [EPR3633(2)] - ER Membrane Marker ab133615 observed at 90 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) observed at 37 kDa.
Anti-Calnexin antibody [EPR3633(2)] - ER Membrane Marker ab133615 was shown to react with CANX in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab255368 (knockout cell lysate Human CANX (Calnexin) knockout HEK-293T cell lysate ab263805) was used. Wild-type HEK-293T and CANX knockout HEK-293T cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. Anti-Calnexin antibody [EPR3633(2)] - ER Membrane Marker ab133615 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) overnight at 4°C at a 1 in 1000 dilution and a 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-Calnexin antibody [EPR3633(2)] - ER Membrane Marker (Anti-Calnexin antibody [EPR3633(2)] - ER Membrane Marker ab133615) at 1/1000 dilution
Lane 1: Wild-type HEK-293T cell lysate at 20 µg
Lane 2: CANX knockout HEK-293T cell lysate at 20 µg
Lane 2: Western blot - Human CANX (Calnexin) knockout HEK-293T cell line (ab255368)
Performed under reducing conditions.
Predicted band size: 68 kDa
Observed band size: 90 kDa
Lanes 1 - 4: Merged signal (red and green). Green - Anti-Calnexin antibody [EPR3632] ab92573 observed at 80 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.
Anti-Calnexin antibody [EPR3632] ab92573 was shown to react with Calnexin in wild-type HEK-293T cells in Western blot with loss of signal observed in CANX knockout cell line ab255368 (CANX knockout cell lysate Human CANX (Calnexin) knockout HEK-293T cell lysate ab263805). Wild-type HEK-293T and CANX knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with Anti-Calnexin antibody [EPR3632] ab92573 and Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 °C at a 1 in 20000 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 h at room temperature before imaging.
All lanes: Western blot - Anti-Calnexin antibody [EPR3632] (Anti-Calnexin antibody [EPR3632] ab92573) at 1/20000 dilution
Lane 1: Wild-type HEK-293T cell lysate at 20 µg
Lane 2: CANX knockout HEK-293T cell lysate at 20 µg
Lane 2: Western blot - Human CANX (Calnexin) knockout HEK-293T cell line (ab255368)
Lane 3: U-2 OS cell lysate at 20 µg
Lane 4: MCF7 cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 68 kDa
Observed band size: 80 kDa
Lanes 1 - 4: Merged signal (red and green). Green - Anti-Calnexin antibody [CANX/1543] ab238078 observed at 80 kDa. Red - loading control Anti-GAPDH antibody [EPR16891] - Loading Control ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) observed at 37 kDa.
Anti-Calnexin antibody [CANX/1543] ab238078 was shown to react with Calnexin in wild-type HEK-293T cells in Western blot with loss of signal observed in CANX knockout cell line ab255368 (CANX knockout cell lysate Human CANX (Calnexin) knockout HEK-293T cell lysate ab263805). Wild-type HEK-293T and CANX knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with Anti-Calnexin antibody [CANX/1543] ab238078 and Anti-GAPDH antibody [EPR16891] - Loading Control ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) overnight at 4 °C at 1 µg/ml and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed ab216777) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
All lanes: Western blot - Anti-Calnexin antibody [CANX/1543] (Anti-Calnexin antibody [CANX/1543] ab238078) at 1 µg/mL
Lane 1: Wild-type HEK-293T cell lysate at 20 µg
Lane 2: CANX knockout HEK-293T cell lysate at 20 µg
Lane 2: Western blot - Human CANX (Calnexin) knockout HEK-293T cell line (ab255368)
Lane 3: U-2 OS cell lysate at 20 µg
Lane 4: MCF7 cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 68 kDa
Observed band size: 80 kDa
Homozygous: 19 bp deletion in exon2
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