PTPN11 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 1 bp insertion in exon 1 and 2 bp deletion in exon 1.
BPTP3, JMML, METCDS, MGC14433, OTTHUMP00000166107, OTTHUMP00000166108, PTN11_HUMAN, PTP-1D, PTP-2C, PTPN11, Protein tyrosine phosphatase 2, Protein tyrosine phosphatase non receptor type 11, Protein-tyrosine phosphatase 1D, Protein-tyrosine phosphatase 2C, SAP-2, SH-PTP2, SH-PTP3, SH2 domain containing protein tyrosine phosphatase 2, SHP-2, Tyrosine-protein phosphatase non-receptor type 11
PTPN11 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 1 bp insertion in exon 1 and 2 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, 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.
SHP2 also known as PTPN11 is a protein tyrosine phosphatase with a molecular mass of approximately 68 kDa. It is expressed in various tissues including the heart liver and immune cells. SHP2 belongs to the non-receptor class of protein tyrosine phosphatases and plays a critical role in cell signaling by acting as a regulator of signal transduction processes. SHP2 mediates these processes by dephosphorylating specific phosphotyrosine residues on target proteins influencing various cellular functions like proliferation differentiation and survival.
The role of SHP2 extends to involvement in several signaling cascades such as the Ras/MAPK and PI3K/AKT pathways. It functions as an essential component within protein complexes that facilitate cell communication and response to external signals. The protein modulates growth factor signaling and cytokine signaling highlighting its significance in normal cell function and development. SHP2's statement in signaling processes makes it an important regulator of cellular dynamics.
SHP2 participates in the Ras/MAPK and PI3K/AKT signaling pathways which are important for regulating cell growth survival and differentiation. Within these pathways SHP2 interacts with various signaling molecules including Grb2 Sos and Gab family adaptors. These interactions coordinate cellular responses to growth factors and other extracellular cues ensuring proper pathway activation and control. By serving as a critical mediator SHP2 integrates signals that are necessary for appropriate cellular outcomes.
SHP2 is associated with several conditions such as Noonan syndrome and various cancers. Mutations in the PTPN11 gene which encodes SHP2 often result in aberrant signaling that leads to developmental anomalies or tumorigenesis. In Noonan syndrome the mutated SHP2 protein results in disrupted Ras/MAPK pathway signaling. As for cancers SHP2 is often found to be overactive leading to enhanced cell proliferation and survival. In these contexts SHP2 is interconnected with other proteins like RAS and RAF which also contribute to oncogenic pathway activation and disease progression.
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Anti-SHP2 antibody [Y478] ab32083 was shown to react with SHP2 in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266450 (knockout cell lysate Human PTPN11 (SHP2) knockout HEK-293T cell lysate ab257618) was used. Wild-type HEK-293T and PTPN11 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-SHP2 antibody [Y478] ab32083 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) overnight at 4° 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-SHP2 antibody [Y478] (Anti-SHP2 antibody [Y478] ab32083) at 1/1000 dilution
Lane 1: Wild-type HEK-293T cell lysate at 20 µg
Lane 2: PTPN11 knockout HEK-293T cell lysate at 20 µg
Lane 2: Western blot - Human PTPN11 (SHP2) knockout HEK-293T cell line (ab266450)
Performed under reducing conditions.
Predicted band size: 68 kDa
Observed band size: 68 kDa
Anti-SHP2 antibody [EPR17829-9] ab187040 was shown to react with SHP2 in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266450 (knockout cell lysate Human PTPN11 (SHP2) knockout HEK-293T cell lysate ab257618) was used. Wild-type HEK-293T and PTPN11 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-SHP2 antibody [EPR17829-9] ab187040 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) overnight at 4°C at a 1 in 5000 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-SHP2 antibody [EPR17829-9] (Anti-SHP2 antibody [EPR17829-9] ab187040) at 1/5000 dilution
Lane 1: Wild-type HEK-293T cell lysate at 20 µg
Lane 2: PTPN11 knockout HEK-293T cell lysate at 20 µg
Lane 2: Western blot - Human PTPN11 (SHP2) knockout HEK-293T cell line (ab266450)
Performed under reducing conditions.
Predicted band size: 68 kDa
Observed band size: 68 kDa
Representative images PTPN11 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 a EVOS M5000 microscope.
Allele-1: 2 bp deletion in exon1
Allele-2: 1 bp insertion in exon 1.
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized PTPN11 KO HEK293T (PTPN11 KO knock out human embryonic kidney epithelial cell) (ab266450) cells labelling SHP2 with Anti-SHP2 antibody [EPR26539-45] ab300579 at 1/50 (9.76 ug/ml) dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green). Confocal image showing cytoplasmic and weak nuclear staining in wildtype HEK293T cell line, while showing no staining in PTPN11 knockout HEK293T cell line is observed. Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5 ug/ml dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.
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