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Western blot analysis of Sup-M2 cell lysate, using Phospho-ALK (Tyr1586) (3B4) Rabbit mAb (A and B), and ALK Antibody (C and D).The phospho-specificity of this antibody was characterized by treating the membrane with calf intestinal alkaline phosphatase (CIP) (B and D) after Western transfer.
Western blot analysis of extracts from NK-92 cells, untreated or treated with Human Interleukin-2 (hIL-2) #8907 (10 ng/ml, 15 minutes), using Phospho-Jak3 (Tyr980/981) (D44E3) Rabbit mAb (upper) or Jak3 Antibody #3775 (lower).
Western blot analysis of extracts from UT-7 cells, untreated or treated with erythropoietin (EPO; 3 units/ml; 5 min) and from untreated HEL cells, using Phospho-Jak2 (Tyr1007) (D15E2) Rabbit mAb (upper) or total Jak2 (D2E12) Rabbit mAb #3230 (lower). HEL cells contain a V617F activating mutation in Jak2.
Chromatin immunoprecipitations were performed with cross-linked chromatin from Hep G2 cells starved overnight and treated with IL-6 (100 ng/ml) for 30 minutes and Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795. The figure shows binding across IRF1, a known target gene of Phospho-Sata3 (see additional figure containing ChIP-qPCR data). For additional ChIP-seq tracks, please download the product data sheet.
Flow cytometric analysis of TF-1 cells, untreated (blue, low) or treated with hGM-CSF #8922 (50 ng/ml, 15 min; green, positive) using Phospho-Stat5 (Tyr694) (D47E7) XP® Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Flow cytometric analysis of Jurkat cells, treated with U0126 (10 µM, 2 hrs; green) or treated with TPA #4174 (200 nM, 30 min; blue) using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Flow cytometric analysis of Jurkat cells, untreated (green) or treated with LY294002 #9901, Wortmannin #9951, and U0126 #9903 (50 μM, 1 μM, and 10 μM, 2 hr; blue) using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Western blot analysis of extracts NIH/3T3 cells, serum-starved or treated with human Platelet-Derived Growth Factor BB hPDGF-BB #8912 (100 ng/ml, 15 min), using Phospho-Src Family (Tyr416) (D49G4) Rabbit mAb (upper) or Src (36D10) Rabbit mAb #2109 (lower).
Flow cytometric analysis of NIH/3T3 cells, untreated (blue) or treated with mPDGFbb (200ng/mL, 15 min; green) using Phospho-PLCγ1 (Tyr783) (D6M9S) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
Chromatin immunoprecipitations were performed with cross-linked chromatin from HaCaT cells Hep G2 cells starved overnight and treated with IL-6 (100 ng/ml) for 30 minutes and either Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Plus Sonication Chromatin IP Kit #56383. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human c-Fos Promoter Primers #4663, human IRF1 promoter primers, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
Confocal immunofluorescent analysis of A-431 cells, EGF-treated (left) or untreated (right), using Phospho-Stat5 (Tyr694) XP®(D47E7) Rabbit mAb (green) and Pan-Keratin (C11) Mouse mAb #4545 (red).
Confocal immunofluorescent analysis of Drosophila egg chambers, untreated (top) or λ phosphatase-treated (bottom), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb #4370 (green) and S6 Ribosomal Protein (54D2) Mouse mAb #2317 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Confocal immunofluorescent analysis of C2C12 cells, LY294002-treated (left) or insulin-treated (right), using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (green). Actin filaments have been labeled with Alexa Fluor® 555 phalloidin #8953 (red). Blue pseudocolor = DRAQ5®#4084 (fluorescent DNA dye).
Immunoprecipitation of phospho-PLCγ1 (Tyr783) from A-431 cell extracts stimulated with hEGF #8916 using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or Phospho-PLCγ1 (Tyr783) (D6M9S) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using Phospho-PLCγ1 (Tyr783) (D6M9S) Rabbit mAb.
Chromatin immunoprecipitations were performed with cross-linked chromatin from Hep G2 cells starved overnight and treated with IL-6 (100 ng/ml) for 30 minutes, and either Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb or of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using human IRF-1 promoter primers, SimpleChIP® Human c-Fos Promoter Primers #4663, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
Western blot analysis of extracts from UT-7 cells, untreated or treated with erythropoietin (EPO; 3 units/ml for 5 min), TF-1 cells, untreated or treated with Human Granulocyte Macrophage Colony Stimulating Factor #8922 (hGM-CSF; 100 ng/ml for 10 min), and NK-92 cells, untreated or treated with Human Interleukin-2 #8907 (hIL-2; 100 ng/ml for 10 min), using Phospho-Stat5 (Tyr694) (D47E7) XP® Rabbit mAb.
Confocal immunofluorescent analysis of HT1080 cells, starved overnight then treated with U0126 #9903 (10 uM, 2 h; left) or PDBu (Phorbol 12,13-Dibutyrate) #12808 (100 nM, 15 m; right) using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb #4370 (green) and β-Actin (8H10D10) Mouse mAb #3700 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Immunohistochemical analysis of paraffin-embedded MDA-MB-468 xenograft using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (left) or PTEN (138G6) Rabbit mAb #9559 (right). Note the presence of P-Akt staining in the PTEN deficient MDA-MB-468 cells.
Western blot analysis of extracts from NIH/3T3 cells, untreated (-) or stimulated with hPDGF-BB #8912 (5 min; +), and from A-431 cells, untreated (-) or stimulated with hEGF #8916 (5 min; +), using Phospho-PLCγ1 (Tyr783) (D6M9S) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).
Flow cytometric analysis of U266 cells, untreated (blue) or treated with IFNalpha (50 ng/ml, 15 min; green) using Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb (solid lines) or concentration matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb.
Immunohistochemical analysis of paraffin-embedded human breast carcinoma comparing SignalStain® Antibody Diluent #8112 (left) to TBST/5% normal goat serum (right) using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060.
Confocal immunofluorescent analysis of HeLa cells, IFN-alpha treated (left) or untreated (right), labeled with Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb (green).
Immunohistochemical analysis of paraffin-embedded human lung carcinoma, untreated (left) or λ phosphatase-treated (right), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb.
Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb.
Immunohistochemical analysis of paraffin-embedded human neuroendocrine lung carcinoma using Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb performed on the Leica® BOND™ Rx.
Immunohistochemical analysis using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb on SignalSlide™ Phospho-p44/42 MAPK (Thr202/Tyr204) IHC Controls #8103 (paraffin-embedded NIH/3T3 cells, treated with U0126 #9903 (left) or TPA #4174 (right).
Immunohistochemical analysis using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb on SignalSlide® Phospho-Akt (Ser473) IHC Controls #8101 (paraffin-embedded LNCaP cells, untreated (left) or LY294002-treated (right)).
Immunohistochemical analysis of paraffin-embedded human prostate adenocarcinoma using Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb performed on the Leica® BOND™ Rx.
Western blot analysis of extracts from COS cells, untreated or treated with either U0126 #9903 (10 µM for 1h) or TPA #4174 (200 nM for 10 m), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb #4370 and p44/42 MAPK (Erk1/2) (3A7) Mouse mAb #9107.
Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb.
Immunohistochemical analysis of paraffin-embedded Apc (min/+) mouse intestine, using Phosho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb.
Western blot analysis of extracts from 293, NIH/3T3 and C6 cells, treated with λ phosphatase or TPA #4174 as indicated, using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb (upper), or p44/42 MAPK (Erk1/2) (137F5) Rabbit mAb #4695 (lower).
Immunohistochemical analysis of paraffin-embedded PTEN heterozygous mutant mouse endometrium using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb. (Tissue section courtesy of Dr. Sabina Signoretti, Brigham and Women's Hospital, Harvard Medical School, Boston, MA.)
Immunohistochemical analysis of paraffin embedded human breast carcinoma, specifically endothelial cells, untreated (left) or lambda phosphatase treated (right), using Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb.
Immunohistochemical analysis of paraffin-embedded U-87MG xenograft, untreated (left) or lambda phosphatase-treated (right), using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb.
Immunnohistochemical analysis of paraffin-embedded human lung carcinoma, showing nuclear localization, using Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb.
Immunoprecipitation of phospho-Akt (Ser473) from Jurkat extracts treated with Calyculin A #9902 (100nM, 30 min). Lane 1 is 10% input, lane 2 is Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb, and lane 3 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900. Western blot analysis was performed with Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb. Anti-rabbit IgG, HRP-linked Antibody #7074 was used as a secondary antibody.
Immunohistochemical analysis using Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb on SignalSlide® HeLa -/+ IFNa IHC Controls #55861 (paraffin-embedded HeLa cell pellets, untreated (left) or treated with Human Interferon-α1 (hIFN-α1) #8927 (right)).
Western blot analysis of extracts from PC-3 cells, untreated or LY294002/wortmannin-treated, and NIH/3T3 cells, serum-starved or PDGF-treated, using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (upper) or Akt (pan) (C67E7) Rabbit mAb #4691 (lower).
Western blot analysis of extracts from IFN-alpha treated Jurkat cells and HeLa cells (left), as well as EGF treated A431 cells (right), using Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb. Note that the basal phospho-Stat3 in A431 is detected by the antibody.
| Product # | Size | Price |
|---|---|---|
| 8336T | 1 Kit (9 x 20 µl) | $ 626 |
| Product Includes | Quantity | Applications | Reactivity | MW(kDa) | Isotype |
|---|---|---|---|---|---|
| Phospho-ALK (Tyr1586) (3B4) Rabbit mAb 3348 | 20 µl |
|
H | 80 (NPM-ALK) 220 (ALK) | Rabbit IgG |
| Phospho-Jak3 (Tyr980/981) (D44E3) Rabbit mAb 5031 | 20 µl |
|
H M | 115 | Rabbit IgG |
| Phospho-Jak2 (Tyr1007) (D15E2) Rabbit mAb 4406 | 20 µl |
|
H M | 125 | Rabbit IgG |
| Phospho-Stat3 (Tyr705) (D3A7) XP® Rabbit mAb 9145 | 20 µl |
|
H M R Mk | 79, 86 | Rabbit IgG |
| Phospho-Stat5 (Tyr694) (D47E7) XP® Rabbit mAb 4322 | 20 µl |
|
H M | 90 | Rabbit IgG |
| Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb 4370 | 20 µl |
|
H M R Hm Mk Mi Dm Z B Dg Pg Sc | 44, 42 | Rabbit IgG |
| Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb 4060 | 20 µl |
|
H M R Hm Mk Dm Z B | 60 | Rabbit IgG |
| Phospho-Src Family (Tyr416) (D49G4) Rabbit mAb 6943 | 20 µl |
|
H M R Mk | 60 | Rabbit IgG |
| Phospho-PLCγ1 (Tyr783) (D6M9S) Rabbit mAb 14008 | 20 µl |
|
H M | 155 | Rabbit IgG |
| Anti-rabbit IgG, HRP-linked Antibody 7074 | 100 µl |
|
Goat |
Product Information
The ALK Activation Antibody Sampler Kit provides an economical means to evaluate the activation status of multiple members of the ALK pathway, including phosphorylated ALK, Jak2, Jak3, Stat3, Stat5, PLCγ1, Akt, Src, and p44/42 MAPK. The kit includes enough antibody to perform two western blot experiments with each primary antibody.
Each antibody in the ALK Activation Antibody Sampler Kit recognizes the phosphorylated form of its specific target. Phospho-ALK (Tyr1586) (3B4) Rabbit mAb may cross-react with other activated protein tyrosine kinases including EGFR. Phospho-Jak2 (Tyr1007) (D15E2) Rabbit mAb may cross react with phosphorylated Jak3 and Tyk2. Phospho-Src Family (Tyr416) (D49G4) Rabbit mAb may cross react with overexpressed phosphorylated RTKs.
Rabbit monoclonal antibodies are produced by immunizing animals with synthetic phosphopeptides corresponding to residues surrounding Tyr1586 of human ALK, Tyr1007 of human Jak2, Tyr980/981 of human and mouse Jak3, Tyr705 of mouse Stat3, Tyr694 of human Stat5a, Thr202/Tyr204 of human p44 MAP kinase, Ser473 of human Akt, Tyr416 of human Src, or Tyr783 of human PLCγ1 protein.
Anaplastic lymphoma kinase (ALK) is a tyrosine kinase receptor for pleiotrophin (PTN), a growth factor involved in embryonic brain development (1-3). In ALK-expressing cells, PTN induces phosphorylation of both ALK and the downstream effectors IRS-1, Shc, PLCγ, and PI3 kinase (1). ALK was originally discovered as a nucleophosmin (NPM)-ALK fusion protein produced by a translocation (4). Investigators have found that the NPM-ALK fusion protein is a constitutively active, oncogenic tyrosine kinase associated with anaplastic lymphoma (4). Research literature suggests that activation of PLCγ by NPM-ALK may be a crucial step for its mitogenic activity and involved in the pathogenesis of anaplastic lymphomas (5).
A distinct ALK oncogenic fusion protein involving ALK and echinoderm microtubule-associated protein like 4 (EML4) has been described in the research literature from a non-small cell lung cancer (NSCLC) cell line, with corresponding fusion transcripts present in some cases of lung adenocarcinoma. The short, amino-terminal region of the microtubule-associated protein EML4 is fused to the kinase domain of ALK (6-8).
Explore pathways + proteins related to this product.
STRING - Known and Predicted Protein-Protein Interactions.
UniProt ID: P31749 , P31751 , Q9Y243 , Q9UM73 , P27361 , P28482 , P06241 , P08631 , O60674 , P52333 , P06239 , P07948 , P19174 , P12931 , P40763 , P42229 , P51692 , P07947
Entrez-Gene Id: 207 , 208 , 10000 , 238 , 5595 , 5594 , 2534 , 3055 , 3717 , 3718 , 3932 , 4067 , 5335 , 6714 , 6774 , 6776 , 6777 , 7525
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