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Phospho flt3

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Phospho-FLT3 is a laboratory product used to detect and quantify the phosphorylated form of the Fms-like tyrosine kinase 3 (FLT3) protein. This protein plays a critical role in cell signaling pathways and is commonly studied in various research applications.

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Lab products found in correlation

Phospho stat5, by Cell Signaling Technology (3 mentions) Stat5, by Cell Signaling Technology (2 mentions) Phospho gsk 3β ser9, by Cell Signaling Technology (1 mentions) Phospho 4e bp1 thr37 46, by Cell Signaling Technology (1 mentions) Phospho foxo1, by Cell Signaling Technology (1 mentions) Phospho gsk3β, by Cell Signaling Technology (1 mentions) C myc, by Cell Signaling Technology (1 mentions) Gapdh, by Cell Signaling Technology (1 mentions) P70s6k, by Cell Signaling Technology (1 mentions) Phospho akt ser473, by Cell Signaling Technology (1 mentions) Phospho akt thr308, by Cell Signaling Technology (1 mentions) Phospho nf κb p65, by Cell Signaling Technology (1 mentions) Phospho pras40, by Cell Signaling Technology (1 mentions) Flt3 ligand, by R&D Systems (1 mentions) Phospho p44 42 mapk erk1 2, by Cell Signaling Technology (1 mentions) Phospho 4ebp1, by Cell Signaling Technology (1 mentions) Phospho p70s6k thr389, by Cell Signaling Technology (1 mentions) Dynabead, by Thermo Fisher Scientific (1 mentions) Protein a g agarose bead, by Santa Cruz Biotechnology (1 mentions)

3 protocols using phospho flt3

1

Investigating FLT3 Signaling Pathways

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MOLM14, MV4-11 cells were treated with DMSO, serially diluted A674563, TCS359 (1 μM), MK2206 (1 μM) for 4 hours. Cells were then washed in PBS and lysed in cell lysis buffer. FLT3, Phospho-FLT3 (Tyr589/591), AKT, Phospho-AKT Ser473, Phospho-AKT Thr308, GSK-3β, Phospho-GSK-3β (Ser9), Phospho-FoxO1 (Thr24), FoxO1, PRAS40, Phospho-PRAS40 (Thr246), STAT5, Phospho-STAT5 (Tyr694), NF-ΚB-P65, Phospho-NF-ΚB-P65 (Ser536), P70S6K, Phospho-P70S6K Thr389, 4EBP1, Phospho-4EBP1 (Thr37/46), ERK, Phospho-p44/42MAPK (Erk1/2) (Thr202/Tyr204), C-Myc and GAPDH antibodies (Cell Signaling Technology) were used for immunoblotting.
For FL addition experiment, MV4-11 cells were treated with FLT3 inhibitors in the presence of 10 ng/mL of FL (FLT3 Ligand, R&D Systems) or absence of FL for 2 hrs. Cells were then washed in PBS and lysed in cell lysis buffer. FLT3, Phospho-FLT3 (Tyr589/591), AKT, GSK-3β, Phospho-GSK-3β (Ser9), Phospho-FoxO1 (Thr24), FoxO1, STAT5, Phospho-STAT5 (Tyr694), P70S6K, Phospho-P70S6K (Thr389), 4EBP1, Phospho-4EBP1 (Thr37/46), C-Myc and GAPDH antibodies (Cell Signaling Technology) were used for immunoblotting.
Wang A., Wu H., Chen C., Hu C., Qi Z., Wang W., Yu K., Liu X., Zou F., Zhao Z., Wu J., Liu J., Liu F., Wang L., Stone R.M., Galinksy I.A., Griffin J.D., Zhang S., Weisberg E.L., Liu J, & Liu Q. (2016). Dual inhibition of AKT/FLT3-ITD by A674563 overcomes FLT3 ligand-induced drug resistance in FLT3-ITD positive AML. Oncotarget, 7(20), 29131-29142.
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2

FLT3 Phosphorylation and Signaling Assay

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Cells were treated with pacritinib or DMSO for 4 hours and lysed in RIPA buffer supplemented with protease and phosphatase inhibitors. Immunoprecipitation was carried out overnight with anti-FLT3 antibodies (Santa Cruz Biotechnology, Dallas, TX). Dynabeads (ThermoFisher, Waltham, MA) were added the next day and incubated for 4 hours. Samples were then prepared according to manufacturer’s instructions. Eluted lysates or total cell lysates were separated by SDS-polyacrylamide gel electrophoresis and transferred to PVDF membranes followed by western blot analysis using the indicated primary antibodies: FLT3, phospho-FLT3, STAT5, and phospho-STAT5 (Cell Signaling Technology, Danvers, MA).
Jeon J.Y., Zhao Q., Buelow D.R., Phelps M., Walker A.R., Mims A.S., Vasu S., Behbehani G., Blachly J., Blum W., Klisovic R.B., Byrd J.C., Garzon R., Baker S.D, & Bhatnagar B. (2019). Preclinical activity and a pilot phase I study of pacritinib, an oral JAK2/FLT3 inhibitor, and chemotherapy in FLT3-ITD-positive AML. Investigational new drugs, 38(2), 340-349.
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3

Profiling FLT3 and STAT5 Signaling

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Whole-cell lysates were made
with radioimmunoprecipitation assay buffer (20 mM Tris–HCl
(pH 7.5), 150 mM NaCl, 1 mM Na2EDTA, 1 mM EGTA, 1% NP-40,
1% sodium deoxycholate, 0.1% SDS, and protease and phosphatase inhibitors).
Immunoprecipitation was carried out with anti-FLT3 antibody and Protein
A/G agarose beads from Santa Cruz Biotechnology (Dallas, TX). Invitrogen
Bis–tris-gradient mini or midi gels were used for western blot
analysis, followed by detection with enhanced chemiluminescence (ECL)
reagent. Primary antibodies used were from Cell Signaling Technology
(Danvers, MA): FLT3, phospho-FLT3, STAT5, and phospho-STAT5. Secondary
antibodies were from Jackson Immunoresearch and Cell Signaling Technology.
Jarusiewicz J.A., Jeon J.Y., Connelly M.C., Chen Y., Yang L., Baker S.D, & Guy R.K. (2017). Discovery of a Diaminopyrimidine FLT3 Inhibitor Active against Acute Myeloid Leukemia. ACS Omega, 2(5), 1985-2009.
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