Anti aml1

Manufactured by Cell Signaling Technology

Sourced in United States

Anti-AML1 is a lab equipment product from Cell Signaling Technology. It is an antibody that detects the AML1 (RUNX1) protein, which plays a key role in the regulation of gene expression during hematopoiesis.

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Lab products found in correlation

Anti gapdh, by Cell Signaling Technology (4 mentions) Pvdf membrane, by Merck Group (3 mentions) Anti runx1 aml1, by Abcam (2 mentions) Supersignal west pico chemiluminescent substrate, by Thermo Fisher Scientific (2 mentions) Complete protease inhibitor cocktail, by Roche (2 mentions) Chemidoc xrs imaging system, by Bio-Rad (2 mentions) Anti mouse alexa fluor 488, by Thermo Fisher Scientific (2 mentions) Anti lamin a c, by Santa Cruz Biotechnology (1 mentions) Conjugated to hrp, by Santa Cruz Biotechnology (1 mentions) Ne per nuclear and cytoplasmic extraction kit, by Thermo Fisher Scientific (1 mentions) Non fat dry milk, by Bio-Rad (1 mentions) Enhanced chemiluminescent detection reagent, by Cytiva (1 mentions) Anti α actin, by Merck Group (1 mentions) Anti acetyl histone h3, by Merck Group (1 mentions) Anti rabbit hrp, by Agilent Technologies (1 mentions) Anti mouse horseradish peroxidase hrp, by Agilent Technologies (1 mentions) Anti histone h3, by Abcam (1 mentions) Anti ythdf2 24744 1 ap, by Proteintech (1 mentions) Anti igfbp 2, by Cell Signaling Technology (1 mentions) Anti fto, by Cell Signaling Technology (1 mentions)

7 protocols using anti aml1

Western Blot Analysis of AML1 and GAPDH

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For Western blots, protein samples were run on a 4-20% gradient polyacrylamide gel (Biorad 456-8093) and transferred to a nitrocellulose membrane. The membrane was blocked in 5% milk in TBS for 1 hour at room temperature. The blots were hybridized with primary antibody overnight at 4 °C and washed with TBST, 4 times, 5min each. The blots were then incubated with an appropriate HRP conjugated secondary antibody (1:20,000) for 1 hour at room temperature followed by 4 washes with TBST 5 minutes each. The blot was developed using the chemiluminescent reagent (SuperSignal West Pico Chemiluminescent substrate, Thermo scientific 34080). The primary antibodies and dilutions used in this study were anti-AML1 (Cell Signalling Technology #4334, 1:1000), anti-Runx1/AML1 (Abcam ab23980, 1:3000) and anti-GAPDH (Cell Signaling Technology 14C10, 1:20,000).
Uncropped images of all blots can be found in the Supplementary Information (Supplementary Fig. 8)

Regha K., Assi S.A., Tsoulaki O., Gilmour J., Lacaud G, & Bonifer C. (2015). Developmental-stage-dependent transcriptional response to leukaemic oncogene expression. Nature Communications, 6, 7203.

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Western Blot Protein Detection Protocol

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For western blots, protein samples were run on a 4–20% gradient polyacrylamide gel (Biorad 456-8093) and transferred to a nitrocellulose membrane. The membrane was blocked in 5% milk in Tris Buffered Saline (TBS) for 1 h at room temperature. The blots were hybridized with primary antibody overnight at 4 °C and washed with TBS Tween (TBST), four times, 5 min each. The blots were then incubated with an appropriate horseradish peroxidase-conjugated secondary antibody (1:20,000) for 1 h at room temperature followed by four washes with TBST 5 min each. The blot was developed using the chemiluminescent reagent (SuperSignal West Pico Chemiluminescent substrate, Thermo scientific 34080). The primary antibodies and dilutions used in this study were anti-AML1 (Cell Signalling Technology #4334, 1:1,000), anti-Runx1/AML1 (Abcam ab23980, 1:3,000) and anti-GAPDH (Cell Signaling Technology 14C10, 1:20,000).
Uncropped images of all blots can be found in the Supplementary Information (Supplementary Fig. 8)

Regha K., Assi S.A., Tsoulaki O., Gilmour J., Lacaud G, & Bonifer C. (2015). Developmental-stage-dependent transcriptional response to leukaemic oncogene expression. Nature Communications, 6, 7203.

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Whole Cell Protein Isolation and Western Blotting

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Whole cell protein lysates were generated using RIPA buffer; nuclear and cytoplasmic protein lysates were generated using the NE-PER nuclear and cytoplasmic extraction kit (Thermo Scientific, Lafayette, CO, USA). Both protein isolation reagents were supplemented with 25μM MG132 and complete protease inhibitor cocktail (Roche Diagnostics, Indianapolis, IN, USA). Lysates were separated on a 10% acrylamide gel and immobilized on PVDF membranes (Millipore, Billerica, MA, USA). Blots were blocked using 5% non-fat dry milk (Bio-Rad Laboratories, Hercules, CA, USA) before being incubated overnight at 4°C with the following primary antibodies: anti-AML1 (rabbit polyclonal, 1:1000) and anti-GAPDH (rabbit monoclonal, 1:5000) (Cell Signaling, Danvers, MA, USA) and anti-Lamin A/C (goat polyclonal, 1:2000) (Santa Cruz Biotechnology, Dallas, TX, USA). GAPDH and Lamin A/C were used as loading controls. Secondary antibodies conjugated to HRP (Santa Cruz Biotechnology, Dallas, TX, USA) were used to detect proteins in conjunction with an enhanced chemiluminescence kit and Chemidoc XRS+ imaging system (both Bio-Rad Laboratories, Hercules, CA, USA).

Browne G., Taipaleenmäki H., Bishop N.M., Madasu S.C., Shaw L.M., van Wijnen A.J., Stein J.L., Stein G.S, & Lian J.B. (2015). Runx1 is associated with breast cancer progression in MMTV-PyMT transgenic mice and its depletion in vitro inhibits migration and invasion. Journal of cellular physiology, 230(10), 2522-2532.

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Western Blot Protein Expression Analysis

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Protein lysates were prepared using cell lysis buffer (150 mmol/L NaCl, 10 mmol/L Tris-HCl [pH 7.4], 5 mmol/L EDTA, 1% Triton X-100) supplemented with PMSF and the cOmplete protease inhibitor cocktail (EDTA-free; Roche). Lysates were subjected to SDS-PAGE and transferred to PVDF membrane (Millipore). Membranes were blocked in skimmed milk or BSA and probed overnight with the following antibodies: anti-AML1 (Cell Signaling Technology; cat. #4336); anti-acetyl-histone H3 (Millipore; cat. #06-599); anti-histone H3 (Abcam; cat. #1791); and anti-α-actin (Sigma; cat. #AC-74). Filters were then washed, and probed with either anti-mouse horseradish peroxidase (HRP) or anti-rabbit HRP (both DakoCytomation) secondary antibodies. Signals were visualized using enhanced chemiluminescent detection reagent (Amersham).

Salmon J.M., Todorovski I., Stanley K.L., Bruedigam C., Kearney C.J., Martelotto L.G., Rossello F., Semple T., Arnau G.M., Zethoven M., Bots M., Bjelosevic S., Cluse L.A., Fraser P.J., Litalien V., Vidacs E., McArthur K., Matthews A.Y., Gressier E., de Weerd N.A., Lichte J., Kelly M.J., Hogg S.J., Hertzog P.J., Kats L.M., Vervoort S.J., De Carvalho D.D., Scheu S., Bedoui S., Kile B.T., Lane S.W., Perkins A.C., Wei A.H., Dominguez P.M, & Johnstone R.W. (2022). Epigenetic Activation of Plasmacytoid DCs Drives IFNAR-Dependent Therapeutic Differentiation of AML. Cancer Discovery, 12(6), 1560-1579.

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Comprehensive Western Blot Analysis of Protein Expression

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Western blotting was performed with equal amounts of cell lysates (~ 20 μg), using the following antibodies: anti-FTO (#45980, 1:1000; Cell Signaling Technology), anti-AML1 (#4334, 1:1000; Cell Signaling Technology), anti-PU.1 (#2266, 1:1000; Cell Signaling Technology), anti-YTHDF2 (24744-1-AP, 1:1000; Proteintech), anti-YTHDF3 (sc-377119, 1:200; Santa Cruz Biotechnology), anti-IGFBP2 (#3922, 1:1000; Cell Signaling Technology), anti-IGFBP2 (sc-515134, 1:200; Santa Cruz Biotechnology), anti-β-actin hFAB rhodamine (12004163, 1:1000; Bio-Rad), and anti-β-actin (#4970, 1:1000; Cell Signaling Technology).

Zhou W., Li S., Wang H., Zhou J., Li S., Chen G., Guan W., Fu X., Nervi C., Yu L, & Li Y. (2024). A novel AML1-ETO/FTO positive feedback loop promotes leukemogenesis and Ara-C resistance via stabilizing IGFBP2 in t(8;21) acute myeloid leukemia. Experimental Hematology & Oncology, 13, 9.

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Immunodetection of Neural Markers

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The following mouse monoclonal antibodies (mAbs) were used: anti-glyceraldehyde-3-phosphate dehydrogenase (GAPDH, Santa Cruz Biotechnology, Heidelberg, Germany) and anti-transient receptor potential cation channel A1 (TRPA1) (Santa Cruz Biotechnology), anti-class III-beta-tubulin (β_III-tubulin) (Millipore, Billerica, MA, USA), anti-Aml1 (Cell Signaling Technology, Danvers, MA, USA). The following polyclonal Abs were used: rabbit anti-glial fibrillary acidic protein (GFAP) and rabbit anti-transient receptor potential vanilloid 1 (TRPV1) (ThermoFisher Scientific, Rodano, Italy). The following secondary Abs were used: horseradish peroxidase (HRP)-conjugated sheep anti-mouse IgG and HRP-conjugated donkey anti-rabbit IgG (Cell Signaling Technology), anti-mouse Alexa Fluor-488 (ThermoFisher Scientific), anti-rabbit Alexa Fluor-594 (ThermoFisher Scientific), PE-conjugated anti-rabbit (Santa Cruz Biotechnology), PE-conjugated anti-mouse (Santa Cruz Biotechnology). The following isotypes were used: PE-conjugated mouse IgG1 and PE-conjugated rabbit IgG1 (ThermoFisher Scientific).

Santoni G., Nabissi M., Amantini C., Santoni M., Ricci-Vitiani L., Pallini R., Maggi F, & Morelli M.B. (2021). ERK Phosphorylation Regulates the Aml1/Runx1 Splice Variants and the TRP Channels Expression during the Differentiation of Glioma Stem Cell Lines. Cells, 10(8), 2052.

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Quantitative Western Blot Analysis

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Whole cell protein lysates were generated using an SDS lysis buffer as described previously [49 (link)]. Western blots were performed and quantitated in biological triplicates and blots representative of each experiment are shown. Briefly, lysates were separated in a 10% polyacrylamide gel and immobilized on PVDF membranes (Millipore, Billerica, MA). Blots were blocked using 5% bovine serum albumin (Sigma-Aldrich, St. Louis, MO) before being incubated overnight at 4°C with the following primary antibodies: anti-AML1 (rabbit polyclonal, 1:800); anti-GAPDH (rabbit monoclonal, 1:5000) (Cell Signaling, Danvers, MA); or anti-PPARGC1B (rabbit polyclonal, 1:1000) (Novus Biologics, Littleton, CO). GAPDH was used as a loading control. Secondary antibodies conjugated to HRP (Santa Cruz Biotechnology, Dallas, TX) were used to detect proteins in conjunction with an enhanced chemiluminescence kit and Chemidoc XRS+ imaging system (both Bio-Rad, Hercules, CA).

Browne G., Dragon J.A., Hong D., Messier T.L., Gordon J.A., Farina N.H., Boyd J.R., VanOudenhove J.J., Perez A.W., Zaidi S.K., Stein J.L., Stein G.S, & Lian J.B. (2016). MicroRNA-378-mediated suppression of Runx1 alleviates the aggressive phenotype of triple negative MDA-MB-231 human breast cancer cells. Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine, 37(7), 8825-8839.

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