Ascl1

Manufactured by BD

Sourced in United States

ASCL1 is a laboratory equipment product manufactured by BD. It is a device designed to assist with specific laboratory procedures. The core function of ASCL1 is to perform certain tasks within a controlled laboratory environment.

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Lab products found in correlation

710 confocal microscope, by Zeiss (3 mentions) Neurod1, by Abcam (2 mentions) Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions) Phospho 4e bp1, by Abcam (1 mentions) Phospho s6, by Abcam (1 mentions) Phospho h2ax, by Abcam (1 mentions) β catenin, by Cell Signaling Technology (1 mentions) Vinculin, by Cell Signaling Technology (1 mentions) Pou3f2, by Cell Signaling Technology (1 mentions) P pras40, by Cell Signaling Technology (1 mentions) P akt, by Cell Signaling Technology (1 mentions) Hsp90, by Cell Signaling Technology (1 mentions) Meis1, by Abcam (1 mentions) Bromodeoxyuridine (brdu), by BD (1 mentions) H3k4me1, by Merck Group (1 mentions) H3k4me2, by Merck Group (1 mentions) Vinculin, by Merck Group (1 mentions) β tubulin, by Merck Group (1 mentions) Ha tag, by Abcam (1 mentions)

Close spelling variants of this product

Mouse anti-Ascl1 (8 citations) Anti-ASCL1 (5 citations) ASCL1 (clone 24B72D11.1, (3 citations) Anti-MASH1 (Ascl1; (3 citations) Mammalian achaete-scute complex-like1 (ASCL1) (2 citations) Mouse anti-Ascl1 antibody (2 citations) Anti-ASCL1 antibody (2 citations)

12 protocols using ascl1

Immunoblot Primary Antibody Panel

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Primary antibodies for immunoblot include: MYC (CST #13987, 1:1,000), MYCN (Santa Cruz sc-791, 1:200), ODC1 (EMD Millipore, MABS36, 1:100), ASCL1 (BD Bioscience, BDB556604, 1:300), phospho-4EBP1 (CST #2855, 1:1,000), 4EBP1 (CST #9644, 1:2,000), phospho-S6 (CST #2211, 1:1,000), S6 (CST #2217, 1:1,000), phospho-H2AX (CST #9718, 1:1000), PARP (CST #9532, 1:1000), ASL (Abcam#201026), ASS1 for mouse tumors (Abcam #170952, 1:1,000), ASS1 for human PDX and cell lines (Polaris Pharmaceuticals, 1:1000), and HSP90 (CST #4877, 1:1,000).

Chalishazar M.D., Wait S.J., Huang F., Ireland A.S., Mukhopadhyay A., Lee Y., Schuman S.S., Guthrie M.R., Berrett K.C., Vahrenkamp J., Hu Z., Kudla M., Modzelewska K., Wang G., Ingolia N.T., Gertz J., Lum D.H., Cosulich S.C., Bomalaski J., DeBerardinis R.J, & Oliver T.G. (2019). MYC-driven small cell lung cancer is metabolically distinct and vulnerable to arginine depletion. Clinical cancer research : an official journal of the American Association for Cancer Research, 25(16), 5107-5121.

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Protein Expression Analysis by Western Blot

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Protein extraction and western blot were performed as previously described(68 (link)). Antibodies for FOXN4 (#PA539174, ThermoFisher), ONECUT2 (#ab28466, Abcam), POU3F2 (#12137, Cell Signaling Technology), EGFR (#4267, Cell Signaling Technology), ASCL1 (#556604, BD), NEUROD1 (#ab109224, Abcam), pAKT (S473, #4060S, Cell Signaling Technology), pPRAS40 (T246, #13175, Cell Signaling Technology), Beta catenin (#8480, Cell Signaling Technology), Vinculin (#13901, Cell Signaling Technology) and actin (#3700, Cell Signaling Technology) were used.

Quintanal-Villalonga A., Taniguchi H., Zhan Y.A., Hasan M.M., Chavan S.S., Meng F., Uddin F., Manoj P., Donoghue M.T., Won H.H., Chan J.M., Ciampricotti M., Chow A., Offin M., Chang J.C., Ray-Kirton J., Tischfield S.E., Egger J., Bhanot U.K., Linkov I., Asher M., Sinha S., Silber J., Iacobuzio-Donahue C.A., Roehrl M.H., Hollmann T.J., Yu H.A., Qiu J., de Stanchina E., Baine M.K., Rekhtman N., Poirier J.T., Loomis B., Koche R.P., Rudin C.M, & Sen T. (2021). Multi-omic analysis of lung tumors defines pathways activated in neuroendocrine transformation. Cancer discovery, 11(12), 3028-3047.

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Quantitative Protein Profiling in Cell Lines

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Protein levels of Nfib and Ascl1 for the tumor-derived cell lines used in IC50 experiments were determined using the Simple Western™ quantitative immunoassay and the Compass software, according to the manufacturer’s protocol. Cells were lysed in TNESV lysis buffer (50 mM Tris·HCl at pH 7.6, 1% IGEPAL®, 20 mM EDTA at pH 8.0, 100mM NaCl), supplemented with proteasome and phosphatase inhibitors, and lysates were cleared by centrifugation at maximum speed for 10 minutes. Total protein was quantified using the Pierce BCA Protein Assay Kit (Thermo Fisher, Cat. #23277). Whole-cell lysates were diluted to a final concentration of 0.2 μl/ml. The antibodies and dilutions used were as follows: Nfib (Abcam, ab186738) 1:20000; Ascl1 (BD Pharmingen, 556604) 1:1000; Hsp90 (Cell Signaling Technology, 4877S) 1:2000.

Yang D., Denny S.K., Greenside P.G., Chaikovsky A.C., Brady J.J., Ouadah Y., Granja J.M., Jahchan N.S., Lim J.S., Kwok S., Kong C.S., Berghoff A.S., Schmitt A., Reinhardt H.C., Park K.S., Preusser M., Kundaje A., Greenleaf W.J., Sage J, & Winslow M.M. (2018). Inter-tumoral heterogeneity in SCLC is influenced by the cell-type of origin. Cancer discovery, 8(10), 1316-1331.

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Immunohistochemical Subtyping for SCLC

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All cases were stained for ASCL1 (clone: 24B72D11.1; BD Biosciences), NEUROD1 (clone: EPR17084; Abcam), and POU2F3 (clone: polyclonal; Atlas Antibodies) for subtyping. The nuclear expression of each marker was semiquantitatively recorded for percentage of positive cells (0%–100%) and intensity of staining (0 = no stain, 1 = weak, 2 = moderate, 3 = strong). A combination H score (range 0–300) was calculated as previously described: H score = (0 × % negative) + (1 × % weakly positive) + (2 × % moderately positive) + (3 × % strongly positive cells).²⁰ (link) Marker with H score less than 50 was interpreted as negative, as previously suggested.⁹ (link) Each slide was scored by two pathologists independently. Average scores were calculated and recorded for analysis. Cases with substantial discrepancy leading to subtype difference were re-reviewed by both pathologists together until consensus was reached.

Lo Y.C., Rivera-Concepcion J., Vasmatzis G., Aubry M.C, & Leventakos K. (2023). Subtype of SCLC Is an Intrinsic and Persistent Feature Through Systemic Treatment. JTO Clinical and Research Reports, 4(9), 100561.

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Cryosectioning and Immunostaining of Embryos

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Fixed embryos were rinsed in PBS, pH 7.4, transferred to 10%, 20%, then 30% sucrose in 0.1M phosphate buffer (PB) for cryoprotection, embedded in 2.5% agar in 30% PB-sucrose to position them consistently, and flash-frozen in cryoembedding media (OCT) using liquid nitrogen-cooled 2-methyl butane. Blocks were sectioned at 10 μm, serial sections mounted onto slides stored at −20°C before immunostaining. Primary antibodies against tdTomato/red fluorescent protein (RFP; Abcam, rabbit), Pax7 (Developmental Hybridoma Studies Bank, mouse), Meis1 (Abcam, rabbit), Ascl1 (BD Bio, mouse; Chemicon, rabbit), Six1 (Proteintech, rabbit; Atlas, mouse), eGFP (Abcam), pSMAD (Millipore, mouse), BrdU (BD, mouse; Novus, rat), PH3 (Cell Signaling; rabbit), βIII-tubulin (Aves, chicken), NCAM (Millipore, rat), and OMP (Pierce Biotech; chicken) followed by 2° antibodies for single, double or triple label as described previously (Karpinski et al., 2016 (link)). Images were collected on a Leica Tiling epifluorescence or Zeiss 710 confocal microscope.

Paronett E.M., Bryan C.A., Maynard T.M, & LaMantia A.S. (2023). Identity, Lineage and Fates of a Temporally Distinct Progenitor Population in the Embryonic Olfactory Epithelium. Developmental biology, 495, 76-91.

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Molecular Profiling of Stem Cell Differentiation

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TGFβ was acquired from Millipore (GF111). Antibodies used were anti: H3K27me3 (Millipore, 07449), H3K4me1 (Millipore, 07436), H3K4me2 (Millipore, 07030), H3K27me2 (Cell signaling, 9728S), H3K27ac (Abcam, ab4729), H3K4me3 (Abcam, ab8580), SMAD3 (Abcam, ab28379), phospho-SMAD3 (Cell Signaling, mAb9520), SMAD2/3 (BD Bioscience, 610842), ASCL1 (BD Pharmingen, 556604), JMJD3 (raised in the laboratory using amino acids 798–1095), CHD8 was raised in Dr José Carlos Reyes laboratory (32 (link)), β-TUBULIN III (TUJ1, Covance, MMS-435P), GFAP (Dako, z0334), NESTIN (Abcam, ab5968), HuC/D (MP, A21271), DAPI (ThermoFisher, D1306), β-TUBULIN (Millipore, MAB3408), VINCULIN (Sigma, V9131), HA tag (Abcam, ab20084) and MYC tag (Abcam, ab9132).

Fueyo R., Iacobucci S., Pappa S., Estarás C., Lois S., Vicioso-Mantis M., Navarro C., Cruz-Molina S., Reyes J.C., Rada-Iglesias Á., de la Cruz X, & Martínez-Balbás M.A. (2018). Lineage specific transcription factors and epigenetic regulators mediate TGFβ-dependent enhancer activation. Nucleic Acids Research, 46(7), 3351-3365.

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Histological Analysis of Organs

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Animals were sacrificed when they acquired respiratory distress. Tissues and organs were collected and fixed in EAF fixative (ethanol/acetic acid/formaldehyde/saline at 40:5:10:45 v/v) and embedded in paraffin. Sections were prepared at 2 μm thickness from the paraffin blocks and stained with hematoxylin and eosin (HE) according to standard procedures. For immunohistochemistry (IHC), 4 μm-thick sections were made on which the following antibodies were applied: Synaptophysin/SYP (Abcam, ab32127), E-cadherin/CDH1 (Cell Signaling Technology, 3195), NFIB (Thermo Fisher Scientific, PA5-28299), CGRP (Sigma-Aldrich, C8198), Ki67 (Abcam, ab15580), keratin wide-spectrum/KWS (Agilent, Z0622), phospho-histone H3/p-HH3 (Millipore, 04-746), Podoplanin/PDPN (Abcam, ab11936), ASCL1 (BD Biosciences, 556604), ALDH1A1 (Abcam, ab23375), NEUROD1 (Proteintech, 12081-1-ap), phospho-AKT (Cell Signaling Technology, 4060) and phospho-4EBP1 (Cell Signaling Technology, 2855). The sections were reviewed with a Zeiss Axioskop2 Plus microscope (Carl Zeiss Microscopy) and images were captured with a Zeiss AxioCam HRc digital camera and processed with AxioVision 4 software (both from Carl Zeiss Vision).

Böttger F., Semenova E.A., Song J.Y., Ferone G., van der Vliet J., Cozijnsen M., Bhaskaran R., Bombardelli L., Piersma S.R., Pham T.V., Jimenez C.R, & Berns A. (2019). Tumor Heterogeneity Underlies Differential Cisplatin Sensitivity in Mouse Models of Small-Cell Lung Cancer. Cell Reports, 27(11), 3345-3358.e4.

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ASCL1 Expression in Rhabdomyosarcomas

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Tumors were immunostained for desmin (Leica, Novocastra, Buffalo Grove, IL, clone DE-R-11, 1:50 – 1:100), myogenin (Dako, Carpinteria, CA, clone F5D, 1:25 – 1:50), MyoD1 (Ventana, Tucson, AZ, clone EP212, 1μg/mL), synaptophysin (Leica, Novocastra, Buffalo Grove, IL, clone 27G12, 1:100 – 1:200), chromogranin A (Ventana, Tucson, AZ, clone LK2H10, 1μg/mL), CD56 (Dako, Carpinteria, CA, clone 123C3, 1:50 – 1:100), TTF1 (Leica, Novocastra, Buffalo Grove, IL, clone SPT24, 1:200) and ASCL1 (BD Pharmingen, San Jose, CA, Clone: 24B72D11.1, 1:100). Immunohistochemical results were dichotomized into cases with absent expression and those with positive expression, defined by staining noted in at least 5% of neoplastic cells. Immunophenotyping of all RMS was performed on representative whole slide sections.
ASCL1 expression in rhabdomyosarcomas was compared to 70 patients with HGNEC and 110 patients with typical urothelial carcinomas (UC) from specimens obtained from patients treated with radical cystectomy at our institution between 1987 and 2014 (18 (link)). Tissue microarrays, with four 1.0mm cores representing each HGNEC and typical UC, were immunostained for ASCL1 for this purpose. In addition, whole slide sections of 47 cases of non-urinary bladder rhabdomyosarcomas were immunostained for ASCL1, to assess specificity.

Gupta S., Sosa C.P., Kosari F., Folpe A., Bhinge K.N., Yang L., Agahi A., Johnson S.H., Frank I., Boorjian S.A., Hansel D.E., Al-Ahmadie H.A., Reuter V.E., Vasmatzis G., Jimenez R.E., Herrera-Hernandez L, & Cheville J.C. (2019). A Comparison of Adult Rhabdomyosarcoma and High Grade Neuroendocrine Carcinoma of the Urinary Bladder Reveals Novel PPP1R12A Fusions in Rhabdomyosarcoma. Human pathology, 88, 48-59.

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Western Blotting of SCLC Proteins

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Protein lysates from SCLC lines and murine tumors were prepared in RIPA buffer, supplemented with PhosSTOP phosphatase and cOmplete protease inhibitor cocktail (Roche #11873580001, #04906845001). Protein samples were quantified with the BCA assay. Samples were separated on 4–20% SDS-PAGE gels, transferred to PVDF membranes, and probed with antibodies against ASCL1 (BD Bioscience #556604); ACTB (Sigma #A3854); IMPDH1 (Sigma #SAB2101156); IMPDH2 (Abcam #ab131158); MAT2A (Abcam #ab77471); MYC (Cell Signaling #5605); GMPS (Cell Signaling #14602); Cleaved Caspase-3 (Cell Signaling #9664); mTOR (Cell Signaling #9862); Phospho-mTOR S2448 (Cell Signaling #9862); Phospho-4EBP1 T37/46 (Cell Signaling #9862); Phospho-p70 S6 Kinase T389 (Cell Signaling #9234); Phospho-S6 S235/236 (Cell Signaling #4858). Bands were detected with the ECL blotting system (Pierce #32106).

Huang F., Ni M., Chalishazar M.D., Huffman K.E., Kim J., Cai L., Shi X., Cai F., Zacharias L.G., Ireland A.S., Li K., Gu W., Kaushik A.K., Liu X., Gazdar A.F., Oliver T.G., Minna J.D., Hu Z, & DeBerardinis R.J. (2018). Inosine monophosphate dehydrogenase dependence in a subset of small cell lung cancers. Cell metabolism, 28(3), 369-382.e5.

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Protein Expression Analysis by Western Blot

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Lysate preparation and Western blot analyses were performed as described previously [34 (link)]. Antibodies used are MYCL (AF4050, R&D), MYC (sc-40, Santa Cruz), MYCN (#9405, Cell Signaling), MAX (sc-197, Santa Cruz), ASCL1 (556604, BD Pharmingen), p21 (#2947, Cell Signaling), PARP-1 (#9542, Cell Signaling), and α-Tubulin (CP06, CalBiochemicals).

Kato F., Fiorentino F.P., Alibés A., Perucho M., Sánchez-Céspedes M., Kohno T, & Yokota J. (2016). MYCL is a target of a BET bromodomain inhibitor, JQ1, on growth suppression efficacy in small cell lung cancer cells. Oncotarget, 7(47), 77378-77388.

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