A 1331852

Manufactured by Selleck Chemicals

Sourced in United States, Germany

A-1331852 is a laboratory centrifuge used for separating and isolating substances based on their density and size. It is a versatile and reliable piece of equipment designed for a wide range of applications in various scientific and research settings.

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Lab products found in correlation

25 protocols using a 1331852

Evaluating Anti-Apoptotic Proteins in Hematological Malignancies

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All chemicals apart from ABT-199, A1331852, A1155463, A1210477 (Selleck Chemicals, Houston, TX, USA), and S63845 (ApexBio, Taiwan) were from Sigma (Deisenhofen, Germany). Most cell lines used in this study were obtained from Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ; Braunschweig, Germany) except Pfeiffer and SUDHL2 cells (American Type Culture Collection; Manassas, VA, USA), OCI-LY10 (Sandeep Dave, Duke University, Durham, NC, USA), MedB1^{16 (link)} (Peter Moeller, University of Ulm, Ulm, Germany) and Karpas-1106^{17 (link)} (Abraham Karpas, University of Cambridge, Cambridge, UK). All cell lines were authenticated by short tandem repeat profiling and routinely tested for mycoplasma contamination. Primary patient-derived samples were obtained from patients attending the University Hospital of Leicester, UK. Local ethical approval (Leicestershire, Northamptonshire and Rutland REC06/Q2501/122) and patients’ consent were obtained through the Haematological Tissue Bank of the Ernest and Helen Scott Haematological Research Institute, Leicester, UK. Peripheral blood mononuclear cells were isolated from the blood of patients presenting in leukemic phase and the CellTiterGlo assay (Promega, Mannheim, Germany) was used to assess these cells’ viability.

Smith V.M., Dietz A., Henz K., Bruecher D., Jackson R., Kowald L., van Wijk S.J., Jayne S., Macip S., Fulda S., Dyer M.J, & Vogler M. (2020). Specific interactions of BCL-2 family proteins mediate sensitivity to BH3-mimetics in diffuse large B-cell lymphoma. Haematologica, 105(8), 2150-2163.

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Evaluating AML Cell Sensitivity to BCL-2 Inhibitors

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The sensitivity of AML cell lines towards ABT-199 (Selleck), A1331852 (Selleck) and S63845 (Appexbio) was assessed with CellTiter-Glo viability assay (Promega). Cells were seeded in a density of 1 × 10⁵ cells/ml in a white 96-well plate. After 72 h of incubation 5 μl/well of CellTiter-Glo reagent was added and luminescence was measured with a Tecan Infinite M200 plate reader. Values were normalized to the untreated control sample.

Ewald L., Dittmann J., Vogler M, & Fulda S. (2019). Side-by-side comparison of BH3-mimetics identifies MCL-1 as a key therapeutic target in AML. Cell Death & Disease, 10(12), 917.

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Mitochondrial Dynamics Regulation Assay

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Apogossypol and Leflunomide from ApexBio (Boston, MA, USA), A-1331852, A-1210477, ABT-199, Z-VAD.FMK and CCCP from Selleck (Houston, TX, USA), Teriflunomide, norhydroguaiaretic acid (NDGA), Ivermectin, Terfenadine, Suloctidil, orotate and uridine from Sigma Aldrich (St Louis, MO, USA), MitoTracker Deep Red FM from Thermo Fisher (Loughborough, UK) were used. Antibodies against BAP31, RTN4, BiP, PDI, CHOP, DHODH and tubulin from Abcam (Cambridge, UK), CLIMP-63 and BAX (6A7) from Enzo Life Sciences (Exeter, UK), TIM22 and KNT-1 from Sigma, HSP60, Cytochrome c, BAX, OPA1 and DRP-1 from BD Biosciences (San Jose, CA, USA), phospho-DRP-1 (S616), phospho-DRP-1 (S637), MFN1 and MFN2 from Cell Signaling Technologies (Danvers, MA, USA), BAK (AB-1) from Calbiochem (Watford, UK), MFF, MID49 and MID51 from ProteinTech (Manchester, UK) and GAPDH from Santa Cruz Biotechnologies (Santa Cruz, CA, USA) were used. For RNA interference, cells were transfected with 10 nM of siRNAs against DHODH (SI00363384 and SI00363391) purchased from Qiagen Ltd. (Manchester, UK), using Interferin (Polyplus Transfection Inc, NY), according to the manufacturer’s protocol and processed 72 h after transfection.

Yedida G., Milani M., Cohen G.M, & Varadarajan S. (2019). Apogossypol-mediated reorganisation of the endoplasmic reticulum antagonises mitochondrial fission and apoptosis. Cell Death & Disease, 10(7), 521.

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BH3 Profiling of Cell Lines

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BH3 Profiling was performed using the iBH3 plate-based method as previously published (56 (link)). In brief, cell lines were seeded at a density of 1 x 10⁶ cells/mL 24 hours before profiling. Four million cells of each cell line were pelleted at 300 x g for 5 minutes and resuspended in 2mL MEB-P25 (150mM Mannitol, 10mM HEPES-KOH pH 7.5, 150mM KCl, 1mM EGTA, 1mM EDTA, 0.1% BSA, 5 mM Succinate, 0.25% Polaxamer 188[Fisher, MT61161RM]). Cells were permeabilized with digitonin (Sigma, D5628) exposed to BH3 peptides for 60 minutes at 25°C and mitochondrial Cytochrome C release was measured by flow cytometry using a FITC-conjugated antibody (BioLegend, 983502). BH3 peptides were synthesised by New England Peptide using published sequences (100 (link)). The Bcl-XL-selective inhibitor A-1331852 (Selleckchem, S7801), the Bcl-2-selective inihibtor ABT-199 (Selleckchem, S8048) and the MCL-inhibitor AZD5991 (Selleckchem, S8643) were used at a concentration of 10 μM. Results were only deemed valid where cell cytochrome C release in presence of DMSO control was <10% and cytochrome C release in presence of 50ug/mL(25uM) Alamethicin (Enzo, BML-A150-0005) was >90%. Figure represents mean of three independent experiments.

Flümann R., Rehkämper T., Nieper P., Pfeiffer P., Holzem A., Klein S., Bhatia S., Kochanek M., Kisis I., Pelzer B.W., Ahlert H., Hauer J., da Palma Guerreiro A., Ryan J.A., Reimann M., Riabinska A., Wiederstein J., Krüger M., Deckert M., Altmüller J., Klatt A.R., Frenzel L.P., Pasqualucci L., Béguelin W., Melnick A.M., Sander S., Montesinos-Rongen M., Brunn A., Lohneis P., Büttner R., Kashkar H., Borkhardt A., Letai A., Persigehl T., Peifer M., Schmitt C.A., Reinhardt H.C, & Knittel G. (2020). An Autochthonous Mouse Model of Myd88- and BCL2-Driven Diffuse Large B-cell Lymphoma Reveals Actionable Molecular Vulnerabilities. Blood Cancer Discovery, 2(1), 70-91.

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Phagocytosis Assay for Apoptotic Cancer Cells

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For induction of tumor cell apoptosis, the BC cell line Cal51 was treated with 70nM paclitaxel (Sigma-Aldrich) plus 100nM of the BCLxL antagonist A1331852 (SelleckChem) for 24h, as previously described in (24 (link)). Meso13 cells were treated with 1.6 µg/ml cisplatin (Merck) and 80 µM pemetrexed (Sigma-Aldrich) for 72h, as previously described in (16 (link)). Apoptotic and control untreated cells were then harvested, washed and labelled with 1µg/ml of the pH sensitive dye pHrodo^® (Incucyte pHrodo Red cell labelling Kit for phagocytosis, Sartorius, 4649) following the manufacturer’s protocol. In parallel, differentiated macrophages were seeded at 10⁴ cells in 96-well plates in 200µl of complete medium and treated with 10µg/ml αChemR23 or hIgG1 antibodies for 24h at 37°C. Then, labelled apoptotic or untreated cancer cells were added to macrophages at a ratio of 1:5 for 24h and phagocytosis was measured by live cell imaging using Incucyte^® technology (Sartorius). Phagocytosis was quantified using fluorescence intensity (“Total integrated intensity”) in integrated software.

Lavy M., Gauttier V., Dumont A., Chocteau F., Deshayes S., Fresquet J., Dehame V., Girault I., Trilleaud C., Neyton S., Mary C., Juin P., Poirier N., Barillé-Nion S, & Blanquart C. (2023). ChemR23 activation reprograms macrophages toward a less inflammatory phenotype and dampens carcinoma progression. Frontiers in Immunology, 14, 1196731.

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Compound Screening for Cellular Assays

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ABT-199, ABT-263, ABT-737, WEHI-539, A-1331852, A-1155463, obatoclax, and gemcitabine were purchased from Selleck Chemicals (Houston, TX, USA), Saliphenylhalamide (SaliPhe) was synthesized as described previously [24 (link)]. 10 mM solutions of the compounds were prepared in 100% dimethyl sulfoxide (Sigma-Aldrich, St. Louis, MO, USA) and stored at −80 °C. Lyophilized lipopolysaccharide (LPS, 10 mg/mL) was purchased from Sigma-Aldrich (St. Louis, MO, USA). Plasmid pEGFP was from Clontech (Mountain View, CA, USA) (cat #: HLP309). Hoechst 33342 (20 mM solution; cat #: 62249) and ATP (10 mM solution; cat #: PV3227) were from Thermo Fisher Scientific (Waltham, MA, USA). Genomic RNA was isolated from influenza A/WSN/1933 strain as described previously [11 (link)].

Bulanova D., Ianevski A., Bugai A., Akimov Y., Kuivanen S., Paavilainen H., Kakkola L., Nandania J., Turunen L., Ohman T., Ala-Hongisto H., Pesonen H.M., Kuisma M.S., Honkimaa A., Walton E.L., Oksenych V., Lorey M.B., Guschin D., Shim J., Kim J., Than T.T., Chang S.Y., Hukkanen V., Kulesskiy E., Marjomaki V.S., Julkunen I., Nyman T.A., Matikainen S., Saarela J.S., Sane F., Hober D., Gabriel G., De Brabander J.K., Martikainen M., Windisch M.P., Min J.Y., Bruzzone R., Aittokallio T., Vähä-Koskela M., Vapalahti O., Pulk A., Velagapudi V, & Kainov D.E. (2017). Antiviral Properties of Chemical Inhibitors of Cellular Anti-Apoptotic Bcl-2 Proteins. Viruses, 9(10), 271.

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Colorectal Cancer Cell Line Cultivation

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The human CRC cell lines Colo205, HT29, CaCo2 and SW480 as well as the nontransformed human colon cell line CCD 841 CoN were purchased from ATCC (Manassas Virginia, USA). CRC cells were maintained in RPMI + GlutaMAX medium (Gibco, Karlsruhe, Germany) and CCD 841 CoN cells in DMEM medium (Gibco), in each case supplemented with 10% heat-inactivated fetal calf serum (PAA Laboratories, Cölbe, Germany) and 1% penicillin/streptomycin (PAA Laboratories) and cultured in a humidified atmosphere at 37 °C and 5% CO₂. Staurosporine and chemotherapeutic reagents 5-fluorouracil (5FU) and irinotecan were purchased from Sigma–Aldrich (Munich, Germany). Selective inhibitors ABT-199 (BCL-2 inhibitor), S63845 (MCL-1 inhibitor), and A-1331852 (orally available BCL-XL inhibitor, used for murine experiments) were purchased from Selleckchem (Munich, Germany) and WEHI-539 (BCL-XL inhibitor, used for in vitro and ex vivo experiments) was purchased from Hycultec (Beutelsbach, Deutschland).

Scherr A.L., Mock A., Gdynia G., Schmitt N., Heilig C.E., Korell F., Rhadakrishnan P., Hoffmeister P., Metzeler K.H., Schulze-Osthoff K., Illert A.L., Boerries M., Trojan J., Waidmann O., Falkenhorst J., Siveke J., Jost P.J., Bitzer M., Malek N.P., Vecchione L., Jelas I., Brors B., Glimm H., Stenzinger A., Grekova S.P., Gehrig T., Schulze-Bergkamen H., Jäger D., Schirmacher P., Heikenwalder M., Goeppert B., Schneider M., Fröhling S, & Köhler B.C. (2020). Identification of BCL-XL as highly active survival factor and promising therapeutic target in colorectal cancer. Cell Death & Disease, 11(10), 875.

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Antibody-Conjugate and Small-Molecule Study

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Polatuzumab vedotin and rituximab were obtained from Chugai Pharmaceutical Co., Ltd. A‐1331852, MK‐2206 2HCl and MMAE were purchased from Selleck Chemicals. Human IgG (HuIgG) was purchased from MP Biomedicals. Verapamil hydrochloride was obtained from Sigma‐Aldrich. U0126 was purchased from Promega. Anti‐human CD79b antibody (SN8) was purchased from BD Biosciences.

Kawasaki N., Nishito Y., Yoshimura Y, & Yoshiura S. (2022). The molecular rationale for the combination of polatuzumab vedotin plus rituximab in diffuse large B‐cell lymphoma. British Journal of Haematology, 199(2), 245-255.

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Comprehensive Antibody Source Protocol

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Pevonedistat was provided by Millennium Pharmaceuticals, Inc. (Cambridge, MA), a wholly owned subsidiary of Takeda Pharmaceutical Company Limited. Venetoclax, Selinexor, A‐1331852, and Ibrutinib were obtained by Selleck Chemicals (Houston, TX). All other drugs were provided by the RPCCC Pharmacy.
Primary mouse anti‐human antibodies raised against Bak and Bax were obtained from Sigma Chemicals (St. Louis, MO), Mcl‐1 from Santa Cruz Biotechnology (Santa Barbara, CA). Primary mouse/rabbit anti‐human antibodies raised against Bid, Bim, Noxa, Beclin‐1, p65, Bcl‐2, Bcl‐XL, Puma, PARP‐1, cleaved PARP‐1 and actin were obtained from Cell Signaling (Danvers, MA). Alkaline phosphatase (AP) or horseradish peroxidase (HRP) conjugated anti‐mouse/rabbit secondary antibodies were purchased from Jackson ImmunoResearch (West Grove, PA).
Sodium ⁵¹chromate (⁵¹Cr) (Perkin‐Elmer Life Inc., Boston MA) was used in functional assays assessing antibody‐mediated complement cytotoxicity (CMC) or antibody dependent cellular cytotoxicity (ADCC). Triton X‐100, trypan blue and histopaque‐1077 were obtained from Sigma‐Aldrich Inc. (St Louis, MO). Cell Titer‐Glo Luminescent Viability Assay reagent was purchased from Promega (Madison, WI).

Torka P., Mavis C., Kothari S., Belliotti S., Gu J., Sundaram S., Barth M, & Hernandez‐Ilizaliturri F.J. (2020). Pevonedistat, a NEDD8‐activating enzyme inhibitor, induces apoptosis and augments efficacy of chemotherapy and small molecule inhibitors in pre‐clinical models of diffuse large B‐cell lymphoma. EJHaem, 1(1), 122-132.

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Evaluating Cell Death Mechanisms

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Cells were treated with indicated concentrations of ABT-199 (Selleck Chemicals, Houston, TX), A1331852 (Selleck) or S63845 (Appexbio, Taiwan) prior to analysis of death by staining with propidium iodide and microscopic analysis using ImageXpress (Molecular Devices, Biberach, Germany). A representative image of this analysis is displayed in Supplementary Fig. 1. Alternatively, cell death was investigated by flow cytometric analysis of FSC/SSC (forward/side scatter) properties using FACScanto II (BD Bioscience, Heidelberg, Germany). Viability was assessed using CellTiterGlo (Promega, Mannheim, Germany) and Tecan Infinite M200 plate reader. Loss of mitochondrial membrane potential (MMP) was investigated following staining with 100 nM TMRM (tetramethylrhodamine methyl ester) (Thermo Fisher, Waltham, MA) and flow cytometry. To inhibit caspases, the broad-range caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (zVAD.fmk) (Bachem, Heidelberg, Germany) was used.

Bierbrauer A., Jacob M., Vogler M, & Fulda S. (2020). A direct comparison of selective BH3-mimetics reveals BCL-XL, BCL-2 and MCL-1 as promising therapeutic targets in neuroblastoma. British Journal of Cancer, 122(10), 1544-1551.

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