Hs27a

Manufactured by American Type Culture Collection

Sourced in United States

The HS27A is a laboratory centrifuge designed for general-purpose applications. It features a maximum speed of 4,500 RPM and can accommodate a variety of sample tubes and microplates. The centrifuge is equipped with a digital display that provides information on speed, time, and other operational parameters.

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19 protocols using hs27a

Co-culture Assay for MCL Cell Lines

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The human MCL cell lines Granta-519, Jeko-1, Maver-1, Mino, and Z138 were obtained from DSMZ (Braunschweig, Germany) and cultured in Iscove’s modified Dulbecco’s medium (IMDM; Invitrogen Life Technologies, Carlsbad, CA, USA). The stromal cell line HS-27a and the kidney cell line HEK-293T/17 were obtained from ATCC (Manassas, VA, USA), and cultured in Dulbecco’s Modified Eagle’s Medium (DMEM; Invitrogen).
For co-culture assays, bone-marrow stromal cells were seeded in 96-well plates 4 hours (h) prior to the addition of MCL cells, to allow cell attachment. MCL cells were pre-incubated for 2 h with stromal cells, followed by a 3-day co-culture in the presence of the indicated drug concentrations. Cell viability and specific cell death were measured by flow cytometry.
The following small-molecule inhibitors were used: venetoclax/ABT-199, Q-VD-OPh, IPTG (MedChemExpress, Princeton, NJ, USA), silmitasertib/CX-4945, S63845, MK2206 (Selleckchem, Houston, TX, USA), cycloheximide, puromycin (Sigma-Aldrich, St. Louis, MO, USA), and blasticidin (Thermo Fisher Scientific, Waltham, MA, USA).

Thus Y.J., de Rooij M.F., Swier N., Beijersbergen R.L., Guikema J.E., Kersten M.J., Eldering E., Pals S.T., Kater A.P, & Spaargaren M. (2022). Inhibition of casein kinase 2 sensitizes mantle cell lymphoma to venetoclax through MCL-1 downregulation. Haematologica, 108(3), 797-810.

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Characterization of Multiple Myeloma Cell Lines

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AMO1 WT, AMO1-BtzR, AMO1-CfzR were kind gifts from Christophe Driessen. U266, JIM-3, KMS12BM, MMM1, LP-1, ANBL6-BtzR, U266-BtzR, RPMI-8226 BtzR, KMM1, KMS18, MM1144 were kind gifts from Brian Van Ness. KMS11 and KMS34 were obtained from the Japanese Collection of Research Bioresources Cell Bank. JJN3, CMK, L363, RPMI-8226, MM.1S, HS27a, and HS5 cells were obtained from ATCC. Cell lines were maintained in RPMI-1640 medium with 10% FBS (Gemini Benchmark). IL-6 dependent cell lines were cultured in the presence of 50 ng/mL recombinant human IL-6 (ProSpec). Proteasome inhibitor resistant cell lines were cultured in 90nM Bortezomib or Carfilzomib as previously described (Mitra et al., 2017 (link); Mitra et al., 2016 (link); Soriano et al., 2016 (link)). Cell lines were authenticated by DNA short tandem repeat profiling at ATCC.

Ferguson I.D., Lin Y.H., Lam C., Shao H., Tharp K.M., Hale M., Kasap C., Mariano M.C., Kishishita A., Escobar B.P., Mandal K., Steri V., Wang D., Phojanakong P., Tuomivaara S.T., Hann B., Driessen C., Van Ness B., Gestwicki J.E, & Wiita A.P. (2022). Allosteric HSP70 inhibitors perturb mitochondrial proteostasis and overcome proteasome inhibitor resistance in multiple myeloma. Cell chemical biology, 29(8), 1288-1302.e7.

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HIV-1 Latent Infection in T Cell and Stromal Lines

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J1.1 and ACH-2, derivatives of Jurkat and A3.01 T cell lines in which the HIV-1 LAV strain was latently infected, respectively [23 (link),24 (link)], were obtained from the NIH AIDS Reagent Program and were grown in RPMI1640 medium supplemented with 10% fetal bovine serum (FBS). The parental Jurkat and A3.01 cell lines were obtained from American Type Culture Collection (ATCC) and were similarly grown in RPMI1640 medium. EA.hy926, a human umbilical vein endothelial cell (HUVEC)-derived endothelial cell line [25 (link)], and HS-5, a bone-marrow-derived stromal cell line [26 (link)], were from ATCC and were grown in Dulbecco’s modified Eagle’s medium (DMEM) with 10% FBS. HS-27A, another bone-marrow-derived stromal cell line (ATCC), was grown in RPMI1640 with 10% FBS. Primary FRC (ScienCell Research Laboratories) were grown in fibroblast medium (ScienCell Research Laboratories) supplemented with 1% fibroblast growth supplement and 2% FBS.

Tanabe R, & Morikawa Y. (2021). Efficient Transendothelial Migration of Latently HIV-1-Infected Cells. Viruses, 13(8), 1589.

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Mesenchymal Stem Cell Culture Protocol

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MSCs cultures were derived from de-identified bone marrow specimens from patients at the West Virginia University Cancer Institute and cultured as previously described [1 (link)]. Patients had no history of exposure to chemotherapy, radiation, or malignancy. HS-5 and HS-27A cell lines were purchased from ATCC (Manassas, VA, USA) and cultured as suggested by the supplier.

Hare I., Evans R., Fortney J., Moses B., Piktel D., Slone W, & Gibson L.F. (2016). Chemotherapy-induced Dkk-1 expression by primary human mesenchymal stem cells is p53 dependent. Medical oncology (Northwood, London, England), 33(10), 113.

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3D Co-culture of Prostate Cell Lines

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Human prostate cancer epithelial cell lines, LNCaP, C4-2, PC3, PC3M, and DU145, and a human bone stromal cell line HS27A (ATCC, Manassas, VA) that were used in our previous studies^{61 (link)–63 (link)} were maintained in T-medium (Invitrogen, Carlsbad, CA) with 5% fetal bovine serum (FBS; Invitrogen). Then, the 3D coculture of HS27A cells mixed with equal numbers of LNCaP or C4-2 cells or a monoculture was performed in an RCCS system (Synthecon, Houston, TX) following an established protocol^{21 (link)}. Each condition was performed in duplicate vessels for three independent experiments. Previously established CAFs and BAFs, derived from cancerous and benign/normal regions of the prostate gland, respectively^{21 (link)}, were confirmed as fibroblasts by positive vimentin but negative cytokeratin expression (Supplemental Fig. s6) and were maintained in PrEGM™ Prostate Epithelial Cell Growth Medium (Lonza, Walkersville, MD). All cells were cultured in a 37 °C incubator with 5% CO₂.

Hsieh C.L., Liu C.M., Chen H.A., Yang S.T., Shigemura K., Kitagawa K., Yamamichi F., Fujisawa M., Liu Y.R., Lee W.H., Chen K.C., Shen C.N., Lin C.C., Chung L.W, & Sung S.Y. (2017). Reactive oxygen species–mediated switching expression of MMP-3 in stromal fibroblasts and cancer cells during prostate cancer progression. Scientific Reports, 7, 9065.

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ATRA and ATO Treatment Protocols for Leukemia Cell Lines

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All cell cultures were maintained in a humidified atmosphere at 37 °C with 5% CO₂. NB4 (all-trans retinoic acid, ATRA-sensitive) and NB4 R2 (ATRA-resistant) cell lines were kindly provided by Dr. Pier Paolo Pandolfi (Harvard Medical School, Boston, MA, USA), and maintained in RPMI 1640 (Gibco, Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (FBS) (Gibco), L-glutamine (2 mM), and penicillin/streptomycin (Invitrogen, Carlsbad, CA, USA). Mycoplasma contamination was routinely tested. All leukemia cell lines were tested by short tandem repeat analysis. The HS5 (ATCC, CRL-11882), HS27A (ATCC, CRL-2496), and HEK293T (ATCC, CRL-3216) cell lines were obtained from American Type Culture Collection and grown in Dulbecco’s modified Eagle medium with 10% FBS. ATRA and arsenic trioxide (ATO) were obtained from Sigma-Aldrich (St. Louis, MO, USA) and used at a concentration of 1 µM for all in vitro assays. Cytarabine (Ara-C) was obtained from Blau Pharmaceutics (Blau Pharmaceutics, Sao Paulo, Brazil). SLIT2 peptide was obtained from PeproTech (produced in 293T cells—PeproTech EC, Ltd. London, UK) and used at the concentration of 50 ng/mL for all in vitro assays.

Weinhäuser I., Pereira-Martins D.A., Ortiz C., Silveira D.R., Simões L.A., Bianco T.M., Araujo C.L., Koury L.C., Melo R.A., Bittencourt R.I., Pagnano K., Pasquini R., Nunes E.C., Fagundes E.M., Gloria A.B., Kerbauy F., Chauffaille M.D., Keating A., Tallman M.S., Ribeiro R.C., Dillon R., Ganser A., Löwenberg B., Valk P., Lo-Coco F., Sanz M.A., Berliner N., Ammatuna E., Lucena-Araujo A.R., Schuringa J.J, & Rego E.M. (2020). Reduced SLIT2 is Associated with Increased Cell Proliferation and Arsenic Trioxide Resistance in Acute Promyelocytic Leukemia. Cancers, 12(11), 3134.

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Myeloma Cell Line Cultivation Protocol

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The human cell lines (MM.1S, RPMI-8226, OPM-2, LP-1, OPM-1, JJN3, L363, AMO-1, OCI-My5, KMS-20, KMS-26, KMS-27) were obtained from ATCC, JCRB, or DSMZ. MM cell lines OPM-2 and KMS11 expressing non-targeting control shRNA or shRNA.CRBN#13 (Zhu et al., 2011 (link), 2013 (link)) were a gift from Keith Stewart (Mayo Clinic, AZ, USA). MM.1S CRBN^−/−cells, generated by CRISPR-knockout (Lu et al., 2014 (link)) were kindly provided by the laboratory of Dr William Kaelin (DFCI, Boston, MA, USA). The MM.1S-Cas9+ cells were generated by the laboratory of Dr Benjamin Ebert (DFCI). KMS11-Cas9+ cells (transduced with pLX 311-Cas9 [Addgene #96924] construct) were obtained from the Broad Institute. The human stromal cell line HS27A was obtained from ATCC. All human MM cell lines were cultured in RPMI 1640 medium supplemented with L-glutamine (Life Technologies, Carlsbad, CA, USA), FBS (10%) (Gemini Bioproducts, Woodland, CA), 20 I.U./mL penicillin and 20μg/mL streptomycin (Fisher Scientific, Springfield, NJ, USA) and cultured at 37°C, with 5% CO2.

Shirasaki R., Matthews G.M., Gandolfi S., de Matos Simoes R., Buckley D.L., Vora J.R., Sievers Q.L., Brüggenthies J.B., Dashevsky O., Poarch H., Tang H., Bariteau M.A., Sheffer M., Hu Y., Downey-Kopyscinski S.L., Hengeveld P.J., Glassner B.J., Dhimolea E., Ott C.J., Zhang T., Kwiatkowski N.P., Laubach J.P., Schlossman R.L., Richardson P.G., Culhane A.C., Groen R.W., Fischer E.S., Vazquez F., Tsherniak A., Hahn W.C., Levy J., Auclair D., Licht J.D., Keats J.J., Boise L.H., Ebert B.L., Bradner J.E., Gray N.S, & Mitsiades C.S. (2021). Functional Genomics Identify Distinct and Overlapping Genes Mediating Resistance to Different Classes of Heterobifunctional Degraders of Oncoproteins. Cell reports, 34(1), 108532.

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Characterization of Cancer and Non-Malignant Cell Lines

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Cancer cell lines (MCF7, T47D, ZR75–1 and LNCaP) and immortalized non-malignant cells lines HS5, HS27A, HOBIT, hFOB, THLE3 and SVGP12 were originally obtained from ATCC. Lenti-X-293T cells were purchased from Clontech (Mountain View, CA). The MCF7 cells stably expressing doxycycline-inducible mutant ER (Y537S) and the respective WT control were established by the lab of Dr. Myles Brown, as previously described (27 (link)). The cells were used for up to 15 passages after thawing. The identity of the cell lines was confirmed by short tandem repeat (STR) analysis. The cells were last tested for the presence of mycoplasma in May 2019, using the MycoAlert mycoplasma detection kit (Lonza). The culture conditions and reagents used in this study are further described in the Supplemental Materials.

Dhimolea E., de Matos Simoes R., Kansara D., Weng X., Sharma S., Awate P., Liu Z., Gao D., Mitsiades N., Schwab J.H., Chen Y., Jeselsohn R., Culhane A.C., Brown M., Georgakoudi I, & Mitsiades C.S. (2020). Pleiotropic mechanisms drive endocrine resistance in the three-dimensional bone microenvironment. Cancer research, 81(2), 371-383.

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Culturing Human Cell Lines for Cancer Research

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Human bone marrow stromal cells (HS27a), human dermal microvascular endothelial cells (HMEC-1), and the prostate cancer cell line MDA-PCa-2b were purchased from ATCC. HS27a were cultured in RPMI 1640 (Gibco, 22400-105) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (Atlanta Biologicals, S11150) and 1% penicillin-streptomycin (Gibco, 15140-122). HMEC-1 were cultured in MCDB131 (Gibco, 10372-019) supplemented with 10% heat-inactivated fetal bovine serum, 10 mM L-glutamine (Gibco, 25030-081), 10 ng/mL epidermal growth factor (EGF)(Gibco, PHG0311), and 1 μg/mL hydrocortisone (Sigma, H0135-1MG). MDA-PCa-2b were cultured in BRFF-HPC1 (AthenaES, NC9970798) supplemented with 20% FBS and 1% penicillin-streptomycin.

, & Jenkins C.G. (2000). The Internet and Healthcare.. Bulletin of the Medical Library Association, 88(1), 89-90.

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Immortalized Cell Line for DSCR4 Expression

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Cell culture The human papillomavirus 16 E6/ E7-transformed cell line HS-27A, an immortalized noncancerous cell type with epithelial morphology, was purchased from ATCC (CRL-2496) and cultured in RPMI-1640 medium (Invitrogen) supplemented with 10% fetal bovine serum (FBS) (Invitrogen) according to ATCC protocols.
Plasmid DNA vector and control vector construction The PTCN-DSCR4 expression vector (BC096162), which contains the full-length cDNA sequence of DSCR4 including upstream and downstream untranslated regions (UTRs), was purchased from transOMIC technologies (Supplementary Fig. S1A). No marker was added to DSCR4 cDNA, to ensure that the function of the extra copy of DSCR4 was not affected. Cytomegalovirusderived promoter and enhancer sequences placed before DSCR4 cDNA in the vector ensure proper expression in eukaryotic cells. For production of a control plasmid (PTCN-control), we removed the DSCR4 cDNA sequence along with UTR elements from PTCN-DSCR4 using a GeneArt Seamless Cloning and Assembly kit (Supplementary Fig. S1B). With DSCR4 cDNA being the only difference between PTCN-DSCR4 and PTCN-control, we sought to minimize the confounding elements in our differential gene expression analysis.

Saber M.M., Karimiavargani M., Uzawa T., Hettiarachchi N., Hamada M., Ito Y, & Saitou N. (2021). Possible roles for the hominoid-specific DSCR4 gene in human cells. Genes & genetic systems, 96(1).

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