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Lhc 8 media

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LHC-8 media is a laboratory culture medium used for the growth and cultivation of various microorganisms, including bacteria and fungi. It provides the necessary nutrients and growth factors required for the proliferation of these microbial species in a controlled laboratory environment.

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Lab products found in correlation

Trypsin edta, by Thermo Fisher Scientific (3 mentions) Tobramycin, by Merck Group (2 mentions) Antibiotic antimycotic, by Thermo Fisher Scientific (2 mentions) Gelatin, by Merck Group (2 mentions) Phosphate buffered saline (pbs), by Thermo Fisher Scientific (2 mentions) Lipofectamine transfection reagent, by Thermo Fisher Scientific (1 mentions) Luciferase assay kit, by Thermo Fisher Scientific (1 mentions) Pmir report luciferase reporter vector, by Thermo Fisher Scientific (1 mentions) Dulbecco s modified eagle media, by Thermo Fisher Scientific (1 mentions) Cultureware, by Thermo Fisher Scientific (1 mentions) Lipofectamine 300, by Thermo Fisher Scientific (1 mentions) Dimethyl sulfoxide (dmso), by Merck Group (1 mentions) α naphthoflavone, by Merck Group (1 mentions) B α p, by Merck Group (1 mentions) 4 abp, by Merck Group (1 mentions) Trypsin, by Thermo Fisher Scientific (1 mentions) Methanol, by Merck Group (1 mentions) Acetyl coa, by Merck Group (1 mentions) Acetonitrile, by Merck Group (1 mentions) Sodium perchlorate, by Merck Group (1 mentions)

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LHC-8 (6 citations)

6 protocols using lhc 8 media

1

Molecular Assays for CFTR Expression

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LHC-8 media, Dulbecco’s Modified Eagle Media, Trypsin-EDTA (0.05%), puromycin and lipofectamine transfection reagents were purchased from Invitrogen (Life Technologies, Carlsbad, CA, USA). The miRVana kit for RNA isolation, miR assays, miR mimics, pMIR-Report luciferase reporter vector, luciferase assay kits and Taqman quantitative PCR (qPCR) reagents were purchased from Applied Biosystems (Life Technologies, Foster City, CA, USA). The C-terminal-specific CFTR antibody was purchased from R&D Systems (Minneapolis, MN, USA).
Kumar P., Bhattacharyya S., Peters K., Glover M., Sen A., Cox R., Kundu S., Caohuy H., Frizzell R., Pollard H, & Biswas R. (2015). miR-16 rescues F508del-CFTR function in native cystic fibrosis epithelial cells. Gene therapy, 22(11), 908-916.
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2

CFBE and 16HBE Cell Culture Protocol

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CFBE cells (CFBE41o-) and human bronchial epithelial cells (16HBE14o-)38 (link) were grown with LHC-8 media (Invitrogen) with 10% FBS, 1X antibiotic antimycotic (Gibco), and tobramycin 40 mg per 500 mL (Sigma). Once grown to confluence, cells were trypsinized by first washing with 0.05% trypsin, then adding 0.25% trypsin for 5 minutes, and harvesting with RPMI medium with 10% FBS. Cells were frozen in 5% DMSO in culture medium as necessary. Nanoparticles were resuspended in culture media by vigorous vortexing and water sonication, then added directly to cells at concentrations of 2 mg/mL/1×10^6 cells (corresponding to approximately 10^9 PNA/DNA molecules delivered to each cell assuming 100% efficiency).
To test primers, a 712 base pair region of the CFTR gene, with either the F508DEL or corrected sequence (including our silent modifications), was cloned into plasmids. PCR reactions were first tested on plasmids. Gradient and step-down PCR at varying conditions was performed to ensure that F508del primers only amplified the F508del plasmid, and the donor-specific primers only amplified the donor-sequence-containing plasmids. Primers are included in Supplementary Table 2.
McNeer N.A., Anandalingam K., Fields R.J., Caputo C., Kopic S., Gupta A., Quijano E., Polikoff L., Kong Y., Bahal R., Geibel J.P., Glazer P.M., Saltzman W.M, & Egan M.E. (2015). Correction of F508del CFTR in airway epithelium using nanoparticles delivering triplex-forming PNAs. Nature communications, 6, 6952.
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3

CFBE and 16HBE Cell Culture Protocol

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CFBE cells (CFBE41o-) and human bronchial epithelial cells (16HBE14o-)38 (link) were grown with LHC-8 media (Invitrogen) with 10% FBS, 1X antibiotic antimycotic (Gibco), and tobramycin 40 mg per 500 mL (Sigma). Once grown to confluence, cells were trypsinized by first washing with 0.05% trypsin, then adding 0.25% trypsin for 5 minutes, and harvesting with RPMI medium with 10% FBS. Cells were frozen in 5% DMSO in culture medium as necessary. Nanoparticles were resuspended in culture media by vigorous vortexing and water sonication, then added directly to cells at concentrations of 2 mg/mL/1×10^6 cells (corresponding to approximately 10^9 PNA/DNA molecules delivered to each cell assuming 100% efficiency).
To test primers, a 712 base pair region of the CFTR gene, with either the F508DEL or corrected sequence (including our silent modifications), was cloned into plasmids. PCR reactions were first tested on plasmids. Gradient and step-down PCR at varying conditions was performed to ensure that F508del primers only amplified the F508del plasmid, and the donor-specific primers only amplified the donor-sequence-containing plasmids. Primers are included in Supplementary Table 2.
McNeer N.A., Anandalingam K., Fields R.J., Caputo C., Kopic S., Gupta A., Quijano E., Polikoff L., Kong Y., Bahal R., Geibel J.P., Glazer P.M., Saltzman W.M, & Egan M.E. (2015). Correction of F508del CFTR in airway epithelium using nanoparticles delivering triplex-forming PNAs. Nature communications, 6, 6952.
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4

Cytochrome P450 Induction Assay

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Sodium chromate, α-naphthoflavone, benzo[a]pyrene (B[a]P), DMSO, para-aminobenzoic acid (PABA), sulfamethazine (SMZ), 4-ABP, BNA, 7-ethoxy-resorufin, resorufin, gelatin, and glucose were obtained from Millipore-Sigma (St. Louis, MO, USA). Trypsin/EDTA, PBS, LHC-8 media, lipofectamine™ 300, and cultureware were purchased from Thermo Fisher Scientific (Carlsbad, CA).
Wise J.T., Salazar-González R.A., Walls K.M., Doll M.A., Habil M.R, & Hein D.W. (2022). Hexavalent Chromium Increases the Metabolism and Genotoxicity of Aromatic Amine Carcinogens 4-Aminobiphenyl and β-Naphthylamine in Immortalized Human Lung Epithelial Cells. Toxicology and applied pharmacology, 449, 116095.
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5

Bronchial Epithelial Cell Culture Protocol

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Deoxyguanosine, acetyl-CoA, β-naphthylamine (BNA), 4-aminobiphenyl
(4-ABP), para-aminobenzoic acid, gelatin, sulfamethazine, sodium perchlorate,
sodium phosphate, methanol, and acetonitrile were purchased from MilliporeSigma
(Burlington, MA). Cell culture supplies, trypsin, phosphate buffered saline
(PBS), and LHC-8 media were purchased from ThermoFisher (Waltham, MA).
N-hydroxy-4-aminobiphenyl and dG-C-8-ABP adduct standard
were purchased from Toronto Research Chemicals (North York, ON, Canada) BEP2D
cells an E6/E7 HPV immortalized bronchial epithelial airway cell line were
generously gifted by Curtis Harris at the NIH and were cultured in LHC-8 [20 (link)]. HBEC2-KT cells a cdk24 and hTERT
immortalized bronchial epithelial airway cell line were used in collaboration
with John P. Wise Sr. at University of Louisville [21 (link)].
Wise J.T., Salazar-González R.A., Habil M.R., Doll M.A, & Hein D.W. (2022). Expression of Arylamine N-Acetyltransferase 2 Activity in Immortalized Human Bronchial Epithelial Cells. Toxicology and applied pharmacology, 442, 115993.
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6

Cell Line Culture Protocols

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All cell lines were purchased from the American Type Culture Collection (ATCC) (except HMLE and BxPC-3) and all tested negative for mycoplasma. A549 (CCL-185, male), NMuMG (CRL-1636, female) and MCF-7 (HTB-22, female) cells were supplemented with Dulbecco’s Modified Eagle’s Media (DMEM) with 10% fetal bovine serum (Hyclone). BEAS-2B (CRL-9609, male) were supplemented with LHC-8 media (Thermo-fisher scientific) with and without FBS (Hyclone, Canada, characterized). Panc 10.05 (CRL-2547, male) and BxPC-3 cells (CRL-1687, female), a generous gift from Dr. David Hoskin, Dalhousie University, were supplemented with Roswell Park Memorial Institute (RPMI) with 10% fetal bovine serum. HMLE (human mammary epithelial cell line, female) cells were a generous gift from Dr. Robert Weinberg (Whitehead Institute for Biomedical Research, Cambridge, Massachusetts) and were cultured in a 1:1 ratio of DMEM F12 1:1 and mammary epithelial cell growth medium (MEGM, Lonza) supplemented with 13 µg/mL bovine pituitary extract, 20 µg/mL human epidermal growth factor,10 µg/mL insulin, 1 µg/mL gentamicin/amphotericin and 2 µg/mL hydrocortisone (Lonza Clonetics) and 10% FBS. All cells were cultured in the presence of 1% pencillin/streptomycin (Hyclone) and were maintained at 37 °C with 5% CO2.
Bydoun M., Sterea A., Weaver I.C., Bharadwaj A.G, & Waisman D.M. (2018). A novel mechanism of plasminogen activation in epithelial and mesenchymal cells. Scientific Reports, 8, 14091.
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