Human glioma cell lines, U-118, U-251; colon cancer cell line DLD-1; lung cancer cell lines H-1299 and H460 and breast cancer cell lines Hs 578T and MDA-MB-251 were obtained from the American Type Culture Collection (Manassas, VA, USA). U-118 is a permanent cell line derived from a grade IV human glioblastoma—astrocytoma, and U-251 was derived from a grade III–IV human malignant glioblastoma multiforme. DLD-1 was derived from a Dukes’ type C, colorectal adenocarcinoma, H-1299 is an NSCLC cell line, H460 is a large cell lung cancer cell line, and Hs 578T and MDA-MB-231 are triple-negative breast cancer (TNBC) cell lines. All human cell lines used in the study were authenticated by STR DNA analysis (DDC Medical, Fairfield, OH, USA; Center for Applied Genomics, SickKids, Toronto, Canada). Mouse 4T1 mammary tumour cells were obtained from Dr Jean Marshall (Dalhousie University, Halifax, Canada). All cell lines were maintained in high glucose Dulbecco’s modified Eagle’s medium (DMEM) (Invitrogen, ON, Canada), supplemented with 10% foetal bovine serum (FBS) (Corning, VA, USA) and 1:100 Antibiotic–Antimycotic (×100; ThermoFisher Scientific). MEK inhibitors U0126 and selumetinib were purchased from Cell Signaling Technology (Danvers, MA, USA) and Selleckchem (Houston, TX, USA), respectively; and 5-Aminolevulinic acid from Sigma (Oakville, ON, Canada).
5 aminolevulinic acid
Manufactured by Merck Group
Sourced in United States
5-aminolevulinic acid is a chemical compound used in laboratory settings. It is a precursor in the biosynthesis of various important molecules, including heme, chlorophyll, and other tetrapyrroles. The compound serves as a useful tool for researchers studying these biological processes.
Automatically generated - may contain errors
Lab products found in correlation
U0126, by Cell Signaling Technology (3 mentions)
Dulbecco s modified eagle s medium dmem, by Thermo Fisher Scientific (2 mentions)
Hs578t, by American Type Culture Collection (2 mentions)
Dld 1, by American Type Culture Collection (2 mentions)
Gw274150, by GlaxoSmithKline (2 mentions)
Restriction enzyme, by New England Biolabs (2 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (2 mentions)
L glutamine, by Merck Group (2 mentions)
Trypan blue, by Merck Group (2 mentions)
Streptomycin, by Solarbio (2 mentions)
Antibiotic antimycotic 100, by Thermo Fisher Scientific (1 mentions)
H1299, by American Type Culture Collection (1 mentions)
Foetal bovine serum fbs, by Corning (1 mentions)
Control igg, by Cell Signaling Technology (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
N ethylmaleimide, by Merck Group (1 mentions)
L alanine, by Merck Group (1 mentions)
Sirt1, by Cell Signaling Technology (1 mentions)
L histidine, by Merck Group (1 mentions)
β actin, by Cell Signaling Technology (1 mentions)
18 protocols using 5 aminolevulinic acid
1
Comprehensive Cancer Cell Line Protocol
2
Inhibition of iNOS and Akt Signaling Pathways
3
Bacterial Oxygen Sensing Behavior
4
Cell Authentication and Reagent Provenance
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Human cancer and normal cells used in this study were obtained from the American Type Culture Collection (Manassas, VA). Mouse 4T1 mammary tumor cells were obtained from Dr. Jean Marshall (Dalhousie University, Halifax, Canada). All human cancer cell lines used in this study were authenticated by STR DNA analysis (DDC Medical (Fairfield, OH)) or by the Center for Applied Genomics (SickKids, Toronto, Canada). U0126 was purchased from Cell Signaling Technology (Danvers, MA), PD98059, LY294002, SB203580 and SP600125 from Calbiochem (La Jolla, CA), Selumetinib from Selleckchem (Houston, TX) and 5-Aminolevulinic acid from Sigma (Oakville, ON).
5
Expression and Purification of PAS Mutants
6
Mitochondrial Activity and Protoporphyrin IX Imaging
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In parallel to the MitoTracker staining of living cells, CD14+ monocytes isolated from the peripheral blood of the study participants were incubated with 100 mg/L of 5-aminolevulinic acid (5-ALA) (Sigma-Aldrich, St. Louis, MO, USA) for 4 h at 37 °C in 5% CO2. Thirty minutes prior to the microscopic examination, 100 nM of MitoTracker Orange CMTMRos was added to the cells. Images were taken under the LSM-710 laser scanning microscope (Zeiss, Germany), equipped with a 20× Plan-Apochromat NA 0.8 objective. Cells were placed in a glass-bottom plate for microscopy. The fluorescent signals of the MitoTracker dye were recorded at wavelengths of 561 nm (excitation) and 580–620 nm (emission). The fluorescent signals of 5-ALA-induced protoporphyrin IX (PpIX) were recorded at the wavelengths of 633 nm (excitation) and 650–720 nm (emission). As a result, an overlay of the transmitted light image with fluorescent images of the MitoTracker distribution and PpIX was obtained.
7
Bacterial Strains and Growth Conditions
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The Escherichia coli strains TG1 and CC118 λ pir were used for plasmid maintenance. The E. coli ST18 strain [31 (link)] was obtained from the German Collection of Microorganisms and Cell Cultures (DSM 22074) and used for conjugal transfer of plasmids into S. proteamaculans 94 [32 (link)]. E. coli and S. proteamaculans strains were grown on Luria broth (LB) supplemented with 100 µg/mL ampicillin, 50 μg/mL kanamycin or 100 µg/mL streptomycin as needed. When E. coli ST18 was cultured, the medium contained 50 μg/mL 5-aminolevulinic acid (Sigma Aldrich, Darmstadt, Germany). To plot S. proteamaculans strains’ growth curves, overnight cultures were diluted with LB to OD600 of 0.01. The bacterial suspensions (100 μL) were transferred to a 96-well plate and cultivated at 30 °C and 200 rpm using a microplate reader, Infinite M200 Pro (Tecan, Switzerland). The optical density was read at a wavelength of 600 nm. E. coli AB1157 (thr-1 leuB6 ara14 proA2 hisG4 argE3 thi-1 supE44 rpsL31) from R. Devoret’s collection was co-transformed with pProPlnHis6 plasmid, which contains the protealysin gene under control of the T7 promoter [20 (link),33 (link)] and the pT7POL26 construct for T7 promoter-driven protein expression from Cox M.M. The presence of both plasmids was controlled by adding a mixture of ampicillin and kanamycin.
8
Synthesis and Characterization of Silver Nanoparticles
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Porcine pancreatic α-amylase (PPA), 5-aminolevulinic acid (ALA), Rhodamine 6G (R6G), acetylacetone, formaldehyde and Dulbecco’s Modified Eagle Medium (DMEM) were purchased from Sigma-Aldrich. SM (PEG)12 crosslinkers and Slide-A-Lyzer™ Dialysis Cassettes (MWCO: 10 KDa) were purchased from Thermo Fisher Scientific Co., Ltd. Silver nitrate (AgNO3), sodium hydroxide (NaOH), sodium borohydride (NaBH4) and dimethyl sulfoxide (DMSO) were purchased from Sinopharm Chemical Reagent Co., Ltd. All chemicals were used as received without further purification. All solutions were prepared using purified water with resistivity of 18 mΩ cm−1 as the solvent. The water was deoxygenated before use with a nitrogen purge for 1 h at the flow rate of 25 mL/s.
9
Cholesterol Oxidation Enzymatic Assay
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All solvents and reagents were of the highest purity commercially available, unless noted otherwise. Isopropyl-β-d -thiogalactopyranoside (IPTG), acrylamide, N,N′-methylenebisacrylamide, ammonium persulfate, 30% acrylamide, TEMED, glycine, Tris, DTT, ampicillin sodium, kanamycin sulfate, chloramphenicol, hydroxypropyl NADH, NADPH, glucose-6-phosphate, glucose-6-phosphate dehydrogenase, catalase and ammonium sulfate were all purchased from Sangon Biotech (Shanghai, China). Cholesterol, Cholesterol propionate, NADPH, 5-aminolevulinic acid, spinach ferredoxin (spFDX) and spinach ferredoxin reductase (spFDR) were obtained from Sigma-Aldrich (St. Louis, MO, USA). 4-Cholesten-3-one and methyl-β-cyclodextrin were purchased from J&K Scientific Ltd. (Beijing, China). Taq DNA polymerase, PrimeSTAR HS DNA Polymerase (with GC buffer), T4 DNA ligase, restriction endonucleases Nde I and Xho I, T-A cloning kits and DNA MW markers were purchased from Takara (Dalian, China). Cholesterol sodium sulfate was purchased from Ark Pharm Inc. (Libertyville, IL, USA).
10
Characterization of Cancer Cell Lines
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NIH3T3 cells were obtained from the American Type Culture Collection (ATCC; Manassas, VA, USA). H-Ras-transformed NIH3T3 cells were generated in-house and was described previously40 (link),41 (link). Human colon cancer cell line DLD-1, glioblastoma cell line SNB-75, and breast cancer cell lines Hs 578 T and MDA MB 231 were obtained from ATCC and were authenticated by STR DNA analysis (SickKids, Toronto). All cells were maintained in high glucose Dulbecco's modified Eagle's medium (DMEM) (Invitrogen, Ontario, Canada), supplemented with 10% fetal bovine serum (FBS) and antibiotic–antimycotic mixture (Invitrogen) (100 units/mL penicillin G sodium) at 37 °C and 5% CO2.
U0126 was purchased from Cell Signaling Technology (Danvers, MA); 5-Aminolevulinic acid from Sigma (Oakville, ON); HIF-1α inhibitor (methyl 3-[[2-[4-(2-adamantyl)phenoxy]acetyl]amino]-4-hydroxybenzoate) from Santa Cruz Biotechnology (Dallas, TX); ABCB1 inhibitor, zosuquidar from Selleckchem, and pan-RSK inhibitor, SL0101 from Calbiochem (Darmstadt, Germany). Anti-phospho-ERK-1/2 and anti-phospho RSK antibodies were purchased from Cell Signaling (Danvers, MA), anti-ABCB1 antibody from Alomone Labs (Israel), anti-FECH, anti-phospho-S6, anti-RSK2, anti-total ERK antibodies, and the FITC-tagged Anti-ABCB1 antibody from Santa Cruz Biotechnology; anti-HIF-1α antibody, and anti-GAPDH antibody were purchased from Abcam (US).
U0126 was purchased from Cell Signaling Technology (Danvers, MA); 5-Aminolevulinic acid from Sigma (Oakville, ON); HIF-1α inhibitor (methyl 3-[[2-[4-(2-adamantyl)phenoxy]acetyl]amino]-4-hydroxybenzoate) from Santa Cruz Biotechnology (Dallas, TX); ABCB1 inhibitor, zosuquidar from Selleckchem, and pan-RSK inhibitor, SL0101 from Calbiochem (Darmstadt, Germany). Anti-phospho-ERK-1/2 and anti-phospho RSK antibodies were purchased from Cell Signaling (Danvers, MA), anti-ABCB1 antibody from Alomone Labs (Israel), anti-FECH, anti-phospho-S6, anti-RSK2, anti-total ERK antibodies, and the FITC-tagged Anti-ABCB1 antibody from Santa Cruz Biotechnology; anti-HIF-1α antibody, and anti-GAPDH antibody were purchased from Abcam (US).
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