The source information of 10 batches of PC extract used for the experiment is shown in Table 4 . Reference substances (purity ≥ 98%) including resveratrol, quercetin, emodin, physcion and emodin-8-β-D-glucoside were purchased from the National Institute for the Control of Pharmaceutical and Biological Products (NICPBP). Dimethyl sulfoxide (DMSO) was purchased from Beijing Solarbio Science & Technology Co., Ltd. (Beijing, China). TritonX-100 was bought from GBCBIO Technologies lnc. (Guangzhou, China). L-DOPA was purchased from Yingxin Laboratories (Shanghai, China). Trypsin, penicillin-streptomycin solution and DMEM were purchased from Cytiva (Marlborough, MA, USA). Fetal bovine serum (FBS) was bought from Zhejiang Tianhang Biological Technology Co. (Huzhou, China). The murine melanoma cell line B16F10 (C57BL/6) and RPMI-1640 complete medium were obtained from Procell Life Science & Technology Co., Ltd. (Wuhan, China). The SweScript RT II First Strand cDNA Synthesis Kit and 2xUniversal SYBR Green qPCR Master Mix were purchased from Wuhan Servicebio Technology Co., Ltd. (Wuhan, China).
Penicillin streptomycin solution
Manufactured by Cytiva
Sourced in United States
Penicillin-streptomycin solution is a sterile liquid media supplement used in cell culture applications. It contains the antibiotics penicillin and streptomycin, which help prevent bacterial contamination of cell cultures.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Cytiva (10 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (6 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (6 mentions)
Opti mem, by Thermo Fisher Scientific (3 mentions)
Cell counting kit 8 cck 8, by Dojindo Laboratories (3 mentions)
Phosphate buffered saline (pbs), by Cytiva (3 mentions)
Penicillin, by Cytiva (3 mentions)
Insulin, by Merck Group (3 mentions)
Fetal bovine serum (fbs), by Atlas Biologicals (3 mentions)
Dexamethasone (dex), by Merck Group (3 mentions)
Trypsin, by Cytiva (2 mentions)
Streptomycin, by Cytiva (2 mentions)
Rpmi 1640, by Cytiva (2 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions)
Rpmi 1640 medium, by Corning (2 mentions)
Dulbecco s modified eagle s medium, by Cytiva (2 mentions)
Dulbecco s modified eagle s medium dmem, by Cytiva (2 mentions)
L glutamine, by Cytiva (2 mentions)
Fetal bovine serum (fbs), by Atlanta Biologicals (2 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (2 mentions)
34 protocols using penicillin streptomycin solution
1
Phytochemical Analysis of PC Extract
2
Culturing Hepatocellular Carcinoma Cell Lines
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HepG2 (CoBioer Biosciences Co., Ltd.; cat. no. CBP60199), Hep3B (CoBioer Biosciences Co., Ltd.; cat. no. CBP60197), PLC (CoBioer Biosciences Co., Ltd.; cat. no. CBP60223), Huh7 (CoBioer Biosciences Co., Ltd.; cat. no. CBP60202) and MIHA (Hunan Fenghui Biotechnology Co., Ltd.; cat. no. CL0469) cells were cultured in DMEM (cat. no. 12430104; Invitrogen; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (cat. no. 10091148; HyClone; Cytiva) and 1% penicillin-streptomycin solution (cat. no. 15140163; Gibco; Thermo Fisher Scientific, Inc.; ratio 9:1:1) at 37°C and 5% CO2. Cross-contamination of the cell lines was excluded by short tandem repeat profiling (17 (link),18 (link)).
3
Isolation and Priming of Primary Murine Peritoneal Macrophages
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Primary peritoneal macrophages (PMs) were obtained from C57BL/6N mice intraperitoneally injected with 2 mL of 3% fluid thioglycollate medium (Merck, Germany) three days before cell collection. PMs were cultured in 3.5 mm dishes (Nunc, ThermoFisher Scientific, USA) or 6-well plates (Nunc, ThermoFisher Scientific, USA) with RPMI 1640 medium (Hyclone, Cytiva, USA) containing 10% fetal bovine serum (FBS) (Gibco, ThermoFisher Scientific, USA) and 1% penicillin-streptomycin Solution (Cytiva, USA). After 3 hours, the medium was replaced by a fresh RPMI 1640 medium (Hyclone, Cytiva, USA), after which the cells were cultured overnight. The medium was changed to opti-MEM (Gibco, ThermoFisher Scientific, USA) for 1 hour. Cells were then transfected with CLP/siRNA for 48 hours. After that, the cells were primed with ultrapure lipopolysaccharide (LPS) (AdipoGen, Switzerland) for 2 hours and subsequently stimulated with nigericin (20 μM) (AdipoGen, Switzerland) for 45 min. Finally, cell extracts and precipitated supernatants were analyzed by Western blot (WB).
4
Hierarchical Vessel-Supported Tumor Model
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Pooled HUVECs (Lonza) were cultured in
EGM-2 MV medium (CC-3202, Lonza) and used for network formation between
passages 3 and 7. Normal human lung fibroblasts (NHLFs) (Lonza) were
cultured in FBM-2 medium (CC-3132, Lonza) and used for the experiments
between passages 5 and 12. The NHLFs were used here as Margolis et
al. reported that NHLFs can facilitate vessel network formation.42 (link) To construct the hierarchical vessel-supported
tumor model, the HCT116 cells (ATCC) were cultured in high-glucose
DMEM (Gibco) with 10% fetal bovine serum (R&D systems) and 1%
penicillin–streptomycin solution (Cytiva). All of the cells
were proliferated in 25 cm2 flasks, and the culture medium
was changed every other day.
EGM-2 MV medium (CC-3202, Lonza) and used for network formation between
passages 3 and 7. Normal human lung fibroblasts (NHLFs) (Lonza) were
cultured in FBM-2 medium (CC-3132, Lonza) and used for the experiments
between passages 5 and 12. The NHLFs were used here as Margolis et
al. reported that NHLFs can facilitate vessel network formation.42 (link) To construct the hierarchical vessel-supported
tumor model, the HCT116 cells (ATCC) were cultured in high-glucose
DMEM (Gibco) with 10% fetal bovine serum (R&D systems) and 1%
penicillin–streptomycin solution (Cytiva). All of the cells
were proliferated in 25 cm2 flasks, and the culture medium
was changed every other day.
5
Culturing MCF-7 Breast Cancer Cells
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MCF-7, an ER-positive human breast cancer cell line, was purchased from American Type Culture Collection (ATCC) and cultured in DMEM medium supplemented with 10% fetal bovine serum and penicillin-streptomycin solution (100 units/mL penicillin and 100 µg/mL streptomycin: Hyclone Laboratories Inc., South Logan, UT, USA). The cells were grown at 37°C in a humidified atmosphere of 95% air/5% CO 2 . The medium was renewed 2-3 times per week, and before reaching confluence, the cells were subcultured every 3-4 days in a 1:4 ratio.
6
Murine Brain Cell Culture Protocol
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The complete culture medium for all cell lines was DMEM supplemented with 10% fetal bovine serum (ATLANTA Biologicals, Flowery Branch, GA, USA) and 1% Penicillin/Streptomycin solution (HyClone Laboratories Inc., South Logan, UT, USA). Murine brain cell lines bEnd.3 (endothelial cells), N2a (neuroblastoma cells), and C8-D1A (astrocytes) were purchased from ATCC. Murine brain cell line BV-2 (microglia) was obtained from Dr. G. Jean Harry (National Institute of Environmental Health Sciences, Research Triangle Park, NC). All cell lines were maintained according to provided protocols and previous studies [5 ]. Briefly, the cells were cultured in 75 cm2 flasks (Corning, NY, USA) separately, and medium was refreshed every other day until the cells reached ≥ 90% confluency.
7
Investigating Itraconazole and Autophagy Modulation
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Itraconazole (I129771) was purchased from Aladdin. IR820 (448014) was obtained from J&K Scientific. 3-methyladenine (3-MA) was purchased from MedChem Express. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), dimethyl sulfoxide, crystal violet (C0775), and chloroquine (CQ) diphosphate salt (C6628) were purchased from Millipore Sigma. DAPI was purchased from Invitrogen. Sodium chloride injection was purchased from Sichuan Kelun Pharmaceutical Co., Ltd. Mel (Purity, 99.5%). Penicillin-streptomycin solution was manufactured for HyClone Laboratories. Fetal bovine serum albumin (FBS) was purchased from Biowest. Dulbecco's modified eagle medium (DMEM) was purchased from Thermo Fisher Biochemical Products (Beijing) Co., Ltd. The antibodies used were as follows: anti-β-actin was purchased from Santa Cruz Biotechnology; cleaved-caspase 3 caspase 3, p62, PARP, cleaved-PARP were acquired from Cell Signaling Technology; Ki67 was purchased from Bioss; and LC3 was purchased from Novus. Horseradish peroxidase-conjugated anti-rabbit secondary antibody was obtained from Santa Cruz Biotechnology. For immunofluorescence, goat anti-rabbit Alexa Fluor 488 and goat anti-mouse Alexa Fluor 594 were obtained from Thermo Fisher Scientific.
8
Cultivation of HEK293T Cells and Viral Samples
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HEK293T cells were purchased from BeNa Culture Collection, China. Cells were cultured in a Dulbecco’s modified Eagle’s medium (DMEM; Gibco, New York, NY, USA) containing 10% fetal bovine serum (FBS; Gibco) and 1% penicillin-streptomycin solution (Hyclone Laboratories, Logan, UT, USA).
The NiVpv and HeVpv used in the study were kindly gifted from Dr. Youchun Wang (Division of HIV/AIDS and Sexually Transmitted Virus Vaccines, National Institutes for Food and Drug Control (NIFDC), Beijing, China).
The NiVpv and HeVpv used in the study were kindly gifted from Dr. Youchun Wang (Division of HIV/AIDS and Sexually Transmitted Virus Vaccines, National Institutes for Food and Drug Control (NIFDC), Beijing, China).
9
Culturing Human Gastric Cancer Cells
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Cells of the human gastric cancer cell lines SGC-7901, MGC-803 and MKN-45 were purchased from the Chinese Academy of Medical Science. The cells were cultured in RPMI 1640 (HyClone Laboratories, Logan, USA) supplemented with 10% fetal bovine serum (FBS; HyClone Laboratories, Logan, USA) and 1% penicillin-streptomycin solution (HyClone Laboratories, Logan, USA). The cells were incubated under humidified conditions at 37°C and 5% CO2.
10
Cytotoxicity and ROS Assay Protocol
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CLT (MED80120) was purchased from Medbio Pharmaceutical Technology Company. IR-820 (N119962) and hydroxychloroquine sulfate (H141480) were purchased from Shanghai Aladdin Biochemical Co., Ltd. Hyaluronic acid (H909935) was purchased from Shanghai Macklin Biochemical Co., Ltd. Penicillin-streptomycin solution (SV30010) was manufactured by HyClone Laboratories. Fetal bovine serum albumin (FBS) (FSP500) was purchased from ExCell Bio. Dulbecco’s modified Eagle’s medium (DMEM) (C11995500BT) was purchased from ThermoFisher Biochemical Products Co., Ltd. The ROS assay kit (S0033S), LDH assay (C0016), Calcein AM/PI cell viability/cytotoxicity assay kit (C2015S), and mitochondrial membrane potential assay kit with JC-1 (C2006) were provided by Beyotime Biotechnology. An Annexin V-FITC/PI apoptosis detection kit (40302ES20) was purchased from Yeasen Biotech Co., Ltd. EdU assay kit (C10310) was provided by Guangzhou RiboBio Co., Ltd. 3-(4,5-dimethyl-2-thiazoyl)-2,5-diphenyltetra tetrazolium bromide (MTT) (M2128) and N-acetyl cysteine (NAC) (A9165) were obtained from Sigma-Aldrich Co., Ltd. DQ-BSA Red (D-12,051) was purchased from ThermoFisher Scientific Co., Ltd. We thank the Pub-lab of West China School of Basic Medical Sciences & Forensic Medicine, Sichuan University for providing several laboratory instruments.
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