Intestinal porcine epithelial cells (IPEC-J2; ACC 701, Leibniz Institute DSMZ—German Collection of Microorganisms and Cell Culture, Braunschweig, Germany) were cultivated and routinely maintained using DMEM/F12 (1:1) without L-glutamine (Pan Biotech, Aidenbach, Germany), adjusted to 2.4 g/L NaHCO3 (Pan Biotech) and supplemented with 1% insulin–transferrin–selenium ITS (Gibco/Life Technologies, Thermo Fisher Scientific, Vienna Austria), 2.5 mM Glutamax (Gibco/Life Technologies, Thermo Fisher Scientific), 5 ng/mL epidermal growth factor (Corning Inc, Corning, USA), and 16 mM HEPES (Merck/Sigma Aldrich, Vienna, Austria). The medium was further supplemented with 10% heat-inactivated (30 min at 56 °C) fetal bovine serum FBS (Gibco/Life Technologies, Thermo Fisher Scientific) and 1% penicillin–streptomycin (Merck/Sigma Aldrich) directly before use. Cultures were incubated at 39 °C and 10% CO2 under a humidified atmosphere (CO2 incubator, Binder, Tuttlingen, Germany) and sub-cultivated upon exceeding 90% confluency in the cultivation vessels. Routine testing for mycoplasma contaminations was conducted by PCR (Venor® GEM Classic Mycoplasma Detection Kit, Minerva Biolabs, Berlin, Germany).
Nahco3
Manufactured by PAN Biotech
Sourced in Germany
NaHCO3 is a chemical compound that is commonly known as sodium bicarbonate. It is a white, crystalline solid with a molecular formula of NaHCO3. The core function of NaHCO3 is to act as a pH regulator, buffer, and neutralizing agent.
Automatically generated - may contain errors
Lab products found in correlation
L glutamine, by PAN Biotech (8 mentions)
Hoechst 33342, by Thermo Fisher Scientific (4 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Fetal bovine serum (fbs), by Merck Group (3 mentions)
Glutamine, by PAN Biotech (2 mentions)
Spy555 tubulin, by Spirochrome (2 mentions)
Penicillin streptomycin, by PAN Biotech (2 mentions)
Hfobs, by American Type Culture Collection (2 mentions)
L glutamine, by Thermo Fisher Scientific (2 mentions)
Recombinant human il 17 alpha, by RnD Systems (2 mentions)
Ld540, by Thermo Fisher Scientific (2 mentions)
Williams e medium, by Merck Group (2 mentions)
Epidermal growth factor (egf), by Corning (1 mentions)
Venor gem classic mycoplasma detection kit, by Minerva Biolabs (1 mentions)
Insulin transferrin selenium, by Thermo Fisher Scientific (1 mentions)
Co2 incubator, by Binder (1 mentions)
Hepes, by Merck Group (1 mentions)
Glutamax, by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin, by Merck Group (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
19 protocols using nahco3
1
Cultivation of porcine intestinal epithelial cells
2
Live-cell imaging of Plasmodium falciparum
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For live-cell imaging, NF54_PfCentrin1-GFP cells were seeded on glass bottom dishes as described above. Imaging medium, that is, phenol red-free RPMI 1640 supplemented with stable Glutamine and 2 g/l NaHCO3 (PAN Biotech) with all other supplements as in the parasite culture medium, was equilibrated in the cell culture incubator for several hours. Immediately before imaging, 9 ml of equilibrated imaging medium were supplemented with 4.5 μl (1:2,000 dilution) of the live microtubule dye SPY555-Tubulin (Spirochrome). List of all used dyes can be found in Table S3. Culture medium in the glass bottom dish with seeded cells was replaced by 8 ml imaging medium, the dish closed tightly without creating air bubbles, and sealed completely with parafilm. The imaging dish was directly taken to the incubation chamber of the microscope, prewarmed to 37°C.
3
Culturing Human Hepatoma HepG2 Cells
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Human hepatoma cell line HepG2 (DSMZ-No: ACC-180) was purchased from the German Collection of Microorganisms and Cell Cultures (DSMZ, Braunschweig, Germany). The cells were cultured in RPMI (Roswell Park Memorial Institute) 1640 medium enriched with L-glutamine, 2.0 g/L NaHCO3 (PAN Biotech, Aidenbach, Germany), 10% (v/v) fetal bovine serum (FBS; Fisher Scientific, Schwerte, Germany), 100 IU/mL penicillin, and 100 μg/mL streptomycin (Life Technologies, Darmstadt, Germany). For optimal cell cultivation, we kept the cells at 37 °C under a humidified atmosphere of 95% air and 5% CO2 by replacing the medium every 2–3 days. Once a week, the cells were passaged using the cell passages 20–25 for the following in vitro experiments.
4
Culturing Human Hepatoma Cells
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Human hepatoma cells (HepG2 cells; DSMZ-No: ACC-180) were purchased from the German Collection of Microorganisms and Cell Cultures (Braunschweig, Germany). Cell culture medium consisted of Roswell Park Memorial Institute (RPMI) 1640 medium supplemented with L -glutamine, 2.0 g/L NaHCO3 (all obtained from PAN Biotech, Aidenbach Germany), 10% (v/v) fetal bovine serum (Fisher Scientific, Schwerte Germany), 100 IU/mL penicillin and 100 µg/mL streptomycin. The cells were kept at 37 °C with 5% CO2 in a humidified atmosphere by changing the medium every 2–3 days.
5
Generation of Immortalized Per2-Deficient MEFs
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Mouse embryonic fibroblasts (MEFs) were obtained from mice without functional Per2 in all body cells42 (link), or wild-type mice. Briefly, mice containing the ubiquitously expressed Cre recombinase transgene (European Mouse Mutant Archive (EMMA) EM:01149, B6.129-Cre-Deleter: B6.C(129)-Tg(CMV-cre)1Cgn/CgnIbcm) crossed with a conditional Per2 allele42 (link) were backcrossed over ten generations to obtain Per2 homozygous knock-out mice without Cre recombinase. The MEFs subsequently were serially diluted every three days over 30 passages to yield immortalized cells. All lines were maintained in Dulbecco's modified Eagle's medium (DMEM) containing 4.5 g/l glucose, stable glutamine, sodium pyruvate and 3.7 g/l NaHCO3 (PAN Biotech), supplemented with 10% fetal calf serum (FCS) and 100 U/mL penicillin–streptomycin at 37 °C in a humidified atmosphere containing 5% CO2. Forskolin stimulation (10–100 µM) was used to mimic in vitro the molecular pathway activated by light in mice. Samples were collected at specific time points mentioned in the text.
6
Culturing Human Fetal Osteoblasts and Periodontal Ligament Fibroblasts
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A human fetal osteoblast cell line (hFOBs; American Type Culture Collection—ATCC®, Manassas, VA, USA) was cultured in Dulbecco’s Modified Eagle Medium/Nutrient Mixture F-12 (DMEM-F12), without phenol red (PAN-Biotech GmbH, Aidenbach, Germany), containing 2.5 mM L-glutamine (PanBiotech®, Aidenbach, Germany), 10% (v/v) fetal bovine serum (FBS; Sigma-Aldrich, St. Louis, MO, USA) and 0.3 mg/mL geneticin (G418; PAN-Biotech GmbH, Aidenbach, Germany). The hFOBs were used between 12 and 14 passages.
Human periodontal ligament fibroblasts (hPLFs) were isolated from healthy human periodontal tissue and purchased from Innoprot®, Spain. They were cultured in DMEM-F12 culture medium supplemented with stable glutamine and 1.2 g/L NaHCO3 (PAN-Biotech GmbH, Aidenbach, Germany) with 10% FBS (Sigma-Aldrich) and 1% (v/v) penicillin–streptomycin (PAN-Biotech GmbH, Aidenbach, Germany). The hPLF cells were used at the third passage. Both hFOBs and hPLFs were maintained at 37 °C in a humidified atmosphere of 5% CO2, and the culture medium was changed twice a week.
Human periodontal ligament fibroblasts (hPLFs) were isolated from healthy human periodontal tissue and purchased from Innoprot®, Spain. They were cultured in DMEM-F12 culture medium supplemented with stable glutamine and 1.2 g/L NaHCO3 (PAN-Biotech GmbH, Aidenbach, Germany) with 10% FBS (Sigma-Aldrich) and 1% (v/v) penicillin–streptomycin (PAN-Biotech GmbH, Aidenbach, Germany). The hPLF cells were used at the third passage. Both hFOBs and hPLFs were maintained at 37 °C in a humidified atmosphere of 5% CO2, and the culture medium was changed twice a week.
7
Standardized Rabies Virus Neutralization Assay
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Clinical serum samples were obtained from the routine diagnostics and stored at -20°C. WHO-2 Standard Rabies Immunoglobulin (SRIG) (code RAI, NIBSC 30 IU) and HRIG (Berirab, CSL Behring GmbH, No: 107a/89) were adjusted to 1 IU/ml by dilution and served as references. Baby hamster kidney fibroblasts subclone 13 (BHK-21 C13; ATCC CCL-10) cells were grown in MEM Eagle with Earle’s Balanced Salt Solution (EBSS), L-Glutamine (292 mg/L), and 2.2 g/L NaHCO3 (PAN-Biotech Cat. No. P04-08500) supplemented with 10% (v/v) fetal calf serum (FCS), 1% (v/v) ZellShield (Minerva cat. no. 13–0150), and NaHCO3 (Merck cat. no. 1.06329.1000) solution to get a final pH of 7.5–7.8. Cells were cultivated at 37°C in an atmosphere of 5% CO2 and approved to be free of mycoplasma periodically. Challenge Virus Standard (CVS-11) stock was received from Friedrich-Löffler-Institut, Wusterhausen, Germany. Virus stocks were aliquoted and stored at -80°C. Viral titers were determined by TCID50 titration. All steps in which CVS-11 was handled prior to chemical fixation and virus inactivation were conducted in a BSL2 laboratory specifically dedicated to Rabies lyssavirus work. Only employees who had been successfully vaccinated against Rabies lyssavirus had access to this area.
8
Lingzhi Mushroom Powder Production
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The lingzhi mushrooms (G. lucidum) were supplied by AU Farm, Khlong Toei District, Bangkok, Thailand. The powder from the lingzhi mushrooms was produced using a slightly modified version of the technique proposed by Ketprayoon et al. [25 (link)] Initially, the mushrooms were dried at 60 °C in a hot air oven. A grinder was then used to make a fine powder which was then sieved using a 90 μm mesh. The surface of the mushroom powder then acts to support the hydrolyzing protein. This powder was then stored in a vacuum-sealed polypropylene bag in a desiccator at room temperature until required for further use.
American Type Culture Collection (ATCC, Manassas, VA, USA) provided the human intestinal epithelial cell line, Caco-2 cells (ATCC® HTB-37™) which then underwent culturing in Eagle's Minimum Essential Medium (EMEM) which was supplemented with L-glutamine and 10% fetal bovine serum supplied by Gibco (Rockville, MD, USA). American Type Culture Collection (ATCC, Manassas, VA, USA) also supplied the RAW 264.7 cell lines (ATCC® TIB-71™) which were then grown in Dulbecco's Modified Eagle’s Medium (DMEM) and supplemented using high glucose 4.5 g/L with sodium pyruvate, L-glutamine, and 3.7 g/L NaHCO3 (PAN Biotech, Aidenbach, Bavaria, Germany). The cells were stored until use in a humidified chamber under 5% carbon dioxide at a temperature of 37 °C.
American Type Culture Collection (ATCC, Manassas, VA, USA) provided the human intestinal epithelial cell line, Caco-2 cells (ATCC® HTB-37™) which then underwent culturing in Eagle's Minimum Essential Medium (EMEM) which was supplemented with L-glutamine and 10% fetal bovine serum supplied by Gibco (Rockville, MD, USA). American Type Culture Collection (ATCC, Manassas, VA, USA) also supplied the RAW 264.7 cell lines (ATCC® TIB-71™) which were then grown in Dulbecco's Modified Eagle’s Medium (DMEM) and supplemented using high glucose 4.5 g/L with sodium pyruvate, L-glutamine, and 3.7 g/L NaHCO3 (PAN Biotech, Aidenbach, Bavaria, Germany). The cells were stored until use in a humidified chamber under 5% carbon dioxide at a temperature of 37 °C.
9
In vitro culture of osteoblasts and fibroblasts
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The human fetal osteoblast cell line (hFOBs) was purchased from the American Type Culture Collection (CRL-11372, ATCC®, Manassas, VA, USA). The cells were maintained in Dulbecco’s Modified Eagle Medium/Nutrient Mixture F-12 (DMEM:F12, 1:1) without phenol red (PAN-Biotech GmbH, Aidenbach, Germany), containing 10% (v/v) fetal bovine serum (FBS, Sigma-Aldrich, St. Louis, MO, USA), 2.5 mM L-glutamine (PAN-Biotech®, Germany) and 0.3 mg/mL antibiotic G418 (PAN-Biotech GmbH, Germany).
The human periodontal ligament fibroblasts (hPLFs) were acquired from Innoprot® (P10867, Innoprot®, Derio, Spain). The cells were cultured in culture medium composed of DMEM:F12 Mix (1:1) with stable glutamine and 1.2 g/L NaHCO3 (PAN-Biotech®, Germany), supplemented with 1% (v/v) penicillin/streptomycin and 10% (v/v) FBS.
Both the osteoblasts and fibroblasts were maintained in an incubator at 37 °C and 5% (v/v) CO2, in a humidified atmosphere of 100% relative humidity. The medium of both cell lines was changed twice a week, and both cells were observed every day using an inverted microscope (Kern®, Lohmar, Germany) at 10× magnification to detect morphologic changes.
The human periodontal ligament fibroblasts (hPLFs) were acquired from Innoprot® (P10867, Innoprot®, Derio, Spain). The cells were cultured in culture medium composed of DMEM:F12 Mix (1:1) with stable glutamine and 1.2 g/L NaHCO3 (PAN-Biotech®, Germany), supplemented with 1% (v/v) penicillin/streptomycin and 10% (v/v) FBS.
Both the osteoblasts and fibroblasts were maintained in an incubator at 37 °C and 5% (v/v) CO2, in a humidified atmosphere of 100% relative humidity. The medium of both cell lines was changed twice a week, and both cells were observed every day using an inverted microscope (Kern®, Lohmar, Germany) at 10× magnification to detect morphologic changes.
10
Colorectal Cancer Cell Line Profiling
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For the experiments, the following human colorectal cancer cell lines were used: DLD1, LoVo, LS-174T, SW620, SW480, SW837, HT29, and HCT116. The origins of these cell lines are given in Supplementary Table 1 . All cell lines were genotyped using Multiplex Cell Authentication by Multiplexion (Heidelberg, Germany) and were regularly tested negative for mycoplasma contamination.
Except for SW620 all cell lines were cultured in RPMI 1640 with stable glutamine and 2.0 g/L NaHCO3 (PAN Biotech, Aidenbach, Germany) supplemented with 10% fetal bovine serum (FBS) (PAN-Biotech) and 1% penicillin‒streptomycin (P/S; PAN-Biotech). The cell line SW620 was cultured in DMEM (Gibco/Life Technologies) supplemented with 10% FBS and 1% P/S. The cells were cultured in a humidified atmosphere at 37 °C and 5% CO2. For the collection of cell pellets, cells were seeded at specific cell numbers (1.5 × 106 for HCT116 and HT29; 3.0 × 106 for DLD1 and LoVo; 2.5 × 106 for LS-174T and SW480; 4.0 × 106 for SW837; 7.0 × 106 for SW620) and collected after 48 h.
Except for SW620 all cell lines were cultured in RPMI 1640 with stable glutamine and 2.0 g/L NaHCO3 (PAN Biotech, Aidenbach, Germany) supplemented with 10% fetal bovine serum (FBS) (PAN-Biotech) and 1% penicillin‒streptomycin (P/S; PAN-Biotech). The cell line SW620 was cultured in DMEM (Gibco/Life Technologies) supplemented with 10% FBS and 1% P/S. The cells were cultured in a humidified atmosphere at 37 °C and 5% CO2. For the collection of cell pellets, cells were seeded at specific cell numbers (1.5 × 106 for HCT116 and HT29; 3.0 × 106 for DLD1 and LoVo; 2.5 × 106 for LS-174T and SW480; 4.0 × 106 for SW837; 7.0 × 106 for SW620) and collected after 48 h.
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