THP1 cells and the THP1 Blue−NFκB monocyte cell line, carrying a stable integrated NFκB-inducible Secreted Embryonic Alkaline Phosphatase (SEAP) reporter construct used to analyze NFκB induction, were purchased from InvivoGen. The THP1 cells were maintained in supplemented RPMI medium (10% fetal bovine serum, 1% gentamycin, and 1% b-mercaptoethanol), and the THP1 Blue−NFκB cell line was maintained in the same medium supplemented with 100 μg/ml normicin (InvivoGen) and 100 U/ml-100 μg/ml pen-strep (InvivoGen). Cell handling and preparation were performed in accordance with the manufacturer's protocol (InvivoGen).
Thp1 blue nf κb cell line
Manufactured by InvivoGen
Sourced in United States
The THP1-Blue™ NF-κB cell line is a human monocytic cell line that stably expresses an NF-κB-inducible secreted embryonic alkaline phosphatase (SEAP) reporter gene. This cell line is a useful tool for screening and studying NF-κB activation in response to various stimuli.
Automatically generated - may contain errors
Lab products found in correlation
Streptomycin, by Merck Group (3 mentions)
Penicillin, by Merck Group (3 mentions)
Fetal bovine serum (fbs), by Merck Group (3 mentions)
Rpmi 1640 medium, by Merck Group (2 mentions)
Thp 1 cells, by InvivoGen (1 mentions)
Normicin, by InvivoGen (1 mentions)
Pen strep, by InvivoGen (1 mentions)
Wst 1 cell proliferation reagent kit, by Roche (1 mentions)
Prism 8, by GraphPad (1 mentions)
L glutamine, by Merck Group (1 mentions)
Roswell park memorial institute (rpmi), by Biosera (1 mentions)
Cell counting kit 8 (cck8), by Merck Group (1 mentions)
Penicillin, by Euroclone (1 mentions)
Streptomycin, by Euroclone (1 mentions)
Atcc crl 1573, by LGC (1 mentions)
Hepes, by Euroclone (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
L glutamine, by Euroclone (1 mentions)
β mercaptoethanol, by Merck Group (1 mentions)
5 protocols using thp1 blue nf κb cell line
1
Analyzing NF-κB Induction in THP1 Cells
2
Cytotoxic Effect of Compounds on THP1-Blue™ NF-κB Cells
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The cytotoxic effect of the tested compounds was specified on THP1-Blue™ NF-κB cell line (Invivogen; San Diego, CA, USA), as described previously [28 (link)]. Briefly, cells resuspended in serum-free RPMI 1640 medium (Merck, Darmstadt, Germany) supplemented with antibiotics (100 U/mL penicillin and 100 mg/mL streptomycin (Merck)) and 10% FBS (Merck) were seeded into 96-well plates (100 µL/well, i.e., 50,000 cells per each well). After 2 h, tested compounds dissolved in DMSO (1.25–20 µM) were added to the cells. The final concentration of DMSO was 0.1% (v/v) in each well. The viability analysis was performed after 24 h incubation with the tested substances using the WST-1 Cell Proliferation Reagent kit (Roche Diagnostics, Basel, Switzerland) according to the manufacturer’s manual. The amount of formazan formed, which corresponded to the number of metabolically active cells in the culture, was calculated as a percentage of the control cells, which were treated only with serum-free RPMI 1640 medium and were assigned as 100%. The IC50 values were calculated by four-parameter logistic (4PL) analysis from obtained viability curves by GraphPad Prism 8.0.1 (San Diego, CA, USA) software. The results are summarized in Table 1 as mean ± SEM (n = 6).
3
THP1-Blue NF-κB Cell Culture Protocol
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THP1-Blue™ NF-κB cell line was purchased from Invivogen (San Diego, CA, USA). Cells were cultured in RPMI 1640 medium (Merck, Darmstadt, Germany) containing stabilized 2 mM l -glutamine (Merck) supplemented with antibiotics [100 U/mL penicillin and 100 mg/mL streptomycin (Merck)] and 10% FBS (Merck). The cultures were passaged twice a week. Cells were kept in an incubator at 37 °C in a water-saturated atmosphere of air containing 5% CO2.
The viability of the used cell lines (5th–18th passage) was over 95% for each experiment. The cell number and viability were determined by staining with Trypan Blue solution. Cells were counted manually using a haemocytometer and an optical microscope. Cells that remained unstained were considered viable, while light red cells were considered non-viable. The viability percentage was calculated as the ratio of the number of all viable cells to the number of all cells.
The viability of the used cell lines (5th–18th passage) was over 95% for each experiment. The cell number and viability were determined by staining with Trypan Blue solution. Cells were counted manually using a haemocytometer and an optical microscope. Cells that remained unstained were considered viable, while light red cells were considered non-viable. The viability percentage was calculated as the ratio of the number of all viable cells to the number of all cells.
4
Anti-NF-κB and Pro-PPARγ Evaluation
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THP1-Blue™ NF-κB cell line (InvivoGen; San Diego, CA, USA) was used to evaluate anti-NF-κB and pro-PPARγ actions of the tested compounds, i.e., complex (1 ) and its precursors [Au(PPh3)Cl] and free kinetin, as well as the reference drug Auranofin (Merck, Rahway, NJ, USA). The cells were cultivated in RPMI (Roswell Park Memorial Institute) 1640 medium (Biosera; Nuaille, France) supplemented with 10% fetal bovine serum (FBS) and antibiotics (100 U/mL penicillin and 100 mg/mL streptomycin) (both from Merck). All further experiments were performed in serum-free medium. The effect of the test compounds dissolved in DMF on cell viability after 24 h incubation was determined using a Cell Counting Kit-8 (CCK-8; Merck) according to the manufacturer’s instructions, similarly as previously described [18 (link)].
5
Cell culture protocols for cancer research
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Chicken ovalbumin (OVA)-transfected EL-4 (EG7) cell line derived from C57BL/6J mice and human embryonic kidney cell line 293 (HEK293, ATCC® CRL-1573™, LGC Standards S.r.l., Milano, Italy), THP-1 cell line (ATCC® TIB-202™, LGC Standards S.r.l., Milano, Italy), THP1-Blue™ NF-κB cell line (InvivoGen, CA, USA), MCA203 cell line, a fibrosarcoma induced by 3-methylcholanthrene (a kind gift by Prof. M.P.Colombo) and MCA205 cell line, a mouse sarcoma (a kind gift by Prof. L. Zitvogel) were grown in DMEM (Invitrogen, Carlsbad, CA, USA) supplemented with 2 mM L-glutamine (Euroclone, Milano, Italy), 10 mM HEPES (Euroclone, Milano, Italy), 20 µM β-mercaptoethanol (Sigma-Aldrich, Saint Louis, MO, USA), 150 U/ml streptomycin (Euroclone, Milano, Italy), 200 U/ml penicillin (Euroclone, Milano, Italy) and 10% heat-inactivated fetal bovine serum (FBS; Invitrogen, Carlsbad, CA, USA). MSC2 is an immortalized cell line derived from Balb/c mice26 (link) and it was cultured in RPMI 1640 (Life Technologies, Carlsbad, CA, USA) supplemented with 2 mM L-glutamine (Euroclone, Milano, Italy), 10 mM HEPES (Euroclone, Milano, Italy), 150 U/ml streptomycin (Euroclone, Milano, Italy), 200 U/ml penicillin (Euroclone, Milano, Italy) and 10% heat-inactivated FBS (Superior, Merck, Darmstadt, Germany). All cell lines were tested to be free from Mycoplasma contamination by PCR screening.
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