Ultrapure lps

Manufactured by Thermo Fisher Scientific

Sourced in United States

Ultrapure LPS is a laboratory instrument designed to produce high-purity water by removing lipopolysaccharides (LPS) and other endotoxins. It utilizes a specialized filtration process to purify water, ensuring low endotoxin levels for sensitive applications.

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Lab products found in correlation

Nigericin, by Merck Group (5 mentions) Adenosine triphosphate (atp), by Merck Group (4 mentions) Mitosox, by Thermo Fisher Scientific (4 mentions) Mitotracker, by Thermo Fisher Scientific (4 mentions) Anti mouse caspase 1 p20 ag 20b 0042, by Adipogen (3 mentions) Anti mouse il 1β p17 af 401 na, by R&D Systems (3 mentions) Anti nlrp3 ag 20b 0014, by Adipogen (3 mentions) Pam3csk4, by Thermo Fisher Scientific (3 mentions) Anti flag f2555, by Merck Group (3 mentions) Poly a t, by Merck Group (3 mentions) Mcc950, by Selleck Chemicals (2 mentions) Protein g agarose, by Merck Group (2 mentions) Anti nek7 sc 50756, by Santa Cruz Biotechnology (2 mentions) Streptavidin coated beads, by Merck Group (2 mentions) Anti asc sc 22514 r, by Santa Cruz Biotechnology (2 mentions) Anti β actin p30002, by Abmart (2 mentions) Anti human cleaved il 1β a5208206, by Sangon (2 mentions) Anti human caspase 1, by Cell Signaling Technology (2 mentions) Anti mouse nlrp3 ag 20b 0014, by Adipogen (2 mentions) Anti β actin antibody 66009 1 ig, by Proteintech (2 mentions)

Close spelling variants of this product

Ultrapure Escherichia coli 0111:B4 LPS free of lipoproteins (2 citations)

12 protocols using ultrapure lps

Inflammasome Activation in Immune Cells

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The following reagents and chemicals were used: ultrapure LPS (Invitrogen, Waltham, MA, USA), ATP, DMSO, glucose, nigericin (Nig), trypan blue (all Sigma-Aldrich, St. Louis, MO, USA), Cytofix/Cytoperm™ Fixation/Permeabilization Kit (BD Bioscience, Franklin Lakes, NJ, USA), EDTA (Invitrogen), fetal calf serum (FCS), Ficoll, HEPES, PBS, Penicillin/Streptomycin, RPMI 1640 (all PAN-Biotech, Aidenbach, Germany), potassium chloride (Merck, Darmstadt, Germany) and β-mercaptoethanol (Thermo Fisher Scientific, Waltham, MA, USA). The following Antibodies (Abs) were used: rabbit polyclonal anti-ASC (AL177, Adipogen, San Diego, CA, USA), anti-rabbit-IgG-AlexaFluor488 (Invitrogen, Waltham, MA, USA), and lineage-specific antibodies CD3 PerCP, CD4 APC, CD14 APC-Cy7, CD16 PE-Cy7 (Biolegend, San Diego, CA, USA).
Reagents for immunofluorescence microscopy were: methanol (J.T. Baker, Phillipsburg, NJ, USA), bovine serum albumin (BSA) (Serva, Heidelberg, Germany), saponin, sodium pyruvate (Sigma-Aldrich, St. Louis, MO, USA), Image-iT™ FX Signal Enhancer and ProLong™ Diamond Antifade Mountant with DAPI (Invitrogen), and the following Abs were used: rabbit anti-ASC (AL177, Adipogen), mouse anti-beta-actin (Sigma-Aldrich), anti-rabbit-IgG-AlexaFluor488 (Invitrogen) and anti-mouse-IgG-AlexaFluor594 (Dianova, Hamburg, Germany).

Wittmann N., Behrendt A.K., Mishra N., Bossaller L, & Meyer-Bahlburg A. (2021). Instructions for Flow Cytometric Detection of ASC Specks as a Readout of Inflammasome Activation in Human Blood. Cells, 10(11), 2880.

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Inflammasome Activation Assay Protocol

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HDM was from Greer Laboratories. Anti-mouse IL-1β (p17) (AF-401-NA) was from R&D. Anti-mouse caspase-1 (p20) (AG-20B-0042) and anti-NLRP3 (AG-20B-0014) antibodies were from Adipogen. Anti-ASC (67824) antibody was from Cell Signaling Technology. Anti-β-actin (66009-1-Ig) was from Proteintech.
Anti-mouse antibodies used for flow cytometry were: CD3-FITC (BD, 553062, 145-2C11), CD19-FITC (eBioscience, 11-0193-82, 1D3), Ly-6G-PE (Biolegend, 127608, 1A8), CD45-PE (eBioscience, 12-0451-81, 30-F11), CD11c-PerCP-Cy5.5 (Biolegend, 117328, N418), CD11b-PerCP-Cy5.5 (BD, 550993, M1/70), CD11b-PE-Cy7 (eBioscience, 25-0112-82, M1/70), CD3e-PE-Cy7 (BD, 552774, 145-2C11), CD19-PE-Cy7 (Biolegend, 115520, 6D5), SiglecF-Alexa Fluor 647 (BD, 562680, E50-2440), MHCII-APC-eFluor 780 (eBioscience, 47-5321-82, M5/114.15.2), Ly-6G-BV421 (Biolegend, 127628, 1A8), CD45-BV510 (Biolegend, 103137, 30-F11), CD11c-BV510 (Biolegend, 117338, N418). Ultrapure LPS was obtained from Invitrogen. Nigericin was obtained from Sigma-Aldrich. RRx-001 (S8405) was from Selleck.

Ma M., Li G., Qi M., Jiang W, & Zhou R. (2021). Inhibition of the Inflammasome Activity of NLRP3 Attenuates HDM-Induced Allergic Asthma. Frontiers in Immunology, 12, 718779.

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Vagus Nerve Electrophysiology with LPS Challenge

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The electrophysiological signals were digitized from the vagus nerve using either a Neuralynx data acquisition system (Digital Lynx 4SX, Cheetah v5 software, Neuralynx, Bozeman, MT) or a Plexon data acquisition system (Omniplex, Plexon Inc., Dallas, Texas). Recordings were sampled at 32 kHz and band-pass filtered between 10 and 9000 Hz for the Neuralynx, and 40 kHz with a 120 Hz filter and 50 gain for the Plexon. All signals were referenced to the animal ground placed between the right salivary gland and the skin. For recordings with three-lead hook electrodes, the signals from the most proximal lead were referenced with the most distal lead to minimize noise and improve the signal to noise ratio. The experimenter was always grounded while manipulating the animal during recordings. In experiments with LPS challenge, following acquisition of the baseline activity (10 min), 8.0 mg/kg ultra-pure LPS (Invitrogen, San Diego, California) was administered intraperitoneally, and recordings were continued for 10 min post-injection.

Silverman H.A., Stiegler A., Tsaava T., Newman J., Steinberg B.E., Masi E.B., Robbiati S., Bouton C., Huerta P.T., Chavan S.S, & Tracey K.J. (2018). Standardization of methods to record Vagus nerve activity in mice. Bioelectronic Medicine, 4, 3.

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Isolation of Bone Marrow-Derived Macrophages

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Bone marrow derived macrophages (BMDM) were isolated from the femurs and tibias obtained from male C57/Bl6 wild type mice (8–10 weeks old) as previously described (Takeda et al., 2010 (link)). Briefly, cells were plated in DMEM supplemented with 10% heat inactivated FBS and 30% conditioned media (supernatant from M-CSF expressing L929 cells). Adherent BMDM were harvasted using 0.05% Trypsin EDTA after 7 days in culture. Ultrapure LPS was obtained from Invitrogen and R/S 2HG Octyl Ester Sodium salt were obtained from Toronto Research Canada.

Fitzpatrick S.F., Lambden S., Macias D., Puthucheary Z., Pietsch S., Mendil L., McPhail M.J, & Johnson R.S. (2020). 2-Hydroxyglutarate Metabolism Is Altered in an in vivo Model of LPS Induced Endotoxemia. Frontiers in Physiology, 11, 147.

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NLRP3 Inflammasome Activation Pathway

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MSU, nigericin, ATP, PMA, BHB, DL-sulforaphane, poly(dA:dT), and glucose were purchased from Sigma-Aldrich. MCC950 was acquired from Selleck. BAY 11-7082, isoliquiritigenin, parthenolide, flufenamic acid, and mefenamic acid were acquired from Topscience. Human recombinant insulin was bought from Novo Nordisk. The One Touch Ultra Blood glucose Test System kit was bought from Roche. Ultrapure LPS, Pam3CSK4, MitoTracker, and MitoSOX were obtained from Invitrogen. Protein G agarose and streptavidin-coated beads were respectively supplied by Millipore and Pierce Biochemicals. ATP-coupled beads were from BioWorld Company. Anti-mouse caspase-1 (p20; AG-20B-0042) and anti-NLRP3 (AG-20B-0014) were from Adipogen. Anti-ASC (sc-22514-R) and anti-NEK7 (SC-50756) were from Santa Cruz. Anti-β-actin (P30002) was bought from Abmart. Anti-human caspase-1 was from Cell Signaling Technology. Anti-human cleaved IL-1β (A5208206) was from Sangon Biotech. Anti-mouse IL-1β (p17; AF-401-NA) was from R&D Systems. Anti-Flag (F2555) and anti-Mcherry (V4888) were from Sigma-Aldrich. Recombinant human NLRP3 was from Novus Biologicals. A standard HFD (D12492, 60% kcal fat) was from Research Diet Company.

Jiang H., He H., Chen Y., Huang W., Cheng J., Ye J., Wang A., Tao J., Wang C., Liu Q., Jin T., Jiang W., Deng X, & Zhou R. (2017). Identification of a selective and direct NLRP3 inhibitor to treat inflammatory disorders. The Journal of Experimental Medicine, 214(11), 3219-3238.

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NLRP3 Inflammasome Activation Assay

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Apilimod (No. S6414) was bought from Selleck. Nigericin, MSU, and poly (A/T) were purchased from Sigma-Aldrich. Ultrapure LPS, MitoSOX, MitoTracker, DAPI, Lysotracker Green, MitoTracker Deep Red, MitoTracker Green were acquired from Invitrogen. MnTBAP was bought from Santa Cruz Biotechnology. The C3 toxin was a gift from Dr. Tengchuan Jin (University of Science and Technology of China, Hefei, China). CY-09 was synthesized from Dr. Xianming Deng (Xiamen University, Xiamen, China). MCC950 was bought from Selleck. Fluo-4, AM(HY-101896), ML-SI3 were purchased from MedChemExpress. BAPTA-AM(A1076) was bought from Sigma-Aldrich. Protein G agarose (16–266) was bought from Millipore. Anti-Flag (F2555) antibodies were from Sigma-Aldrich. The anti-mouse NLRP3 (AG-20B-0014) and the anti-mouse caspase-1 (AG-20B-0042) antibodies was from AdipoGen. The anti-mouse IL-1β antibody (AF-401-NA) was obtained from R&D Systems. The anti-β-actin antibody (66009-1-Ig) was from Proteintech Group. The anti-mouse ASC antibody (67824S) was from Cell Signaling Technology. The anti-mouse NEK7 antibody (ab133514) was from Abcam.

Hou Y., He H., Ma M, & Zhou R. (2023). Apilimod activates the NLRP3 inflammasome through lysosome-mediated mitochondrial damage. Frontiers in Immunology, 14, 1128700.

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Quantifying Nitric Oxide Levels

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The DAF2-DA assay is a fluorescence-based NO detection assay that uses a cell permanent 4,5-diaminofluorescein diacetate (DAF2-DA) as NO detector (Sigma-Aldrich, USA). Diacetate (DA) part of DAF2-DA is hydrolysed by cellular esterases to produce DAF2 compound that reacts with NO to produce a fluorescent triazolofluorescein (DAF-2T). Therefore, in this study, cells were seeded on coverslips in 6-well plates overnight and then pretreated with LiCl (5 and 10 mM), 10 mM NaCl and activated with 100 ng/ml Ultrapure LPS (Invitrogen, USA) for 24 hrs. Thereafter, cells were stained with 10 μm/ml DAF2-DA for 20 min at RT and washed with 1xPBS. Cells were further stained with 25 μg/ml Hoechst for 20 min at RT, thereafter followed by fixing with 2.7% paraformaldehyde for 30 min. Nitric oxide quantitation was accomplished using a Nikon Ti-E inverted fluorescent microscope (Nikon, Japan).

Makola R.T., Mbazima V.G., Mokgotho M.P., Gallicchio V.S, & Matsebatlela T.M. (2020). The Effect of Lithium on Inflammation-Associated Genes in Lipopolysaccharide-Activated Raw 264.7 Macrophages. International Journal of Inflammation, 2020, 8340195.

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Cytokine-Induced Macrophage Polarization

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Recombinant cytokines were all from Peprotech (Rocky Hill, NJ). To generate of M1 and M2 macrophages, bone marrow cells from the tibia and fibula of C57BL/6 mice (Harlan) were cultured at 37 °C in 5% CO₂ in RPMI supplemented with 1% L-glutamine and 10% foetal bovine serum with 10 ng/ml mouse M-CSF. Medium was replaced on days 3 and 6 and cells were harvested on day 8. For M1 phenotype induction, cells were stimulated for 24 h with 10 ng/ml M-CSF and 100 ng/ml IFN-γ and for an additional 24 h with 10 ng/ml M-CSF and 20 ng/ml ultra-pure LPS (Invitrogen, Carlsbad, CA). For M2 phenotype induction, cells were stimulated twice for 24 h with 10 ng/ml M-CSF and 20 ng/ml IL-4. To determine the effect of HMGB1 on M1 and M2 macrophages, culture medium used to induce M1- or M2-like macrophages was supplemented with 1 μg/ml recombinant HMGB1 (HMGBiotech).

Huber R., Meier B., Otsuka A., Fenini G., Satoh T., Gehrke S., Widmer D., Levesque M.P., Mangana J., Kerl K., Gebhardt C., Fujii H., Nakashima C., Nonomura Y., Kabashima K., Dummer R., Contassot E, & French L.E. (2016). Tumour hypoxia promotes melanoma growth and metastasis via High Mobility Group Box-1 and M2-like macrophages. Scientific Reports, 6, 29914.

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Lipopolysaccharide-Induced Shock in Mice

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Mice were monitored as described above using continuous telemetry for 24 h prior to administration of LPS at a dose of 10 mg/kg via intraperitoneal injection (Ultrapure LPS, Invitrogen). Haemodynamics and physical appearance were observed hourly after injection and culled upon reaching a predetermined humane severity endpoint.
Analysis of systemic haemodynamics during baseline conditions and following LPS treatment was by unpaired t test of the area under the curve for the respective time course of each animal.

Lambden S., Cowburn A.S., Macias D., Garrud T.A., Krause B.J., Giussani D.A., Summers C, & Johnson R.S. (2021). Endothelial cell regulation of systemic haemodynamics and metabolism acts through the HIF transcription factors. Intensive Care Medicine Experimental, 9, 28.

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NLRP3 Inflammasome Activation Assay

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ATP, Nigericin, MSU, and poly (A/T) were purchased from Sigma-Aldrich. Ultrapure LPS, Lipofextamine 2000, Pam3CSK4, MQAE, MitoSOX, MitoTracker, DAPI were acquired from Invitrogen. The Salmonella strain was a gift from Dr. Cai Zhang (Shandong University, Shandong, China). The C3 toxin was a gift from Dr. Tengchuan Jin (University of Science and Technology of China, Hefei, China). CY-09 was synthesized from Dr. Xianming Deng (Xiamen University, Xiamen, China). Protein G agarose (16-266) and anti-Flag (A2220) beads were bought from Millipore and Sigma-Aldrich, respectively. Anti-Flag (F2555) and anti-VSV (V4888) antibodies were from Sigma-Aldrich. The anti-mouse NLRP3 (AG-20B-0014) and the anti-mouse caspase-1 (AG-20B-0042) antibodies was from AdipoGen. The anti-mouse IL-1β antibody (AF-401-NA) was obtained from R&D Systems. The anti-β-actin antibody (66009-1-Ig) was from Proteintech Group. The anti-mouse ASC antibody (67824S) was from Cell Signaling Technology. The anti-mouse NEK7 antibody (ab133514) was from Abcam.

Lin H., Yang M., Li C., Lin B., Deng X., He H, & Zhou R. (2022). An RRx-001 Analogue With Potent Anti-NLRP3 Inflammasome Activity but Without High-Energy Nitro Functional Groups. Frontiers in Pharmacology, 13, 822833.

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