Ultrapure LPS-B5, ATP, nigericin, Pam3CSK4, poly(I:C), CL264 and Hygromycin B Gold were obtained from InvivoGen. P. alecto ASC-specific monoclonal antibody (mouse IgG2b) was generated by GenScript’s monoclonal antibody service. Rabbit polyclonal anti-ASC (AL177) (human/mouse) was purchased from Adipogen. Goat polyclonal anti-dog IL-1β (ab193852) (cross-reactive to P. alecto), rabbit polyclonal anti-mouse IL-1β (ab9722) and monoclonal antibody (mAb) to IAV nucleoprotein (ab20343) were from Abcam. mAb to β-actin (A2228) was from Sigma-Aldrich and mAb to GFP and variants (including mCitrine) were from Roche (11814460001). Anti-dsRNA mAb J2 was purchased from SCICONS and anti-mouse/rabbit/goat/monkey horseradish peroxidase (HRP)-conjugated secondary antibodies were from Santa Cruz. mPlum antibody from Origene was used to stain mPlum. Human mAb to MERS-CoV spike glycoprotein (m336) was provided by D. S. Dimitrov (NIH)58 (link). Alexa 488/568/647 Zenon labelling kits (Invitrogen) were used for direct labelling of antibodies for confocal microscopy and ImageStream. Protein G agarose beads (Millipore) were used to purify P. alecto IL-1β-Fc cloned and expressed from HEK293T cells. Polyclonal macaque serum (#5244 D42, infected by PRV3M, produced in-house) was used for immunoblot analysis.
Lps b5 ultrapure
Manufactured by InvivoGen
Sourced in United States
LPS-B5 Ultrapure is a lipopolysaccharide (LPS) product from InvivoGen. LPS is a major component of the outer membrane of Gram-negative bacteria and is known to be a potent activator of the innate immune response. The LPS-B5 Ultrapure product is a highly purified form of LPS derived from Escherichia coli 055:B5.
Automatically generated - may contain errors
Lab products found in correlation
Pam3csk4, by InvivoGen (3 mentions)
Sp600125, by Merck Group (2 mentions)
Ab20343, by Abcam (1 mentions)
Nigericin, by InvivoGen (1 mentions)
Hygromycin b gold, by InvivoGen (1 mentions)
Anti dsrna mab j2, by Techcomp Instruments (1 mentions)
Protein g agarose beads, by Merck Group (1 mentions)
Ab9722, by Abcam (1 mentions)
Lta bs, by InvivoGen (1 mentions)
Luciferase assay system, by Promega (1 mentions)
L cysteine hydrochloride, by Merck Group (1 mentions)
Anaerobic chamber, by Coy Laboratory Products (1 mentions)
Bhi broth, by BD (1 mentions)
Bhi agar, by BD (1 mentions)
4 octyl itaconate, by Merck Group (1 mentions)
Polyethyleneimine, by Thermo Fisher Scientific (1 mentions)
B2551, by Merck Group (1 mentions)
Ki696, by MedChemExpress (1 mentions)
D5796, by Merck Group (1 mentions)
N n dimethylformamide (dmf), by Merck Group (1 mentions)
11 protocols using lps b5 ultrapure
1
Bat Immune Signaling Reagent Characterization
2
Sepsis Induction and Survival Analysis in Mice
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Septic shock was induced in mice via i.p. injection with ultrapure LPS-B5 (20 mg/kg) (In vivoGen) prepared from E. coli 055:K59(B5). Mice were weighed and sepsis scores were determined at various intervals. The MSS scoring method assesses sepsis severity with scores ranging from 1-4 based on 7 parameters (appearance, consciousness, activity, stimulus (sounds/touch), eyes aspect, respiration rate and respiration quality) (27 (link)). Survival analysis was conducted using the Kaplan-Meier analysis method and log-rank (Mantel-Cox test) was used to determine statistical differences in survival.
3
Luciferase Assay for Cellular Signaling
4
Human TNFα DuSet ELISA Reagents
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human TNFα DuSet ELISA kit (R&D Systems, Catalog Number: DY210), recombinant human ApoM protein (R&D Systems, Catalog Number: 4550-AM-050) and recombinant human LL-37 (TOCRIS, Catalog Number: 5213). Highly purified endotoxins Salmonella typhimurium, E. coli B55, Salmonella minnesota and Vibrio cholerae were described previously [29 (link)] and new lots of endotoxins were purchased from Invivogen; SM Ultrapure (Invivogen, Catalog Number: tlr1-sm lps) and LPS-B5 Ultrapure (Invivogen, Catalog Number: tlrl-pb5lps). Commercial HDL derived from human plasma was purchased from (Sigma-Aldrich, Catalog Number: L8039. St. Louis, USA). Pooled human plasma HDL and LDL fractions were a kind gift from Dr Ngoc-Anh Le (Atlanta Veterans Affairs Medical Center, Decatur, GA; and Emory University School of Medicine, Atlanta, GA, USA).
5
Bacterial Cultivation and LPS Extraction Protocols
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
See Table S1 in the supplemental material for a full list of bacterial strains and plasmids used in this study. All Bacteroides strains were cultured anaerobically at 37°C in peptone-yeast extract-glucose (PYG) liquid medium or brain heart infusion (BHI) agar (BD Biosciences) with defibrinated horse blood (Hardy
Diagnostics) added to 10% vol/vol. The B. thetaiotaomicron transposon mutants were grown in liquid and agar media supplemented with 25 μg/ml erythromycin. B. thetaiotaomicron strains needed for large-scale LPS extraction were grown in BHI broth (BD Biosciences) supplemented with 5 μg/ml hemin and 500 μg/mll -cysteine hydrochloride (Sigma-Aldrich). The gas mix for the anaerobic chamber (Coy Laboratory Products) was 5% hydrogen and 20% carbon dioxide, balanced with nitrogen (Airgas). The E. coli strains used for cloning the pExchange-tdk knockout constructs were cultured in Luria broth (LB) or agar supplemented with carbenicillin. LPS from E. coli O55:B5 and E. coli MG1655 was purchased from InvivoGen (LPS-B5 Ultrapure and LPS-K12 Ultrapure, respectively).
Diagnostics) added to 10% vol/vol. The B. thetaiotaomicron transposon mutants were grown in liquid and agar media supplemented with 25 μg/ml erythromycin. B. thetaiotaomicron strains needed for large-scale LPS extraction were grown in BHI broth (BD Biosciences) supplemented with 5 μg/ml hemin and 500 μg/ml
6
TLR4 Pathway Stimulation with LPS-B5
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
TLR4 signaling pathway was stimulated with its synthetic ligand LPS-B5 Ultrapure (Invivogen San Diego, CA, United States). Previous studies have used this ligand to trigger TLR4 pathway to combat infections[12 (link)]. Cell culture supernatant was collected from HepG2.2.15 cells after treatment with 1, 2 and 4 μg/mL of LPS-B5 Ultrapure for 72 h.
7
Macrophage Polarization Modulation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
L929 cells were obtained from American Type Culture Collection and maintained in Dulbecco’s modified Eagle’s medium (DMEM; D5796, Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; 97068-085, VWR). BMDMs were generated from bone marrow and cultured on petri plates in DMEM containing 10% FBS and 30% L929 culture media for 7 days. PerMacs were collected from peritoneal gavages 4 days after intraperitoneal injection of 3% brewer thioglycolate medium (B2551, Sigma-Aldrich). Transfection of ISD (InvivoGen) into the cytosol of BMDMs was performed using polyethyleneimine (43896, Alfa Aesar). Unless stated, 6 × 105 BMDMs and 1.2 × 106 PerMacs per milliliter were used in in vitro experiments. The LPS-B5 Ultrapure (InvivoGen) concentration used was 200 ng/ml for BMDMs and 20 ng/ml for PerMacs unless otherwise indicated. The 4-octyl-itaconate (Cayman) concentration used was 125 μM, the DMF (Sigma-Aldrich) concentration was 50 μM, and the KI696 (MedChemExpress) concentration was 20 μM. Cells were exposed to both treatments 6 hours before LPS stimulation.
8
Modulating Macrophage Inflammatory Responses
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Murine peritoneal exudate macrophages or human THP-1 cell-derived macrophages were stimulated with 0.1–1 μg/ml of SARS-CoV-2 spike recombinant protein S1 subunit (Arigo Biolaboratories, Hsinchu City, Taiwan), 100 ng/ml of a TLR4 agonist LPS from Escherichia coli 055:B5 (LPS-B5 Ultrapure; InvivoGen, San Diego, CA, USA), or 10 ng/ml of a TLR2 agonist Pam2CSK4 (InvivoGen) to induce pro-inflammatory responses. Cells were treated with 2 μM BAY 11-7082 (Abcam, Cambridge, UK), 10 μM SP600125 (Sigma-Aldrich), 0.1 or 1 μg/ml of LPS from Rhodobacter sphaeroides (LPS-RS Ultrapure; InvivoGen), or 5 μg/ml of anti-murine or anti-human TLR2 neutralizing antibodies (InvivoGen) to block NF-κB, c-Jun N-terminal kinase (JNK), TLR4, or TLR2 signaling, respectively. The final concentrations of vehicles (H2O or dimethyl sulfoxide) to dissolve these agents were equivalent (less than 0.1%) in the culture medium among experimental groups.
9
Investigating Immune Responses with Chemical Modulators
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DMSO (Sigma‐Aldrich #D2650), DMXAA (SelleckChem #S1537), 2′3′‐cGAM(PS)2 (InvivoGen #tlrl‐nacga2srs), LPS‐B5 Ultrapure (InvivoGen #tlrl‐pb5lps), Pam3CSK4 (InvivoGen #tltl‐pms), TAK243 (SelleckChem #S8341), H‐151 (InvivoGen #inh‐h151), doxycycline (Sigma‐Aldrich #D9891).
10
Biochemical Inhibition of NLRP3 Inflammasome
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All chemicals and reagents were purchased from Sigma (St. Louis, MO, USA) and Invitrogen (CA, USA) unless specified otherwise. ATP, N-acetyl-L-cysteine (NAC), diphenyleneiodonium chloride (DPI), U73122, BAPTA-AM, Glyburide, Bay 11-7082, and sp600125 were all from Sigma. Monosodium urate (MSU) was from Invitrogen. Gallein was from Tocris Bioscience (Bristol UK). U0126 was from Promega (Madison, WI, USA). SB203580 was from Cayman Chemical (Ann Arbor, MI, USA). z-YVAD-fmk (ALX-260-074) and Ac-YVAD-CHO (ALX-260-027) were from Enzo Life Sciences (Farmingdale, NY, USA). Muramyl dipeptide (MDP), Pam3CSK4, FLA-ST, and LPS-B5 Ultrapure were obtained from InvivoGen (San Diego, CA, USA). AZD9056 was from MedChemExpress (Princeton, NJ, USA). Monoclonal antibodies (mAbs) used for Western blotting included: anti-caspase-1 (D3U3E) and anti-human IL-1β (D7F10) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-ASC (AL177) was from AdipoGen (San Diego, CA, USA); anti-NLRP3 (ALX-804-819) was from Enzo Life Sciences; Anti-β-actin mAb was purchased from BD Biosciences (San Jose, CA, USA). Anti-GAPDH mAb was from Proteintech (IL, USA). Abs used for cell stimulation (2A1 and mouse IgG1 control) and for the detection of signaling molecules have been described previously (24 (link)).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!