Synovial tissues or BMM homogenates were collected according to the commercial kit instructions (KGE004B, R&D Systems, USA). Standard ELISA kits were used to measure PGE2 (KGE004B, R&D Systems, USA), COX2 (DYC4198-2, R&D Systems, USA), and RGS12 (MBS9326858, MYBIOSOURCE, USA). The plate wells were measured at 450 nm, and the OD readings for the duplicate wells were averaged.
Kge004b
Manufactured by R&D Systems
Sourced in United States, United Kingdom
The KGE004B is a laboratory equipment product. It is designed for general laboratory use.
Automatically generated - may contain errors
Lab products found in correlation
D6050, by R&D Systems (2 mentions)
Elisa kit, by R&D Systems (2 mentions)
Countess cell counter, by Thermo Fisher Scientific (2 mentions)
Bca protein detection kit, by Thermo Fisher Scientific (1 mentions)
Griess reagent system, by Promega (1 mentions)
Quickplex sq 120, by Mesoscale (1 mentions)
U plex kit, by Mesoscale (1 mentions)
Ca6000, by R&D Systems (1 mentions)
Infinite m200 pro, by Tecan (1 mentions)
Exo spin midi columns, by Cell Guidance Systems (1 mentions)
Griess assay kit, by Beyotime (1 mentions)
Kge006b, by R&D Systems (1 mentions)
Dyc4198 2, by R&D Systems (1 mentions)
Ab267646, by Abcam (1 mentions)
Opteia human il 6 elisa set 555220, by BD (1 mentions)
Reagent set b, by BD (1 mentions)
Enzchek gelatinase collagenase assay kit, by Thermo Fisher Scientific (1 mentions)
Monosodium iodoacetate, by Merck Group (1 mentions)
Indomethacin, by Merck Group (1 mentions)
Elisa kit, by Elabscience (1 mentions)
37 protocols using kge004b
1
Synovial Tissue and BMM Analysis
2
Quantifying Inflammatory Markers in Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Nitric Oxide (NO) release was assessed using the Griess assay based on quantification of nitrite (NO2), the stable product of NO degradation. Nitrite content was quantified against a sodium nitrite standard curve using the Galaxy Fluorstar spectrophotometer (BMG Labtech, UK). An immunoassay kit (KGE004B, R&D Systems, UK) was used to quantify PGE2 concentrations in the media according to the manufacturer's instructions. Results were corrected for non-specific binding and calibrated using a PGE2 standard curve. The release of sulphated glycosaminoglycan (sGAG) into the culture media was quantified using the dimethylmethyleneblue (DMMB) assay against a chondroitin sulphate standard curve (6-sulphate:4-sulphate; 0.33:1; Sigma–Aldrich).
3
Quantifying PGE2 Levels via ELISA
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
PGE2 enzyme‐linked immunosorbent assay (ELISA) kit was obtained (KGE004B, R&D Systems) for investigating the presence of PGE2 among all treatment groups. Conditioned media from all trials were diluted 3‐fold with calibrator diluent for use. A competitive horseradish‐peroxide (HRP) enzyme probe reporter was used for its high turnover rate and specific enzyme activity. All absorbance values were read using a spectrophotometer (PerkinElmer, Waltham, MA) set to 450 nm for the main reading, and 570 nm for background noise detection.
4
Quantifying PGE2 Levels in Nevus Samples
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
PGE2 content in plasma samples was determined by ELISA using a PGE2 assay kit (KGE004B, R&D Systems, Minneapolis, MN, USA) according to the manufacturer’s instructions as described previously [22 (link)]. For nevus samples, frozen tissue was cut into sub-millimeter pieces and then homogenized in buffer (RD-556) supplied in the assay kit, using a disposable pestle (Kimble 749521-0590) obtained from Sigma-Aldrich, St. Louis, MO) on ice. After homogenization, nevus tissue lysates were sonicated briefly and then microfuged at 10,000 rpm for 10 min at 4 °C. After centrifugation, supernatants were collected, and protein concentrations were determined using a BCA protein detection kit (Thermo Fisher Scientific, Waltham, MA, USA). PGE2 values were normalized to protein content for each sample.
5
Quantifying PGE2 Levels via ELISA
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The level of PGE2 in supernatants or serum of the mice was measured by ELISA technique (KGE004B; R&D Systems, MN, United States), following the manufacturer’s instructions.
6
Quantifying Secretion of GF Factors
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To quantify the secretion of MMP1, IL6, IL8 and the production of PGE2 from human GFs, ELISA kits for MMP1 (RayBiotech, #ELH-MMP1−1), IL6 (R&D Systems, D6050), IL8 (R&D Systems, D8000C) and PGE2 (R&D Systems, KGE004B) were used. Briefly, human GFs were cultured without serum under each experimental set of conditions. The culture medium was collected and centrifuged for 10 min at 1200 × g, and the supernatant fractions were stored at ‑80 °C until analysis. The MMP1, IL6, IL8, and PGE2 levels were determined following the manufacturer’s instructions.
7
Equine Synovial Fluid and Serum Biomarkers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Blood samples were collected from the jugular vein at baseline and day 84 of supplementation to assess the effect of NXT15906F6 on the levels of RF, ANA, ACC peptide, and proinflammatory cytokines, including IL‐1β, IL‐6, TNF‐α and PGE2. The synovial fluid samples were collected from selected horses (eight animals from each group, including three females) by synoviocentesis aseptically using a 22‐gauge needle from the affected joint for analysis of routine cytology and for the estimation of cytokines, including IL‐1β, IL‐6, TNF‐α, and PGE2. Biomarkers in serum and synovial fluid were analyzed using commercially available kits (Rheumatoid factor: #Cat MBS006299, ACC peptide: #Cat MBS9344438, and IL‐1β: #Cat MBS282065, MyBioSource, San Diego, USA, ANA: #Cat E1384541 [Type II], SiNCERE Biotech, New Taipei City, Taiwan, IL‐6: #Cat EKC41235, Biomatik, Wilmington DE, USA, TNF‐α: #Cat LS‐F32050‐1, LS Bio, Seattle, WA, USA, and PGE2, #Cat KGE004B: R&D Systems, Minneapolis, MN, USA). The assay sensitivities of ACC peptide, IL‐1β, IL‐6, TNF‐α, RF, ANA, and PGE2 were 0.1 ng/ml, 15.6 pg/ml, 0.78 pg/ml, 9.375 pg/ml, 1.0 ng/ml, 0.054 ng/ml, and 41.4 pg/ml, respectively.
8
Quantification of Nitric Oxide and Inflammatory Cytokines
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
NO production was analysed as the accumulation of nitrite in culture supernatants using the Griess Reagent System (Promega, Madison, WI, USA). The supernatant was mixed with the same volume of Griess reagent and incubated at room temperature for 5 min. To ensure accurate NO quantification, reference curves were prepared using nitrite standards. The concentration of nitrite was determined by measuring the absorbance using a microplate reader and compared to the nitrite standards reference curve. The serum levels of the pro-inflammatory cytokines, including interleukin-1 beta (IL-1β, MLB00C), interleukin-6 (IL-6, M6000B), tumour necrosis factor-alpha (TNF-α, MTA00B), and prostaglandin E2 (PGE2, KGE004B), were measured using ELISA kits (R&D Systems, Minneapolis, MN, USA), according to the manufacturer’s protocol.
9
Cytokine and Chemokine Profiling in Cell Cultures
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Supernatant from cultured cells and explants was used to analyse IL-6, IL-8, IL-10, IL-1α, IL-1ß, TNF-α, IL-4, MIP-1α, MIP-1ß, and RANTES production. Supernatant was collected at the following timepoints; baseline, following TLR3 treatment, and following TLR2 or TLR2/6 treatment. Supernatant was also analysed from single agonist treatment and from vehicle control samples. Quantification of the cytokines and chemokines were performed using the Meso Scale Discovery platform. U-Plex kits were used according to manufacturer's instructions, MSD (Meso Scale Diagnostics, Rockville, Maryland). Plates were read by the QuickPlex SQ 120 (Meso Scale Diagnostics).
Supernatant was also used to analyse PGE2 at the same timepoints. The PGE2 ELISA kit was used according to the manufacturer's guidelines (KGE004B, R&D Systems). Briefly supernatant was diluted 1:3 with the assay diluent and plated alongside the calibration curve standards (Range of 39–2,500 pg/ml). The optical density of each well and readings from the absorbance value of 450 nm was subtracted from 540 and 570 nm in order to obtain PGE2 concentrations.
Supernatant was also used to analyse PGE2 at the same timepoints. The PGE2 ELISA kit was used according to the manufacturer's guidelines (KGE004B, R&D Systems). Briefly supernatant was diluted 1:3 with the assay diluent and plated alongside the calibration curve standards (Range of 39–2,500 pg/ml). The optical density of each well and readings from the absorbance value of 450 nm was subtracted from 540 and 570 nm in order to obtain PGE2 concentrations.
10
Quantifying Secreted IL-6 and PGE2
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Secreted IL-6 and PGE2 were evaluated in the cell culture supernatant by commercially available ELISA kits (CA6000, R&D Systems, Minneapolis, MN. USA; KGE004B, R&D Systems) following the manufacturer's protocol. Samples were used undiluted, and each sample was tested in duplicates. The absorbance was measured (Tecan, Infinite M200 PRO) at 450 nm (with subtracted 540 nm absorbance). IL-6 and PGE2 concentrations were calculated based on the standard curve. Results are presented as pg/mL.
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!