Free fatty acid (FFA) levels in each conditioned medium in the co-culture experiments were measured by an enzymatic colorimetric method (NEFA-SS EIKEN kit; Eiken Chemical, Tokyo, Japan), and ketone bodies were measured by an enzymatic cycling method (Kainos ketone kit; Kainos Laboratories, Tokyo, Japan). These measurements were performed by SRL (Tokyo, Japan), an independent testing laboratory. Concentrations of adiponectin were determined with adiponectin human ELISA kit (Abcam) according to the manufacturers’ protocols.
Human adiponectin elisa kit
Manufactured by Abcam
Sourced in United States
The Human Adiponectin ELISA kit is a quantitative assay designed to measure the concentration of adiponectin, an adipokine protein, in human biological samples. It utilizes the enzyme-linked immunosorbent assay (ELISA) technique to detect and quantify the target analyte.
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Lab products found in correlation
Nefa ss eiken kit, by Eiken Chemical (1 mentions)
Human lipocalin 2 ngal quantikine elisa kit, by R&D Systems (1 mentions)
Human lgals3 galectin 3 elisa kit, by Merck Group (1 mentions)
Cobas e411, by Roche (1 mentions)
Qdr 4500a, by Hologic (1 mentions)
Milliplex map human metabolic hormone magnetic bead panel, by Merck Group (1 mentions)
Quantikine elisa kit, by R&D Systems (1 mentions)
Cobas e311, by Roche (1 mentions)
6 protocols using human adiponectin elisa kit
1
Quantifying Adipokine Levels in Co-culture
2
Adiponectin Levels in Cancer Patients
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For laboratory tests, overnight fasting blood samples were provided by the Institutional Tissue Bank at FUSCC (for cases). After the questionnaire interview and physical measurements, 10 ml of overnight fasting blood samples were collected for long-term storage in Taizhou Longitudinal Study (for controls).17 (link) For a subset of 305 cases and 330 matched controls who had genotype data during a similar time period, plasma adiponectin levels were measured with commercial enzyme-linked immunosorbent assay (ELISA) kit (Abcam Adiponectin Human ELISA Kit, USA) according to the manufacturer's manuals. The minimum detectable dose was 0.7 ng ml−1. The intra- and inter-assay coefficients of variation were 4.5% and 8.7%, respectively.
3
Quantification of Kidney Injury Biomarkers
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Daily megalin excretion was estimated using Human LRP2/Megalin ELISA assay (LS-F11978-1, BD BioSciences, Warsaw, Poland). The concentrations of NGAL, Gal-3, MMP12, and TIMP2 were estimated using Human Lipocalin-2/NGAL Quantikine ELISA Kit (R&D Systems, Minneapolis, MN), Human LGALS3/Galectin-3 ELISA Kit (RAB0661, Sigma-Aldrich, Saint Louis, MO), Human MMP-12 ELISA Kit (P39900, Invitrogen, Carlsbad, CA), and TIM2 Human ELISA Kit (P16035, ThermoFisher Scientific, Waltham, US-MA), respectively.
Finally, serum adiponectin concentration was assessed with the use of Human Adiponectin ELISA kit (ab99968, Abcam, Cambridge, UK).
All samples from patients and controls were run in duplicate. The assays were performed according to the manufacturer’s instructions. The optical density (OD) value was determined twice, using a microplate reader, set to 450 nm and 540 nm as a wavelength correction. The readings at 540 nm were subtracted from the readings at 450 nm to correct for optical imperfections in the plate. The concentrations of NGAL, Gal-3, MMP12, TIMP2, and adiponectin were calculated based on the standard curve. These standards were also assayed in duplicate.
Finally, serum adiponectin concentration was assessed with the use of Human Adiponectin ELISA kit (ab99968, Abcam, Cambridge, UK).
All samples from patients and controls were run in duplicate. The assays were performed according to the manufacturer’s instructions. The optical density (OD) value was determined twice, using a microplate reader, set to 450 nm and 540 nm as a wavelength correction. The readings at 540 nm were subtracted from the readings at 450 nm to correct for optical imperfections in the plate. The concentrations of NGAL, Gal-3, MMP12, TIMP2, and adiponectin were calculated based on the standard curve. These standards were also assayed in duplicate.
4
Comprehensive Metabolic Profiling Using DXA and WBC
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Body fat was measured by dual energy x-ray absorptiometry (DXA) (QDR 4500A, fan-beam densitometer, software version 8.21; Hologic, Waltham, MA). Screening fasting blood samples were sent to the National University Hospital, Singapore referral laboratory for biochemical assessment of liver, renal and thyroid function. Fasting and 2-hour postintervention blood samples during the whole-body calorimetry (WBC) session were collected from venous sampling. Plasma concentration of insulin was measured using immunoassay analyzer (Cobas e411, Roche). Plasma concentrations of high-density lipoprotein (HDL) cholesterol, triglycerides, and glucose were measured using a clinical chemistry analyzer (Cobas e311, Roche). Plasma FGF21 was measured using the Quantikine ELISA Kit (R & D System, Minneapolis, MN). Plasma adiponectin concentration was measured using a human adiponectin ELISA Kit (Abcam, Cambridge, UK). Plasma leptin and TNFα were measured using the MILLIPLEX MAP Human Metabolic Hormone Magnetic Bead Panel (Merck, MO).
5
Serum Adiponectin Quantification by ELISA
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Serum Adiponectin was assayed using a solid phase enzyme-linked immunosorbent assay. The assay was done using Abcam’s Human Adiponectin ELISA kit which measures the quantity of adiponectin in serum.
6
Measuring Soluble Adipokines
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Soluble leptin and adiponectin were detected in reserved 2-chamber system medium, or adipocyte medium from monoculture experiments. Frozen samples were thawed and centrifuged before the assay was performed. Leptin was analyzed using a Human Leptin ELISA Kit following the manufacturer’s protocol (Abcam, ab179884). Adiponectin was analyzed using Human Adiponectin ELISA Kit following the manufacturer’s protocol (Abcam, ab99968).
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