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Elisas

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ELISAs (Enzyme-Linked Immunosorbent Assays) are a type of laboratory equipment used for the detection and quantification of specific proteins, hormones, antibodies, or other analytes in biological samples. ELISAs utilize the principle of antigen-antibody interactions to measure the concentration of target molecules in a sample.

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Lab products found in correlation

Spectramax 190, by Molecular Devices (1 mentions) Vacuette, by Greiner (1 mentions) Lypocheck, by Bio-Rad (1 mentions)

7 protocols using elisas

1

Serum Fibrinogen and Estradiol Analysis

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After 8 h overnight fasting, 15 ml venous blood samples were collected in evacuated tubes that were pre-treated with a silica gel clot activator (Vacuette, Greiner Bio-One). The whole blood was allowed to clot for 20 min at room temperature, and the clot was then removed by centrifugation at 2000× g for 10 min at room temperature. The serum was separated from the clot and stored at −80 °C prior to analysis. Plasma fibrinogen and estradiol were determined using enzyme-linked immunosorbent assays (ELISAs; DRG International Inc., Mountainside, NJ, USA), according to the manufacturer’s suggestions. All of the samples were processed in duplicate during the same assay session. The intra-assay coefficient of variation was <5%, and the inter-assay precision was <8%. Optical densities were read on a standard plate reader at 450 nm (SpectraMax 190; Molecular Devices, Sunnyvale, USA).
Izzicupo P., Di Blasio A., Di Credico A., Gaggi G., Vamvakis A., Napolitano G., Ricci F., Gallina S., Ghinassi B, & Di Baldassarre A. (2020). The Length and Number of Sedentary Bouts Predict Fibrinogen Levels in Postmenopausal Women. International Journal of Environmental Research and Public Health, 17(9), 3051.
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2

Ovarian Estradiol ELISA in Mice

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Blood was obtained from mice of all treatment groups during the collection of the ovaries. Sera were subjected to enzyme-linked immunosorbent assays (ELISAs, DRG International Inc., Springfield, New Jersey) for measurement of estradiol (analytical sensitivity was 10.6 pg/mL and both intra- and inter-assay coefficients of variation were below 15%). Estradiol was selected because it is the primary ovarian estrogen and previous studies showed that IAA treatment decreases estradiol levels (Jeong, Gao et al. 2016 (link), Gonsioroski, Meling et al. 2019 , Gonsioroski, Meling et al. 2021 (link)). Assays were run according to the manufacturer’s instructions. Lypocheck from Bio-Rad Laboratories was used as a control with known values in all ELISAs in this study.
Gonsioroski A., Laws M., Mourikes V.E., Neff A., Drnevich J., Plewa M, & Flaws J.A. (2022). Iodoacetic Acid Exposure Alters the Transcriptome in Mouse Ovarian Antral Follicles. Journal of environmental sciences (China), 117, 46-57.
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3

Hormone Assays in Euthanized Mice

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To measure the hormone concentrations for this study, sera were collected from the euthanized mice. The sera were then used for enzyme-linked immunosorbent assays (ELISAs, DRG International Inc., Springfield, New Jersey) to measure concentrations of estradiol, progesterone, and testosterone. Additionally, the levels of follicle-stimulating hormone (FSH) were measured by radioimmunoassay at the University of Virginia Center for Research in Reproduction Ligand Assay and Analysis Core. The lowest limits of detection were 0.083 ng/mL for testosterone, 0.034 ng/ml for progesterone, 10.6 pg/mL for estradiol, and 2 ng/mL for FSH. If the measurement was lower than the lowest limit of detection, the value was substituted with the lowest limit of detection/√2. The intra- and inter-assay coefficients of variability were less than 10%.
Gill S., Brehm E., Leon K., Chiu J., Meling D.D, & Flaws J.A. (2021). Prenatal exposure to an environmentally relevant phthalate mixture alters ovarian steroidogenesis and folliculogenesis in the F1 generation of adult female mice. Reproductive toxicology (Elmsford, N.Y.), 106, 25-31.
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4

Steroid Hormone Quantification in Follicle Culture

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Media were collected after 24 or 96 hours of follicle culture and then
subjected to enzyme-linked immunosorbent assays (ELISAs, DRG International Inc.,
Springfield, New Jersey) for pregnenolone, dehydroepiandrosterone (DHEA),
progesterone, androstenedione, testosterone, and 17β-estradiol. The lower
limit of detection for each assay was 0.05 ng/mL for pregnenolone, 0.07 ng/mL
for DHEA, 0.045 ng/mL for progesterone, 0.021 ng/mL for androstenedione, 0.083
ng/mL for testosterone, and 9.7 ng/mL for 17β-estradiol. The assays were
performed according to the manufacturer’s protocol. Standard curves for
each assay were created using provided standards and standard curves were
created using four-parameter logistic curve fitting. A mean value of duplicate
runs was calculated for all samples. Sample dilution was done with steroid free
serum (DRG International Inc.), if needed, for measurements to be in the range
of the standard curve.
Meling D.D., Warner G.R., Szumski J.R., Gao L., Gonsioroski A.V., Rattan S, & Flaws J.A. (2019). The Effects of a Phthalate Metabolite Mixture on Antral Follicle Growth and Sex Steroid Synthesis in Mice. Toxicology and applied pharmacology, 388, 114875.
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5

Steroid Hormone ELISA Quantification

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Culture media collected at 96 h were subjected to immunosorbent assays (ELISAs, DRG International Inc., Springfield, NJ, USA). Analytical sensitivities for progesterone, androstenedione, testosterone, and estradiol were 0.045, 0.019, 0.083, and 0.0097 ng/mL, respectively. Both intra- and inter-assay coefficients of variation were below 10%, except for androstenedione analyses which were below 15%. Samples were run in duplicate and diluted if needed to fit the dynamic range of the ELISA kits.
Santacruz-Márquez R., Flaws J.A., Sánchez-Peña L.D, & Hernández-Ochoa I. (2023). Exposure to Zinc Oxide Nanoparticles Increases Estradiol Levels and Induces an Antioxidant Response in Antral Ovarian Follicles In Vitro. Toxics, 11(7), 602.
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6

Antral Follicle Steroid Hormone Assay

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Culture media were collected at 96 h of culture from 3 separate antral follicle cultures, and subjected to enzyme-linked immunosorbent assays (ELISAs, DRG International Inc., Springfield, New Jersey) for measurement of progesterone (analytical sensitivity was 0.034 ng/ml and both intra- and inter-assay coefficients of variation were below 8%), pregnenolone (analytical sensitivity was 0.05 ng/ml and both intra- and inter-assay coefficients of variation were below 15%), androstenedione (analytical sensitivity was 0.021 ng/ml and both intra- and inter-assay coefficients of variation were below 10%), testosterone (analytical sensitivity was 0.083 ng/mL and both intra- and inter-assay coefficients of variation were below 10%), and estradiol (analytical sensitivity was 10.6 pg/mL and both intra- and inter-assay coefficients of variation were below 15%). Assays were run according to the manufacturer’s instructions. Samples were diluted if necessary and run in duplicate for measurements of progesterone (no dilution), pregnenolone (no dilution), androstenedione (no dilution), testosterone (no dilution), and estradiol (1:10 dilution) to match the dynamic range of each ELISA kit.
Gonsioroski A., Meling D.D., Gao L., Plewa M.J, & Flaws J.A. (2019). Iodoacetic Acid Inhibits Follicle Growth and Alters Expression of Genes that Regulate Apoptosis, the Cell Cycle, Estrogen Receptors, and Ovarian Steroidogenesis in Mouse Ovarian Follicles. Reproductive toxicology (Elmsford, N.Y.), 91, 101-108.
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7

Hormonal Profiling of Athlete Samples

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These data were a subset of those published previously (Luk et al., 2019 (link)). Serum samples from BL, IMD, 15, 30 and 60 were analyzed for testosterone (EIA1559; intra‐assay % CV = 7.12–9.85; inter‐assay % CV = 9.00), growth hormone (EIA1787; intra‐assay % CV = 8.02–14.23; inter‐assay % CV = 10.32), and cortisol (EIA1887R; intra‐assay % CV = 3.12–9.45; inter‐assay % CV = 5.95) concentrations using commercially available ELISAs (DRG Internation al Inc. Springfield, NJ).
Luk H., Jiwan N.C., Appell C.R., Levitt D.E, & Vingren J.L. (2022). Sex‐specific mitochondrial dynamics and mitophagy response to muscle damage. Physiological Reports, 10(10), e15230.
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