Salivary concentrations of the four hormones were measured using commercial Linked Immunosorbent Assay (ELISA) kits: Oxytocin by ENZO (New-York, USA), CT and T by Salimetrics (Pennsylvania USA), and s-IgA by Euroimmun (Lubeck Germany) according to the kit's instructions and consistent with prior research55 (link),56 (link). Each kit provides a quantitative in vitro assay for the biomarker in human saliva. Measurements were done in duplicate according to the instructions recommended by the respective manufacturer. The concentration of each hormone in the sample was calculated by MEGELAN (Tecan, Germany) according to relevant standard curves. The intra-assay and inter-assay coefficients of samples as measured by the manufacturer's control are as follows: for OT less than 9.7% and 14.5%, for CT less than 4.7% and 9.3%, for T less than 5.97% and 12.1%, and for s-IgA less than 4.1% and 4%.
Oxytocin
Manufactured by Enzo Life Sciences
Sourced in United States, China, Germany
Oxytocin is a peptide hormone that is produced in the hypothalamus and secreted by the posterior pituitary gland. It plays a crucial role in various physiological processes, including the stimulation of uterine contractions during childbirth and the let-down reflex in breastfeeding.
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Lab products found in correlation
Testosterone, by Cusabio (1 mentions)
Corticosterone, by Cusabio (1 mentions)
Estradiol, by Cusabio (1 mentions)
Prolactin, by Cusabio (1 mentions)
Salivabio oral swab, by Salimetrics (1 mentions)
Cd8 pe clone sk1, by BD (1 mentions)
Primovert microscope, by Zeiss (1 mentions)
Cd4 pe cy7 clone sk3, by BD (1 mentions)
Facscanto 2, by BD (1 mentions)
Cd3 apc clone sk7, by Thermo Fisher Scientific (1 mentions)
Facsdiva software, by BD (1 mentions)
4 protocols using oxytocin
1
Salivary Hormone Biomarker Quantification
2
Maternal Immune Activation Alters Hormones
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Serum levels of these hormones in dam were determined at PND 7. Blood was collected via decapitation and serum was separated. The levels of testosterone (CUSABIO, Wuhan, China), estradiol (CUSABIO, Wuhan, China), progesterone (CUSABIO, Wuhan, China), prolactin (CUSABIO, Wuhan, China), corticosterone (CUSABIO, Wuhan, China), oxytocin (Enzo life sciences, PA, USA) and AVP (Enzo life sciences, PA, USA) were assessed according to the kit instruction. All the 8 control dams in part II were included in this section. But among the 9 MIA dams, we failed to get blood in 1 MIA dam, and no enough serum was separated from another MIA dam. Finally, the serum hormone analytes had n = 8 per group except testosterone and corticosterone had n = 7 for the MIA group.
3
Salivary Biomarker Collection and Analysis
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Salivary samples were taken from patients at the same time between 7:00—8:00 AM to reduce diurnal variation. Patients were instructed to keep off eating, drinking except water, and smoking at least 1 h before the collection and to rinse mouth thoroughly with water 10 min before sample preparation. Salivary samples (~ 2 ml per subject) were collected using SalivaBio Oral Swab (Item No. 5001.02, Salimetrics) and were frozen at -80 °C until further analysis within 4 h of collection. According to the manufacturer’s protocol, freezing saliva samples were vortexed and centrifuged at 3,000 rpm for 15 min. Salivary cortisol was analyzed using Salimetrics® Cortisol Enzyme Immunoassay (EIA) Kit (Item No.1–3002, Salimetrics) [33 (link)–35 (link)]. Salivary oxytocin levels were determined using a commercially available oxytocin enzyme-linked immunosorbent assay (ELISA) kit (Product Number: ADI-900-153A-0001, Enzo Life Sciences) following the instructions of the manufacturer. The saliva sample of each participant was assayed in duplicate by a microplate reader, and the mean of the two oxytocin values was calculated according to relevant standard curves provided by the manufacturers [36 (link), 37 (link)].
4
Stress-induced Biomarker Profiling
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Blood samples were drawn from the right arm before, from the left arm after the stress test using heparinized Vacutainer® vials, and centrifuged 15 min at 1000g, 4°C, and supernatants immediately stored at -80°C. Measurements were done in duplicates using enzyme-linked immunoassay (ELISA) kits according to the manufacturers′ instructions; noradrenaline and adrenaline: Labor Diagnostika Nord, Nordhorn, Germany (detection limits 20 pg/ml noradrenaline, 5.2 pg/ml adrenaline); serotonin: DLD Diagnostika, Hamburg, Germany (5 ng/ml); oxytocin: Enzo, Lausen, Switzerland (15 pg/ml); human platelet activation factor (PAF): Abbexa, Cambridge, United Kingdom (2.5 pg/ml); human prostaglandin D2 (PGD2): Bioassay Technology, Shanghai, China (5.02 ng/ml).
Blood cells were isolated from heparinized blood samples using Ficoll-Paque Plus 1.078 g/ml density gradient (GE Healthcare Biosciences, Uppsala, Sweden). PBMCs were deposited in a Neubauer chamber and counted using Primo Vert Microscope (Carl Zeiss, Germany). PBMC subpopulations were stained using the following fluorescent labelled antibodies: CD3-APC (clone SK7; eBioscience Inc.), CD4-PE-CY7 (clone SK3; BD Biosciences), CD8-PE (clone SK1; BD Biosciences) and CD14-FITC (clone MOPC-21; BioLegend), for 30 min at 4°C; after washing, 10.000 single cell events were acquired in BD FACSCanto II with FACSDiva Software (Becton Dickinson).
Blood cells were isolated from heparinized blood samples using Ficoll-Paque Plus 1.078 g/ml density gradient (GE Healthcare Biosciences, Uppsala, Sweden). PBMCs were deposited in a Neubauer chamber and counted using Primo Vert Microscope (Carl Zeiss, Germany). PBMC subpopulations were stained using the following fluorescent labelled antibodies: CD3-APC (clone SK7; eBioscience Inc.), CD4-PE-CY7 (clone SK3; BD Biosciences), CD8-PE (clone SK1; BD Biosciences) and CD14-FITC (clone MOPC-21; BioLegend), for 30 min at 4°C; after washing, 10.000 single cell events were acquired in BD FACSCanto II with FACSDiva Software (Becton Dickinson).
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