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25-HVD3(25-Hydroxy Vitamin D3) ELISA Kit

Cat.No.:E-EL-0015
Reactivity: Universal

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96T
48T
24T
96T*5
96T*10

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Product Details

Properties

Assay type	Competitive-ELISA
Format	96T/48T
Assay time	2.0h
Detection range	3.13-200 ng/mL
Sensitivity	1.88 ng/mL
Sample type &Sample volume	serum, plasma and other biological fluids; 50μL
Specificity	This kit recognizes 25-HVD3 in samples. No significant cross-reactivity or interference between 25-HVD3 and analogues was observed.
Reproducibility	Both intra-CV and inter-CV are < 10%.
Application	This ELISA kit applies to the in vitro quantitative determination of 25-HVD3 concentrations in serum, plasma and other biological fluids.

Test Principle

This ELISA kit uses the Competitive-ELISA principle. The micro ELISA plate provided in this kit has been pre-coated with 25-HVD3. During the reaction, 25-HVD3 in the sample or standard competes with a fixed amount of 25-HVD3 on the solid phase supporter for sites on the Biotinylated Detection Ab specific to 25-HVD3. Excess conjugate and unbound sample or standard are washed away, and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well and incubated. Then a TMB substrate solution is added to each well. The enzyme-substrate reaction is terminated by the addition of stop solution and the color turns from blue to yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The concentration of 25-HVD3 in tested samples can be calculated by comparing the OD of the samples to the standard curve.

Kit components & Storage

An unopened kit can be stored at 2-8℃ for 1 month. If the kit is not supposed to be used within 1 month, store the components separately according to the following conditions once the kit is received.

Item	Specifications	Storage
Micro ELISA Plate(Dismountable)	96T: 8 wells ×12 strips 48T: 8 wells ×6 strips	-20℃, 6 months
Reference Standard	96T: 2 vials 48T: 1 vial
Concentrated Biotinylated Detection Ab (100×)	96T: 1 vial, 120 μL 48T: 1 vial, 60 μL
Concentrated HRP Conjugate (100×)	96T: 1 vial, 120 μL 48T: 1 vial, 60 μL	-20℃(Protect from light), 6 months
Reference Standard & Sample Diluent	1 vial, 20 mL	2-8°C, 6 months
Biotinylated Detection Ab Diluent	1 vial, 14 mL
HRP Conjugate Diluent	1 vial, 14 mL
Concentrated Wash Buffer (25×)	1 vial, 30 mL
Substrate Reagent	1 vial, 10 mL	2-8℃(Protect from light)
Stop Solution	1 vial, 10 mL	2-8℃
Plate Sealer	5 pieces
Manual	1 copy
Certificate of Analysis	1 copy

Technical Data

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level 25-HVD3 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level 25-HVD3 were tested on 3 different plates, 20 replicates in each plate.

	Intra-assay Precision			Inter-assay Precision
Sample	1	2	3	1	2	3
n	20	20	20	20	20	20
Mean (ng/mL)	9.44	28.15	88.19	9.08	27.46	85.89
Standard deviation	0.62	1.47	2.88	0.52	1.21	4.08
CV (%)	6.57	5.22	3.27	5.73	4.41	4.75

Recovery

The recovery of 25-HVD3 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.

Sample Type	Range (%)	Average Recovery (%)
Serum(n=8)	87-100	92
EDTA plasma(n=8)	86-99	92
Cell culture media(n=8)	95-112	102

Target Information

Research Area

Metabolism, Developmental biology

Assay Procedures

	1. Add 50μL standard or sample to the wells, immediately add 50μL Biotinylated Detection Ab working solution to each well. Incubate for 45 min at 37°C
	2. Aspirate and wash the plate for 3 times
	3. Add 100μL HRP conjugate working solution. Incubate for 30 min at 37°C. Aspirate and wash the plate for 5 times
	4. Add 90μL Substrate Reagent. Incubate for 15 min at 37°C
	5. Add 50μL Stop Solution
	6. Read the plate at 450nm immediately. Calculation of the results

Citations

BIOCHEMICAL PHARMACOLOGY (2022) IF: 6.1
Pterostilbene-isothiocyanate impedes RANK/TRAF6 interaction to inhibit osteoclastogenesis, promoting osteogenesis in vitro and alleviating glucocorticoid induced osteoporosis in rats

DOI: 10.1016/j.bcp.2022.115284

PMID: 36209841
Sample: serum
ACS Applied Polymer Materials (2019) IF: 4.089
Hydrophilic and Insoluble Electrospun Cellulose Acetate Fiber-Based Biosensing Platform for 25-Hydroxy Vitamin-D3 Detection

DOI: 10.1021/acsapm.9b00179

Sample: serum
CALCIFIED TISSUE INTERNATIONAL (2022) IF: 4.333
Lactobacillus rhamnosus GG Promotes Intestinal Vitamin D Absorption by Upregulating Vitamin D Transporters in Senile Osteoporosis

DOI: 10.1007/s00223-022-00975-z

PMID: 35616697
Sample: plasma
BIOMEDICINE & PHARMACOTHERAPY (2020) IF: 4.545
A prebiotic, short-chain fructo-oligosaccharides promotes peak bone mass and maintains bone mass in ovariectomized rats by an osteogenic mechanism

DOI: 10.1016/j.biopha.2020.110448

PMID: 32776872
Sample: Serum
NEUROSCIENCE LETTERS (2022) IF: 3.046
Chronic vitamin D3 supplementation alleviates cognition impairment via inhibition of oxidative stress regulated by PI3K/AKT/Nrf2 in APP/PS1 transgenic mice

DOI: 10.1016/j.neulet.2022.136725

PMID: 35697158
Sample: cortex tissue
Frontiers in Neuroscience (2018) IF: 3.877
Postnatal Vitamin D Intake Modulates Hippocampal Learning and Memory in Adult Mice

DOI: 10.3389/fnins.2018.00141

Sample: Serum
Annals of Translational Medicine (2021) IF: 3.932
Conjugated linoleic acid prompts bone formation in ovariectomized osteoporotic rats and weakens osteoclast formation after treatment with ultraviolet B

DOI: 10.21037/atm-21-934

PMID: 33850900
Sample: Serum
NUTRITION RESEARCH (2019) IF: 2.627
Brazil nut intake increases circulating miR-454-3p and miR-584-5p in obese women

DOI: 10.1016/j.nutres.2019.05.004

Sample: serum
ARQUIVOS DE NEURO-PSIQUIATRIA (2019) IF: 1.048
Vitamin D, vitamin D binding protein, vitamin D receptor levels and cardiac dysautonomia in patients with multiple sclerosis: a cross-sectional study

DOI: 10.1590/0004-282x20190182

Sample: serum
JOURNAL OF PEDIATRIC ENDOCRINOLOGY & METABOLISM (2019) IF: 1.239
Determinants for low bone mineral density in pre-school children: a matched case-control study in Wuhan, China

DOI: 10.1515/jpem-2018-0554

Sample: serum
JOURNAL OF INTERNATIONAL MEDICAL RESEARCH (2020) IF: 1.287
Bacillus Calmette-Guérin (BCG) stimulates changes in dendritic cell surface marker expression in vitamin D-deficient mice:

DOI: 10.1177/0300060519896892

Sample: Serum
Cureus (2021)
The Association Between Ferritin and Vitamin D Levels in Premenopausal Fibroid Uterus Cases With Anemia

DOI: 10.7759/cureus.13392

Sample: serum
Turkish Journal of Biochemistry-Turk Biyokimya Dergisi (2021) IF: 0.401
Increased vitamin D binding protein levels are associated with irritable bowel syndrome

DOI: 10.1515/tjb-2020-0305

Sample: serum

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