25-OH Vitamin D ELISA TEST KIT

INTENDED USE

     Detection Kit for 25-OH Vitamin D (Enzyme-Linked ImmunoSorbent Assay , ELISA).It is used in quantitative tests for Vitamin D in human serum.

SUMMARY AND CLINICAL SIGNIFICANCE

   Vitamin D is a fat-soluble hormone precursor that is produced primarily through skin exposure to sunlight. Vitamin D is biologically inert and must undergo a hydroxylation step to become active. Our bodies can only synthesize vitamin D3. Vitamin D2 can be taken through fortified foods or supplements. Physiologically, vitamins D3 and D2 bind to vitamin D binding protein (VDBP) in plasma and are transported to the liver to become 25-hydroxyvitamin D (25-OH vitamin D). Since 25-OH vitamin D is the major storage form, its blood concentration can be used to assess overall vitamin D status. More than 95% of the measurable 25-OH vitamin D in serum is 25-OH vitamin D3, while 25-OH vitamin D2 only reaches measurable levels in patients taking vitamin D2 supplements. Vitamin D is essential for bone health. In children, severe deficiency can lead to rickets. In the elderly, the risk of falls has been attributed to vitamin D deficiency due to muscle weakness. Additionally, low 25-OH vitamin D concentrations are associated with low bone density. Vitamin D insufficiency has also been linked to diabetes, cancer, cardiovascular disease, and autoimmune diseases. The two vitamin D dietary supplements currently on the market (vitamin D2 and vitamin D3) are converted to 25-OH vitamin D in the liver. The sum of the concentrations of 25-OH vitamin D2 and 25-OH vitamin D3 in serum or plasma is referred to as "total 25-OH vitamin D." In clinical settings, accurate monitoring of total 25-OH vitamin D levels is critical. Vitamin D-deficient patients who take daily vitamin D supplements should have their serum or plasma vitamin D levels monitored regularly to achieve optimal levels and prevent their 25-OH vitamin D concentrations from reaching levels that are considered toxic. The total vitamin D assay uses specific antibodies to capture 25-hydroxyvitamins D3 and D2. The test is designed to quantitatively measure total 25-OH vitamin D in human serum and plasma to aid in the assessment of vitamin D sufficiency.

PRINCIPLE

The 25-OH Vitamin D Quantitative Test Kit is based on a solid phase enzyme-linked immunosorbent assay. The assay system utilizes one antibody for solid phase (microtiter wells) immobilization and another monoclonal antibody in the solutions. The third antibody which recognize the second antibody conjugated with an enzyme (horseradish peroxidase). In the assay procedure, standards and test specimen (serum) are added to the 25-OH Vitamin D antibody coated microtiter wells, incubation 30 minutes together with the second antibody. After wash, the third antibody labeled with horseradish peroxidase (conjugate) is added. If human 25-OH Vitamin D is present in the specimen, it will combine with the antibody on the well and the enzyme conjugate resulting in forming sandwiches between the solid phase and enzyme-linked antibodies. After incubation, the wells are washed with water to remove unbound labeled antibodies. A solution of TMB is added and incubated for 20 minutes, resulting in the development of a blue color. The color development is stopped with the addition of Stop Solution.The color is changed to yellow and measured spectrophotometrically at 450 nm. The concentrations of Vitamin D is directly proportional to the color intensity of the test sample.

PREPARATIONS

1.  Allow all specimens and reagents to reach room temperature (~25°C) and mix thoroughly by gentle inversion before use.

2.Prepare Wash Buffer by diluting Wash Concentrate 20-fold with deionized water. The diluted wash solution is stable in room temperature for at least one week.

PRECAUTION FOR USERS

1. Handling should preclude any pipetting by mouth. Use only pipettes with disposable tips for each specimen.

2.Do not mix materials from different master lots. Do not use kit components beyond the expiration date.

3.All materials should be brought to room temperature before use.