Vitamin D combi ImmuTube® LC-MS / MS - the vitamin D multiplex test

For the quantitative determination of 25-hydroxyvitamin D3 / 25-hydroxyvitamin D2 / 1,25-dihydroxyvitamin D3 / 1,25-dihydroxyvitamin D2 / 24,25-dihydroxyvitamin D3

Vitamin D is a group of prohormones, of which the main members are vitamin D3 (cholecalciferol) and vitamin D2 (ergocalciferol). Both are bioequivalent and are converted into hormonally active metabolites in vivo by identical metabolic pathways. Vitamin D3 is mainly formed in the epidermis of the skin from endogenous 7-dehydrocholesterol by the influence of sunlight.
The prohormones are transported to the liver by the vitamin D binding protein (VDBP) and converted into the biologically inactive 25-hydroxyvitamin D (25-OH-vitamin D) by a first hydroxylation. 25-hydroxyvitamin D is the main form of circulating vitamin D and is the parameter for measuring vitamin D status. The synthesis of the active 1,25-dihydroxyvitamin D (1,25-(OH)2-vitaminD) is carried out in a second hydroxylation step by 1alpha-hydroxylase mainly in the kidneys but also locally in many extra-renal tissues (paracrine and autocrine effect). 1,25-dihydroxyvitamin D binds to the intracellular vitamin D receptor (VDR), which is an ubiquitous transcription factor. As a result, 1,25-dihydroxyvitamin D regulates the gene expression of numerous proteins, inhibits cell proliferation and stimulates cell differentiation.
In the case of vitamin D deficiency, the 1,25-dihydroxyvitamin D serum level can be maintained within the normal range for a long time due to the resulting secondary hyperparathyroidism with stimuli of the renal 1alpha-hydroxylase. Vitamin D deficiency manifests much earlier in extra-renal tissues with low 1alpha-hydroxylase activity and lack of compensation mechanisms. Excess 1,25-dihydroxyvitamin D is converted by the 24-hydroxylase into water-soluble calcitroic acid, which is excreted via bile.
1,25-dihydroxyvitamin D has – together with calcium - a central importance for the mineral and bone metabolism. In the kidney, parathyroid hormone (PTH) stimulates the synthesis of 1,25-dihydroxyvitamin D, which increases calcium reabsorption in the distal renal tubules. On the other hand, 1,25-dihydroxyvitaminD inhibits PTH secretion via a feedback mechanism, which reduces the phosphate resorption in the proximal tubules and consequently the serum phosphate level. In the intestine, 1,25-dihydroxyvitamin D increases the calcium and phosphate resorption. In bone, 1,25-dihydroxyvitaminD affects the osteoblasts as well as the osteoclasts and thus contributes to the balance between bone anabolism and bone catabolism. In addition, 1,25-dihydroxyvitaminD is involved in the regulation of the following functions due to pleiotropic effects: cell proliferation and cell differentiation (influencing tumorigenesis by inhibition of proliferation, angiogenesis, cell invasion, and promotion of cell differentiation), hormone secretion (stimulation of the insulin secretion, suppression of renin synthesis), and immune function (stimulation of innate, unspecific immunity e. g. by induction of β-defensin synthesis, inhibition of adaptive, specific immunity and increased immune tolerance threshold by inhibition of maturation of dendritic cells).
Indications for the 25-hydroxyvitamin-D determination are: rickets, osteoporosis, osteomalacia, renal insufficiency, hepatic impairment, malabsorption, pregnancy, obesity, granulomatous diseases, therapy with barbiturates or anti-epileptic drugs, as well as with conspicuous infectious diseases, autoimmune diseases, tumor diseases, diabetes mellitus , fertility restrictions.
Indications for the determination of vitamin D metabolites are: differential diagnosis of hyper- and hypocalcemia and hypercalciuria, therapy control after 1,25-dihydroxyvitaminD substitution (the ratio of 24,25dihydroxyvitamin D to 25-hydroxyvitamin D can lead to the conclusion of supplementation success), chronic granulomatosis, congenital and acquired defects of the vitamin D metabolism.
The ImmuTube® LC-MS / MS kit combines the five vitamin D metabolites:
25-hydroxyvitamin D3 / 2  -  1,25-dihydroxyvitaminD3 / D2  -  24,25-dihydroxyvitamin D3
from 500 μl serum or plasma. It is the only method to differentiate between the vitamin D metabolites by a single measurement. Sensitivity and specificity are significantly higher than with ELISA or HPLC. By a single pre-analytical extraction step interfering matrix components are removed so that an ion suppression effect is avoided. Vitamin D combi ImmuTube® can be used flexibly on many high-end devices.

Vitamin D combi ImmuTube® LC-MS / MS Kit     KM1200

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Claudia Wandesleben / Gudrun Gutschalk
Phone: +49 (0)6251 - 70190-175 /-177
Fax: +49 (0)6251 - 84 94 30

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