Human IGFBP-3 (insulin-like growth factor binding protein 3) ELISA Kit

Sample

Serum, plasma or other body fluids

Species Reactivity

Human

Detection Method

sELISA

Intended Use

This enzyme immunoassay kit is suited for measuring IGFBP-3 in human serum, EDTA and Heparin-plasma

Contents of Kit

1. Microtiter plate: ready for use, coated with rabbit-anti-hIGFBP-3-antibody. Wells are separately breakable. (8x12) wells
2. Calibrators (CAL A-E): lyophilized, (human IGFBP-3), concentrations are given on vial labels and on quality certificate. 5 x 1 mL
3. Control 1 (CTR1): lyophilized, (human serum), concentration is given on quality certificate. 1 x 250 μL
4. Control 2 (CTR2): lyophilized, (human serum), concentration is given on quality certificate. 1 x 250 μL
5. Antibody-HRP-Conjugate (DET): ready for use, contains rabbit biotinylated anti-hIGFBP-3 antibody. 1 x 12 mL
6. Sample Buffer (SB): red color, ready for use. Please shake before use! 1 x 120 mL
7. Dilution Buffer (DIL): ready for use. Please shake before use! 1 x 30 mL
8. Washing Buffer (WB): 20-fold concentrated solution. 1 x 50 mL
9. Substrate (S): ready for use, horseradish-peroxidase-(HRP) substrate, stabilised H₂O₂ Tetramethylbencidine. 1 x 12 mL
10. Stopping Solution (STP): ready for use, 0.2 M sulfuric acid. 1 x 12 mL
11. Sealing Tape: for covering the microtiter plate. 2

Storage

Store the kit at 2-8°C after receipt until its expiry date. The lyophilized reagents should be stored at –20 °C after reconstitution. Avoid repeated thawing and freezing.
The shelf life of the components after initial opening is warranted for 4 weeks, store the unused strips and microtiter wells airtight together with the desiccant at 2-8°C in the clip-lock bag, use in the frame provided. The reconstituted components Calibrators A-E and Controls CTR1 and CTR2 must be stored at –20°C (max. 4 weeks). For further use, thaw quickly but gently (avoid temperature increase above room temperature and avoid excessive vortexing). Up to 3 of the freeze-thaw cycles did not influence the assay. The 1:20 diluted Washing Buffer WP is 4 weeks stable at 2-8°C.

Precision

Intra-Assay Variance
One sample has been measured 10 times in the same assay. The measured coefficient of variation (CV) is on average 1.9%.

Inter-Assay-Variability
Serum samples were measured in independent assays on different days. On average the coefficient of variation was 5.7%.

Sensitivity

Sensitivity was assessed by measuring the blank and calculating the theoretical concentration of the 2fold standard deviation of the blank. The analytical sensitivity of Human IGFBP-3 ELISA Kit is 0.03 ng/mL. According ICH Q2 R1 (CPMP/ICH/381/95) the limit of quantification (LoQ) is reflected by the recalculated IGFBP-3 concentration of the 10fold standard deviation of the blank, which therewith is 0.15 ng/mL.

General Description

Insulin-like growth factors (IGF)-I and -II are bound to specific binding proteins (IGFBPs) in the circulation. To date, at least six binding proteins can be distinguished on the basis of their amino acid sequence. They are designated as IGFBP-1, IGFBP-2, ... IGPBP-6. The predominating IGFBP in blood is IGFBP-3, which largely determines the total IGF-I and IGF-II concentration. In contrast to the other binding proteins, IGFBP-3 has the property to associate with an acid-labile subunit (ALS) after binding of either IGF-I or IGF-II. Most of the IGFBP-3 in plasma is present as high molecular weight ternary complex, however, small amounts of free IGFBP-3 are also found.
The development of a specific immunoassays for IGFBP-3, which detects IGFBP-3 in the ternary complex, provided new in-sights into IGFP-3 regulation. On the basis of these findings serum IGFBP-3 has been proven to be an additional useful test in the repertoire of diagnostic tools for evaluation of growth disorders.
Several factors besides GH influence IGFBP-3 levels: age including sexual development, nutrition, hypothyroidism, diabetes mellitus, liver function and kidney function. IGFBP-3 levels are decreased by malnutrition, although less than IGF-I, in hypothyroidism, in diabetes mellitus and in hepatic failure, but are increased in chronic renal failure. Measurement over 24 hours revealed no circadian rhythm. For clinical practice, the most important regulatory factor is GH. Single IGFBP-3 measurements correlate significantly with the logarithm of the integrated spontaneous GH secretion. In patients with GH deficiency, IGFBP-3 levels are subnormal and increase gradually to within the normal range after several days of GH administration. The slow response to GH and constant circadian levels during chronic daily application of GH suggest that IGFBP-3 reflects the GH secretory state over days.
In normal tall children and adolescents without excessive GH secretion or in patients with Sotos syndrome, IGFBP-3 levels are normal or slightly increased. In contrast, children with pituitary gigantism or adults with acromegaly have clearly elevated levels that normalize on successful treatment. Therefore, IGFBP-3 is also a useful parameter for the detection of excessive GH secretion and monitoring therapy efficacy. In precocious puberty, IGFBP-3 levels are clearly increased by chronological age, whereas patients with premature thelarche have IGFBP-3 levels in the upper normal range.

Standard Curve

The following data is for demonstration only and cannot be used in place of data generation at the time of assay.

The exemplary shown calibration curve in Figure below cannot be used for calculation of your test results. You have to establish a calibration curve for each test you conduct!