Human IGFBP-1 (insulin-like growth factor binding protein 1) 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-1 in human serum, plasma or in other body fluids, for example, amnion fluid, mother´s milk, urine or saliva, as for scientific purposes. It is also suited to quantitate IGFBP-1 in cell culture media.

Contents of Kit

1. Microtiter plate: ready for use, coated with mouse-anti-IGFBP-1-antibody. Wells are separately breakable. (8x12) wells
2. Calibrators (CAL A-G): lyophilized, (native human IGFBP-1), concentrations are given on vial labels and on quality certificate. 7 x 500 μL
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 Conjugate (DET): ready for use, contains mouse biotinylated anti-hIGFBP-1 antibody. 1 x 6 mL
6. Enzyme Conjugate (EC): ready for use, contains HRP (Horseradish-Peroxidase)-labeled Streptavidin. 1 x 12 mL
7. Dilution Buffer (DIL): ready for use. Please shake before use! 1 x 125 mL
8. Washing Buffer (WB): 20-fold concentrated solution. 1 x 50 mL
9. Substrate (S): ready for use, horseradish-peroxidase-(HRP) substrate, stabilised 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-G 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
Serum samples were measured up to 20 times in the same assay and the coefficient was calculated based on the recalculated IGFBP-1 concentration. The mean measured coefficient of variation (CV) is 6.52% (n =6).

Inter-Assay-Variability
Serum samples were measured in independent assays. On average the coefficient of variation was 6.05% (SD 0.46).

Sensitivity

The analytical sensitivity was determined by measuring the null calibration (CAL-A) and calculating the corresponding concentration of the signal intensity of the CAL-A + 2SD. In three different assays analytical sensitivities from 0.03 to 0.08 ng/mL were measured with an average value of 0.055 ng/mL

General Description

The Insulin-like Growth Factors I and – II are free in body fluids and tissues but are bound to specific binding proteins. Until today seven different binding proteins (IGFBP-1 to –7) can be differentiated additionally several IGFBP-related proteins have also been detected. Bioavailability of IGF is regulated by these IGFBPs or their proteolytic cleavage which reduces affinity to IGF. The IGFBPs as well as their proteolytic fragments can also exert IGF-independent effects, like influencing cell migration or proliferation.
IGFBP-1 (Placental Protein 12) consists of 234 amino acids and has a molecular weight of approximately 25kDa. The coding DNA region is located on chromosome 7. IGFBP-1 is mainly synthesized by foetal and adult liver tissue and decidual endometrium. Intensity of Expression varies enduring menstruation with a maximal expression in the late secretory phase. Further IGFBP-1 expression seems to be regulated by Insulin concentration, with Insulin inhibiting the expression. Insulin regulation results in diurnal fluctuations of up to factor 10. IGFBP-1 is posttranslational modified by phosphorylation of serine residues 101, 119 and 169. Phosphorylation has physiological relevance as it increases affinity of IGFBP-1 to IGF. In adult humans phosphorylated IGFBP-1 of the liver is the predominant form in circulation. IGFBP-1 produced by endometrial tissue is significantly less phosphorylated than the liver originated form.
In pregnancy IGFBP-1 maternal serum concentration increases significantly with maximal values in the second trimester or 22-23 week of gestation (75.8 ng/ml) and decreases slowly until term. IGFBP-1 concentration is not only increased in maternal but also in foetal serum, extremely high concentrations are found in amniotic fluid. Here concentration can reach more than the 1000-fold of serum values. Long-term changes of serum IGFBP-1 concentration can also be found in amniotic fluid. After birth the IGFBP-1 level of the child decreases until it reaches the low steady-state level of puberty and adulthood.
Short term IGFBP-1 serum concentration is strongly influenced by nutrition level and therewith by insulin. Decreasing IGFBP-1 levels can be found enduring fasting or in diabetes; IGFBP-1 levels increase in case of intensive exercises.

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!