The Eagle Biosciences IGFBP-3 LIA Assay Kit utilizes specific and high affinity antibodies for IGFBP-3, the antibodies are immobilized on the microtitre plate. The ligand, biotinylated IGF-I is pre-dispensed in excess into the needed wells. The sample is diluted outside within a special dilution buffer (Sample Buffer PP); all naturally bound IGFs are thereby released from their binding proteins. By adding an aliquot of such diluted sample with free IGFBP-3 to the ligand containing wells, the biotinylated IGF-I occupies all existing specific binding sites of the IGFBP-3. All IGFBP-3 molecules are bound afterwards to the microtitre plate, however by using a Strepavidin-Peroxydase-(POD)-Conjugate only the complexes of IGFBP-3/biotinylated IGF-I are involved in signal generation (in the closing POD-substrate reaction), thus, only functional IGFBP-3 is being quantified. Sample Types: serum, plasma. Range: 0.4 - 30 ng/mL. Sensitivity: 0.4 ng/mL. Time: 3.5 hours ? All currently existing IGFBP-3 immunoassays use the binding of specific anti-IGFBP-3 antibodies for signal generation and thus IGFBP-3 quantification. The failure of differentiation between complete IGFBP-3 molecules and their respective fragments (derived physiologically due to the different proteases activities) is unavoidable in this system. Because one molecule IGFBP-3 can be cleaved in several fragments often false high quantitative values are measured. Based on this methodology it is not possible to differentiate between high IGFBP-3 levels in fact, or, a high degree of fragmentation. The incidental attempts to use monoclonal antibodies with a binding region represented only by the intact IGFBP-3 molecule are indirect, imprecise and insufficient. The activities of all effective proteases, which have different sites of action and therefore generate different kind of fragments, are disregarded.