July - September 2007, Volume 3, Issue 3

Tanasescu R, Purice M

The influence of protein concentration on hGH immunoassay in different reaction enviroments

Acta Endo (Buc) 2007, 3 (3): 257-267
doi: 10.4183/aeb.2007.257

As the kits used in routine for assessment of human growth hormone (hGH) status in CSF are originally designed for the serum and plasma, the specifications and limitations imposed by some environmental factor are important to be known. The aim of study was to\r\ninvestigate the influence of different protein concentrations on an immunometric assay to design a suitable and reproducible method for hGH measurement in different matrices, particularly in CSF-like environment.\r\nMaterial and methods. Experiments were performed using an in-house protocol for the determination of hGH by Time Resolved Immunofluorometric method. Standard curves were prepared in different biological matrices (sheep, human or fetal calf serums and CSFlike matrix). The concentrations of albumin and gamma-globulins were the variable parameters. The gold standard was the standard curve in sheep serum.\r\nResults. Constantly high background signal values were obtained at different albumin concentrations, in the absence of IgG, indicating intense non-specific binding and a low sensitivity. This fact allows accurate hGH measurement at over 5-10 ng/ml. Using a reaction environment enriched in IgG with a standard albumin concentration, background signal decreases while increasing IgG concentration. The highest sensibility is obtained for 5 g/l and 2.5 g/l IgG environments, allowing the sample signal to be measured accurately at hGH concentrations of 0.2 ng/ml or 0.5 ng/ml respectivly. This is observed in all biological matrices used.\r\nConclusions. The variation of protein concentration influences hGH determination. A standard curve with high sensibility for low values and a reduction in the influences of the reaction environment by minimising non-specific binding were obtained by enriching the environment with IgG.

Keywords: hGH, immunometric methods, IFMA, albumin, IgG, cerebrospinal fluid

Correspondence: Ruxandra Tanasescu, medical student, grant holder of “Grigore Coculescu” grant for Neuroendocrinology, Department of Endocrinology, "Carol Davila" University of Medicine and Pharmacy, 36 Bdul Aviatorilor, 011863 Bucharest, Romania, e-mail:ruxandra.tanasescu@gmail.com