A. Barbati, L. Cesarini, R. Pellegrino, G.C. Di Renzo, Chia Grenelli
Objective: The measurement of urinary Cystatin C (cyst-C) can be a very good way for early identifying newborns predisposed to renal function impairment. PETIA, PENIA and EIA are the immunometric methods used for quantitative determination of cyst-C in human biologic fluid but they have some interference and do not allow realizing qualitative analysis. The present study performs a validation of Immunoblot SDS-PAGE for the analysis of urinary cyst-C.
Methods: urines were collected from neonates of the nursey at S. Maria della Misericordia Hospital. Urinary cyst-C was investigated by the SDS-PAGE electrophoresis, the immunoblot and the reading of optical density.
Results: the qualitative analysis showed two different molecular forms: a reactivity at about 70 KDa in all samples and a reactivity at 13 KDa in a limited number of samples. The coefficient of variation for assay precision was 10% and for assay accuracy was ± 10%; the detection limit was 0.009 ng/ μL and the calibration curve has a good linearity (range 0.02-0.3 ng/ μL). The stability of urinary cyst-C was acceptable, even without the use of protease inhibitors, if the assay was performed on urine freshly recruited or immediately after thawing the samples, stored for up to six months.
Conclusion: Immunoblot SDS-PAGE analysis is a valid method to obtain a qualitative and a quantitative analysis of urinary cyst-C.