sds-page-based quantitative assay for screening of kidney stone diseasesds-page-based定量测定肾结石疾病的筛查.pdfVIP

SDSBased Quantitative Assay for Screening of Kidney Stone Disease Lau Wai-Hoe, Leong Wing-Seng, Zhari Ismail, and Gam Lay-Harn Abstract Kidney stone disease is a common health problem in industrialised nations. We developed a SDS based method to quantify Tamm Horsfall glycoprotein (THP) for screening of kidney stone disease. Uri- nary proteins were extracted by using ammonium sulphate precipitation at 0.27 g salt/mL urine. The resulted pellet was dissolved in TSE buffer. Ten microliters of the urinary proteins extract was loaded and separated on 10% SDSunder reducing condition. THP migrated as single band in SDS- PAGE. The assay reproducibility and repeatability were 4.8% CV and 2.6% CV, respectively. A total of 117 healthy subjects and 58 stone patients were tested using this assay, and a distinct cut-off (PG 0.05) at 5.6 μg/mL THP concentration was used to distinguish stone patients from healthy subjects. The sen- sitivity and specificity of the method were 92.3% and 83.3%, respectively. Keywords: Kidney stone disease, SDS, Tamm Horsfall Protein and diagnostic device. 1. Introduction Pathogenesis of kidney stone disease involves nucleation, growth, aggregation, and retention of crystals in the kidney. Biochemical tests for urinary electrolytes cannot specifically identify the forma- tion of stone in the kidney. In human, the mechanism against the formation of kidney stone involves macromolecules, i.e., protein and glycosaminoglycan that are polyanionic molecules with sub- stantial amounts of acidic amino acid residues that inhibit crystal aggregation (1). Tamm Horsfall glycoprotein (THP) is one of the inhibitors to crystalize aggregation in kidney (2, 3). Shulin Li (ed