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SDS-PAGE-Based Quantitative Assay for Screening of Kidney Stone Disease

External protocol Created on 09 Apr 2014

Authors

Lau Wai-Hoe, Leong Wing-Seng, Zhari Ismail, Gam Lay-Harn

Summary

Kidney stone disease is a common health problem in industrialised nations. We developed a SDS-PAGE-based method to quantify Tamm Horsfall glycoprotein (THP) for screening of kidney stone disease. Urinary 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% SDS-PAGE under 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 (P < 0.05) at 5.6 μg/mL THP concentration was used to distinguish stone patients from healthy subjects. The sensitivity and specificity of the method were 92.3% and 83.3%, respectively.

Further details

The protocol was imported from Springer Protocols. To see the entire protocol, click on the source link, or download the pdf here.

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