Table 2.

Selected strategies for urine protein collection and analysis

StudyNMP-22 immunoassay (7)Ovarian cancer biomarker discovery (62)Urine protein cataloguing (2)HUPO Kidney and Urine Proteome Project (50)
CollectionSpot urineSpot urineSingle/pooled samplesMidstream 2nd or spot urine
Additive(s)Protease inhibitorNoneProtease inhibitorSodium azide/boric acid
ProcessingCentrifugeStored at −80°C within 8 h Centrifuge after thawingCentrifuge 2,000 × g (10 min)Centrifuge 10,000 × g (10 min)
Protein cleanupBuffer exchangeProtein precipitationUltrafiltrationNA
SeparationNATwo-dimensional GelRPLC, SDS-PAGENA
DetectionImmunoassayMALDI-TOF-MS LC-MS/MS IP and ELISALC-MS/MSNA
ResultDetection of bladder cancer 81% sensitivity, 87% specificityTargets identified by MS and verified with ELISA1,500 proteins identified in urine

NOTE: Workflows are presented for antibody-based quantification techniques as well as proteome cataloging experiments. The recommendations of the HUPO Human Kidney and Urine Proteome Project are also included.

Abbreviations: NA, not applicable; RPLC, reverse-phase liquid chromatography; MALDI-TOF-MS, matrix assisted laser desorption ionization time-of-flight mass spectrometry; IP, immunoprecipitation.