Application of Label-Free Proteomics for Quantitative Analysis of Urothelial Carcinoma and Cystitis Tissue

Methods Mol Biol. 2021;2228:283-292. doi: 10.1007/978-1-0716-1024-4_20.

Abstract

A label-free approach based on a highly reproducible and stable workflow allows for quantitative proteome analysis . Due to advantages compared to labeling methods, the label-free approach has the potential to measure unlimited samples from clinical specimen monitoring and comparing thousands of proteins. The presented label-free workflow includes a new sample preparation technique depending on automatic annotation and tissue isolation via FTIR-guided laser microdissection, in-solution digestion, LC-MS/MS analyses, data evaluation by means of Proteome Discoverer and Progenesis software, and verification of differential proteins. We successfully applied this workflow in a proteomics study analyzing human cystitis and high-grade urothelial carcinoma tissue regarding the identification of a diagnostic tissue biomarker. The differential analysis of only 1 mm2 of isolated tissue cells led to 74 significantly differentially abundant proteins.

Keywords: AHNAK2; Bladder cancer; FTIR imaging; Label-free proteomics; Laser microdissection; Urothelial cell carcinoma (UCC).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatography, High Pressure Liquid
  • Cystitis / metabolism*
  • Humans
  • Laser Capture Microdissection
  • Neoplasm Proteins / analysis*
  • Proteome*
  • Proteomics*
  • Research Design
  • Spectrometry, Mass, Electrospray Ionization*
  • Spectroscopy, Fourier Transform Infrared
  • Tandem Mass Spectrometry*
  • Urinary Bladder Neoplasms / metabolism*
  • Urothelium / metabolism*

Substances

  • Neoplasm Proteins
  • Proteome