title: Comprehensive proteome analysis of fresh frozen and optimal cutting temperature (OCT) embedded primary non-small cell lung carcinoma by LC-MS/MS.
abstract: Clinical tissue samples provide valuable information for understanding human diseases. One major type of clinical tissue sample that is amenable to various kinds of analysis is fresh frozen and optimal cutting temperature (OCT)-embedded primary patient tissue. Recent advances in mass spectrometry (MS) technologies have been widely applied to study human proteomes by using clinical specimens. However, polymeric compounds such as OCT can interfere with MS analyses. Here we present methods that enable the preparation and analysis of fresh frozen and OCT embedded primary tissue samples by LC-MS/MS. A scraping method was first introduced to reduce the heterogeneity of OCT-embedded non-small cell lung carcinoma tumor sections. OCT compound was reproducibly removed by a series of washing steps involving ethanol and water prior to trypsin digestion. In data-dependent acquisition mode, optimized dynamic exclusion duration settings were established to maximize peptide identifications. These sample preparation conditions and MS parameter settings should be utilized or carefully adjusted in order to achieve optimal comprehensive proteome characterization starting from fresh frozen and OCT embedded clinical tissue specimens.
authors.affiliation: Program in Molecular Structure and Function, The Hospital For Sick Children, 686 Bay Street, Toronto, Ontario M5G 0A4, Canada; Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Wen
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Princess Margaret Cancer Centre, Toronto, Ontario, Canada.
authors.forename: Shingo
authors.initials: S
authors.lastname: Sakashita
authors.affiliation: Program in Molecular Structure and Function, The Hospital For Sick Children, 686 Bay Street, Toronto, Ontario M5G 0A4, Canada.
authors.forename: Paul
authors.initials: P
authors.lastname: Taylor
authors.affiliation: Princess Margaret Cancer Centre, Toronto, Ontario, Canada; Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Ming S
authors.initials: MS
authors.lastname: Tsao
authors.affiliation: Program in Molecular Structure and Function, The Hospital For Sick Children, 686 Bay Street, Toronto, Ontario M5G 0A4, Canada; Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada; Princess Margaret Cancer Centre, Toronto, Ontario, Canada. Electronic address: m.moran@utoronto.ca.
authors.forename: Michael F
authors.initials: MF
authors.lastname: Moran
mesh_terms: D002289:Carcinoma, Non-Small-Cell Lung / Q000737:chemistry*; D002853:Chromatography, Liquid / Q000379:methods*; D015925:Cryopreservation / Q000379:methods; D005615:Freezing; D006801:Humans; D008175:Lung Neoplasms / Q000737:chemistry*; D020543:Proteome / Q000032:analysis*; D013048:Specimen Handling / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D020543:Proteome
keywords: Lung primary tumors; Mass spectrometry; OCT compound; Proteomics