Proteomic analysis of formalin-fixed paraffin-embedded renal tissue samples by label-free MS: assessment of overall technical variability and the impact of block age.
Author(s): Craven RA, Cairns DA, Zougman A, Harnden P, Selby PJ, Banks RE
Publication: Proteomics Clin Appl, 2013, Vol. 7, Page 273-82
PubMed ID: 23027403 PubMed Review Paper? No
Purpose of Paper
The purpose of this paper was to assess effects of formalin-fixed paraffin-embedded (FFPE) block storage on the identity and levels of proteins detected in normal and cancerous kidney specimens.
Conclusion of Paper
The duration of FFPE block storage did not alter the number of proteins identified by LC-MS/MS, their identity or their abundance in normal kidney or renal cell carcinoma (RCC) specimens. However, fewer proteins and greater variability were present in RCC FFPE specimens than normal kidney FFPE specimens, which the other authors attribute to patient heterogeneity. A total of 2,663 proteins were collectively identified from all FFPE normal kidney tissue specimens. The number of proteins identified per specimen did not differ among FFPE blocks stored for different durations (range 1889-2153 unique proteins). There was substantial overlap in protein identity both within (65-68%) and between (83%) the shortest (<18 months) and longest (10 y) FFPE storage durations. Protein intensities were very strongly correlated between the shortest and longest FFPE block storage durations (r= 0.954- 0.969). Comparatively, results obtained with FFPE RCC specimens yielded fewer proteins per specimen (1335-1945), and a smaller degree of overlap in protein identity within (47%) storage duration timepoints as well as between (47%) the shortest and longest timepoints in comparison to normal FFPE specimens. Protein intensities in RCC FFPE specimens displayed were strongly correlated between the shortest and longest FFPE block storage durations (r= 0.804- 0.961).
Studies
-
Study Purpose
The purpose of this study was to assess effects of formalin-fixed paraffin-embedded (FFPE) block storage on the identity and levels of proteins detected in normal and cancerous kidney tissue specimens. Kidney specimens from 16 normal individuals and 8 patients diagnosed with renal cell carcinoma (RCC), were collected, processed and subjected to FFPE block storage for <18 months, 5 y, 7 y, or 10 y. Specimens assigned to each storage timepoint were matched for age and gender. The conditions of FFPE block storage were not provided. Protein lysates extracted from 10 µm thick FFPE sections were stored at -80°C then digested with trypsin prior to analysis by LC-MS/MS.
Summary of Findings:
A total of 2,663 proteins were collectively identified from all FFPE normal kidney tissue specimens. The number of proteins identified per sample did not differ among FFPE blocks stored for different durations (range 1889-2153 unique proteins). There was a good overlap in protein identity both within (65-68%) and between (83%) the shortest (<18 months) and longest (10 y) FFPE storage durations. Protein intensities were very strongly correlated between specimens stored for the shortest and longest FFPE block durations (r= 0.954- 0.969). Comparatively, results obtained with FFPE samples obtained from RCC kidney tissue, yielded fewer proteins per specimen (1335-1945), and a smaller degree of overlap in protein identity within (47%) storage duration timepoints as well as between (47%) the shortest and longest timepoints in comparison to normal FFPE specimens. Protein intensities in RCC FFPE specimens were strongly or very strongly correlated between the shortest and longest FFPE block storage durations (r= 0.804- 0.961).
Biospecimens
Preservative Types
- Formalin
Diagnoses:
- Normal
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform Protein LC-MS or LC-MS/MS Peptide LC-MS or LC-MS/MS Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Preaquisition Diagnosis/ patient condition Normal
Renal cell carcinoma
Storage Storage duration < 18 months
5 y
7 y
10 y