Assessment of the 2-d gel-based proteomics application of clinically archived formalin-fixed paraffin embedded tissues.
Hospital tissue repositories possess a vast and valuable supply of disease samples with matched retrospective clinical information. Detection and characterization of disease biomarkers in formalin-fixed paraffin-embedded (FFPE) tissues will greatly aid the understanding of the diseases mechanisms and help in the development of diagnostic and prognostic markers. In this study, the possibility of using full-length proteins extracted from clinically archived FFPE tissues in two-dimensional (2-D) gel-based proteomics was evaluated. The evaluation was done based on two types of tumor tissues (breast and prostate) and two extraction protocols. The comparison of the 2-D patterns of FFPE extracts obtained by two extraction protocols with the matching frozen tissue extracts showed that only 7-10% of proteins from frozen tissues can be matched to proteins from FFPE tissues. Most of the spots in the 2-D FFPE's maps had pl 4-6, while the percentages of proteins with pl above 6 were 3-5 times lower in comparison to the fresh/frozen tissue. Despite the three-fold lower number of the detected spots in FFPE maps compared to matched fresh/frozen maps, 67-78% of protein spots in FFPE could not be matched to the corresponding spots in the fresh/frozen tissue maps indicating irreversible protein modifications. In conclusion, the inability to completely reverse the cross-linked complexes and overcome protein fragmentation with the present day FFPE extraction methods stands in the way of effective use of these samples in 2-D gel based proteomics studies.
Research Centre for Genetic Engineering and Biotechnology "Georgi D Efremov", Macedonian Academy of Sciences and Arts, Krste Misirkov 2, 1000, Skopje, Republic of Macedonia, katarina@manu.edu.mk.
Katarina
K
Davalieva
Sanja
S
Kiprijanovska
Momir
M
Polenakovic
D014408:Biomarkers, Tumor / Q000032:analysis / Q000302:isolation & purification; D001940:Breast / Q000473:pathology; D001943:Breast Neoplasms / Q000473:pathology*; D015925:Cryopreservation; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D005260:Female; D005557:Formaldehyde / Q000737:chemistry; D006801:Humans; D008297:Male; D016612:Paraffin Embedding; D011467:Prostate / Q000473:pathology; D011471:Prostatic Neoplasms / Q000473:pathology*; D011506:Proteins / Q000032:analysis* / Q000302:isolation & purification; D040901:Proteomics / Q000379:methods*; D016707:Tissue Fixation
D014408:Biomarkers, Tumor; D011506:Proteins; D005557:Formaldehyde