Detergent addition to tryptic digests and ion mobility separation prior to MS/MS improves peptide yield and protein identification for in situ proteomic investigation of frozen and formalin-fixed paraffin-embedded adenocarcinoma tissue sections.
The identification of proteins involved in tumour progression or which permit enhanced or novel therapeutic targeting is essential for cancer research. Direct MALDI analysis of tissue sections is rapidly demonstrating its potential for protein imaging and profiling in the investigation of a range of disease states including cancer. MALDI-mass spectrometry imaging (MALDI-MSI) has been used here for direct visualisation and in situ characterisation of proteins in breast tumour tissue section samples. Frozen MCF7 breast tumour xenograft and human formalin-fixed paraffin-embedded breast cancer tissue sections were used. An improved protocol for on-tissue trypsin digestion is described incorporating the use of a detergent, which increases the yield of tryptic peptides for both fresh frozen and formalin-fixed paraffin-embedded tumour tissue sections. A novel approach combining MALDI-MSI and ion mobility separation MALDI-tandem mass spectrometry imaging for improving the detection of low-abundance proteins that are difficult to detect by direct MALDI-MSI analysis is described. In situ protein identification was carried out directly from the tissue section by MALDI-MSI. Numerous protein signals were detected and some proteins including histone H3, H4 and Grp75 that were abundant in the tumour region were identified.
Djidja
Marie-Claude
MC
Biomedical Research Centre, Sheffield Hallam University, Howard Street, Sheffield, UK.
Francese
Simona
S
Loadman
Paul M
PM
Sutton
Chris W
CW
Scriven
Peter
P
Claude
Emmanuelle
E
Snel
Marten F
MF
Franck
Julien
J
Salzet
Michel
M
Clench
Malcolm R
MR
D000230:Adenocarcinoma / Q000737:chemistry*; D000818:Animals; D001943:Breast Neoplasms / Q000737:chemistry; D045744:Cell Line, Tumor; D003902:Detergents; D004195:Disease Models, Animal; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D009363:Neoplasm Proteins / Q000032:analysis* / Q000737:chemistry; D009368:Neoplasm Transplantation; D016612:Paraffin Embedding; D040901:Proteomics / Q000379:methods*; D012680:Sensitivity and Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014357:Trypsin / Q000378:metabolism*
D003902:Detergents; D009363:Neoplasm Proteins; D014357:Trypsin