The potential use of laser capture microdissection to selectively obtain distinct populations of cells for proteomic analysis--preliminary findings.
Proteomics-based studies offer a powerful complementary approach to DNA/RNA-based investigations and are now being applied to investigate aspects of many diseases including cancer. However, the heterogeneous nature of tissue samples often makes interpretation difficult. We have undertaken a study into the potential use of a novel laser capture microdissection (LCM) system to isolate cells of interest for subsequent proteomic analysis. Retrieval of selected cells is achieved by activation of a transfer film placed in contact with a tissue section, by a laser beam (30 or 60 microm diameter) which is focused on a selected area of tissue using an inverted microscope. The precise area of film targeted by the laser bonds to the tissue beneath it and these cells are then lifted free of surrounding tissue. Although the technique has been shown to be readily compatible with subsequent analysis of nucleic acids, little information is yet available regarding the application of protein-based analyses to the captured tissue. We report here preliminary data regarding the potential use of the LCM system in combination with two-dimensional electrophoresis to examine protein profiles of selected tissue areas. Electrophoretic profiles of proteins from normal and malignant renal tissue samples showed little change following LCM, nine selected proteins showed identical mass spectrometric sequencing profiles, and two selected proteins retained antigenicity. Dissection of epithelial tissue from a sample of normal human cervix resulted in enrichment of some proteins compared with analysis of the whole tissue. LCM will be a valuable adjunct to proteomic studies although further detailed validation is necessary.
Banks
R E
RE
ICRF Cancer Medicine Research Unit, St James's University Hospital, Leeds, UK. r.banks@leeds.ac.uk
Dunn
M J
MJ
Forbes
M A
MA
Stanley
A
A
Pappin
D
D
Naven
T
T
Gough
M
M
Harnden
P
P
Selby
P J
PJ
D000595:Amino Acid Sequence; D002584:Cervix Uteri / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D005260:Female; D006801:Humans; D007668:Kidney / Q000737:chemistry; D007680:Kidney Neoplasms / Q000737:chemistry; D007834:Lasers; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000302:isolation & purification; D011506:Proteins / Q000302:isolation & purification
D009363:Neoplasm Proteins; D011506:Proteins