Advances in isolation and characterization of homogeneous cell populations using laser microdissection.
The isolation and characterization of homogeneous cell populations are of great importance for the analysis of gene expression, because normal tissues contain various types of cells, and the differences in the populations of isolated cells exert significant effects on gene expression analysis. Researchers have attempted to develop methods for the isolation of homogeneous cell populations, such as flow cytometry and mechanical dissection. However, the recent emergence of laser-assisted microdissection has revolutionized the isolation of single-cell populations from solid tissues. With the help of a cutting laser, laser microdissection can isolate tissues (cells) of interest without contamination from surrounding tissues with the microscopic visualization field. By combining laser microdissection and subsequent microarray technology, several studies have resulted in the identification of disease-related genes. In this review, we summarize the principle of laser microdissection and provide several successful examples of target-gene identification using the conventional method combining laser microdissection and microarray. Next, we discuss the practical drawbacks of the combinational method, such as the need for a large number of cells and the disturbance of the relative abundance of transcripts during RNA amplification. We introduce our modifications to combined laser microdissection and microarray for detection of disease-related genes; the technique is simple, yet practical and accurate. Finally, versatile applications of laser microdissection, not only to transcript expression analysis, but also to other genomics and proteomics analyses are, also presented.
Mizuarai
S
S
Banyu Tsukuba Research Institute, Tsukuba, Ibaraki 300-2611, Japan.
Takahashi
K
K
Kobayashi
T
T
Kotani
H
H
D000818:Animals; D006801:Humans; D007834:Lasers; D042282:Microdissection; D020411:Oligonucleotide Array Sequence Analysis; D040901:Proteomics