Microdissection Overview 显微解剖
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Microdissection
Overview
The Goals of Microdissection
Human tissues are comprised of multiple interacting cell populations in a complex three dimensional arrangement with each cellular phenotype determined by a unique profile of mRNA and protein expression. Before microdissection techniques were developed, the only analysis tools for phenotypic studies were primarily immunohistochemistry and in-situ hybridization. While useful, these tools are limited to single gene analysis and, in general, do not allow qualitative studies (mutation, deletion analysis) of DNA, mRNA, or proteins to be performed.
One goal of the NCI Prostate Group has been to develop methods and approaches that permit the molecular analysis of histopathologically defined cell types. They have pioneered the development of microdissection technology which, in concert with new molecular analytical methods, permits the study of disease processes as they exist in vivo (see diagram below), in a highly sensitive and specific way. This is proving to be particularly important in two scenarios:
- Pathological processes where the diseased cells comprise only a small percentage of the total cells present in a sample.
- Tumor progression where the disease evolves through stages represented by discrete microscopic foci.
The Development of Microdissection
The concept of tissue microdissection is quite simple, that is, procuring a specific population of cells from a heterogeneous tissue sample under direct microscopic visualization. However, in practice, the approach is technically challenging and has gone through several developmental stages:
- Early efforts entailed grossly removing tissue from a frozen histological slide or utilizing a scalpel blade to scrape sections and procure cells of interest.
- Subsequently, the use of a manual or micromanipulator-guided needle with an adhesive tip was developed, which improved the accuracy and reliability of microdissection.
- A technical advance known as selective ultraviolet radiation fractionation (SURF) was developed using ink to protect against ultraviolet irradiation and obliterating the genetic material of the unwanted surrounding tissue.
- A similar method was developed for isolating single cells, which utilizes a high-energy ultraviolet laser microbeam to obliterate the unwanted cells and the desired cells are then retrieved by a needle.
- Another laser-based method called Non-Contact Laser Microdissection of Membrane-Mounted Native Tissue was developed, which involves placing a tissue section onto a membrane on a glass slide and using a high-intensity ultraviolet beam to cut out an area of tissue/membrane that is subsequently recovered.
- Laser Capture Microdissection (LCM) is a positive selection method that was developed at the National Institutes of Health and designed to permit simple, fast, and reliable microdissection of tissue.
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