Abstract
Recent developments in sensitive genome characterization and quantitative gene expression analyses that permit precise molecular genetic fingerprinting of tumoral tissue are having a huge impact on cancer diagnostics. However, the significance of the data obtained with these techniques strictly depends on the composition of the biological sample to be analyzed and is greatly enhanced by including a preprocessing step that allows the researcher to distinguish and isolate selected cell populations from surrounding undesired material. This may represent a remarkable problem: indeed, genomic and proteomic analysis in the context of cancer investigation is susceptible to contamination by nonneoplastic cells, which can mask some tumor-specific alterations. Moreover, the heterogeneity of the tissues of a histo-logical section, in which the cell population of interest may constitute only a small fraction, can represent an insurmountable difficulty for the use of quantitative techniques that absolutely depend on genomic material stricdy derived from the cells that require analysis. This is obviously not possible if DNA or RNA is extracted from entire biopsies.
In the past, this obstacle was partially overcome by manual dissection from slides with a needle or scalpel; however, this method is feasible only if there is a clear demarcation between the tissue under consideration and its surroundings and moreover, allows only an approximate separation of tissues. The recent development of microdissection systems based on laser technology has largely solved this important problem.
Laser microdissection is a powerful tool for the isolation of specific cell populations (or single cells) from stained sections of both formalin-fixed, paraffin-embedded and frozen tissues, from cell cultures and even of a single chromosome within a metaphase cell. Resulting material is suitable for a wide range of downstream assays such LOH (loss of heterozygosity) studies, gene expression analysis at the mRNA level and a variety of proteomic approaches such as 2D gel analysis, reverse phase protein array and SELDI protein profiling. This chapter describes the characteristics of the most widely utilized laser microdissection systems and their current applications.
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References
Abeln EC, Smit VT, Wessels JW et al. Molecular genetic evidence for the conversion hypothesis of the origin of malignant mixed mullerian tumours. J Pathol 1997;183:424–31.
Perren A, Roth J, Muletta-Feurer S et al. Clonal analysis of sporadic pancreatic endocrine tumours. J Pathol 1998;186:363–71.
Saxena A, Alport EC, Custead S et al. Molecular analysis of clonality of sporadic angiomyolipoma. J Pathol 1999;189:79–84.
Looijenga LH, Rosemberg C, van Gurp RH et al. Comparative genomic hybridization of microdissected samples from different stages in the development of a seminoma and a non-seminoma. J Pathol 2000;191:187–92.
Shibata D. Selective ultraviolet radiation fractionation and polymerase chain reaction analysis of genetic alterations. Am J Pathol 1993;143:1523–6.
Emmert-Buck MR, Bonner RF, Smith PD et al. Laser capture microdissection. Science 1996;274:998–1001.
Vogel A, Venugopalan V. Mechanisms of pulsed laser ablation of biological tissues. Chem Rev 2003;103:577–644.
Srinivasan R, Leigh WJ. Ablative photodecomposition: Action of far ultraviolet (193 nm) laser radiation on poly (ethyleneterephthalate) films. J Amer Chem Soc 1982;104:6784–5.
Trokel SL, Srinivasan R, Braren B. Excimer laser surgery of the cornea. Am J Ophthalmol 1983;96:710–5.
Di Cristofano C, Mrad K, Zavaglia K et al. Papillary lesions of the breast: A molecular progression? Breast Cancer Res Treat 2005;90:71–6.
D’Arrigo A, Belluco C, Ambrosi A et al. Metastatic transcriptional pattern revealed by gene expression profiling in primary colorectal carcinoma. Int J Cancer 2005;115:256–62.
Coco S, Defferrari R, Scaruffi P et al. Genome analysis and gene expression profiling of neuroblastoma and ganglioneuroblastoma reveal differences between neuroblastic and Schwannian stromal cells. J Pathol 2005;207:346–357.
Burgemeister R. New aspects of laser microdissection in research and routine. J Histochem Cytochem 2005;53:409–12.
Schneider-Stock R, Boltze C, Jaeger V et al. Significance of loss of heterozygosity of the RB1 gene during tumour progression in well-differentiated liposarcomas. J Pathol 2002;197:654–60.
Sethi N, Palefsky J. Transcriptional profiling of dysplastic lesions in K14-HPV16 transgenic mice using laser microdissection. FASEB J 2004;18:1243–5.
Cowherd SM, Espina VA, Petricoin IIIrd EF et al. Proteomic analysis of human breast cancer tissue with laser-capture microdissection and reverse-phase protein microarrays. Clin Breast Cancer 2004;5:385–92.
Sugiyama Y, Farrow B, Murillo C et al. Analysis of differential gene expression patterns in colon cancer and cancer stroma using microdissected tissues. Gastroenterology 2005;128:480–6.
Ma XJ, Salunga R, Tuggle JT et al. Gene expression profiles of human breast cancer progression. Proc Natl Acad Sci USA 2003;100:5974–9.
Ren ZP, Sallstrom J, Sundstrom C et al. Recovering DNA and optimizing PCR conditions from microdissected formalin-fixed and paraffin-embedded materials. Pathobiology 2000;68:215–7.
Gloghini A, Canal B, Klein U et al. RT-PCR analysis of RNA extracted from Bouin-fixed and paraffin-embedded lymphoid tissues. J Mol Diagn 2004;6:290–6.
Florell SR, Coffin CM, Holden JA et al. Preservation of RNA for functional genomic studies: A multidisciplinary tumor bank protocol. Mod Pathol 2001;14:116–28.
Roos-van Groningen MC, Eikmans M, Baelde HJ et al. Improvement of extraction and processing of RNA from renal biopsies. Kidney Int 2004;65:97–105.
Kunz Jr GM, Chan DW. The use of laser capture microscopy in proteomics research-a review. Dis Markers 2004;20:155–60.
Ehrig T, Abdulkadir SA, Dintzis SM et al. Quantitative amplification of genomic DNA from histological tissue sections after staining with nuclear dyes and laser capture microdissection. J Mol Diagn 2001;3:22–5.
Burgemeister R, Gangnus R, Haar B et al. High quality RNA retrieved from samples obtained by using LMPC (laser microdissection and pressure catapulting) technology. Pathol Res Pract 2003;199:431–6.
Okuducu AF, Janzen V, Hahne JC et al. Influence of histochemical stains on quantitative gene expression analysis after laser-assisted microdissection. Int J Mol Med 2003;11:449–53.
Craven RA, Totty N, Harnden P et al. Laser capture microdissection and two-dimensional poly-acrylamide gel electrophoresis: Evaluation of tissue preparation and sample limitations. Am J Pathol 2002;160:815–22.
Fend F, Emmert-Buck MR, Chuaqui R et al. Immuno-LCM: Laser capture microdissection of immunostained frozen sections for mRNA analysis. Am J Pathol 1999;154:61–6.
Fend F, Kremer M, Quintanilla-Martinez L. Laser capture microdissection: Methodical aspects and applications with emphasis on immuno-laser capture microdissection. Pathobiology 2000;68:209–14.
Fink L, Kinfe T, Seeger W et al. Immunostaining for cell picking and real-time mRNA quantitation. Am J Pathol 2000;157:1459–66.
Fink L, Kinfe T, Stein MM et al. Immunostaining and laser-assisted cell picking for mRNA analysis. Lab Invest 2000;80:327–33.
Burbach GJ, Dehn D, Nagel B et al. Laser microdissection of immunolabeled astrocytes allows quantification of astrocytic gene expression. J Neurosci Methods 2004;138:141–8.
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© 2007 Landes Bioscience and Springer Science+Business Media
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Esposito, G. (2007). Complementary Techniques. In: Mocellin, S. (eds) Microarray Technology and Cancer Gene Profiling. Advances in Experimental Medicine and Biology, vol 593. Springer, New York, NY. https://doi.org/10.1007/978-0-387-39978-2_6
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DOI: https://doi.org/10.1007/978-0-387-39978-2_6
Publisher Name: Springer, New York, NY
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