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Laser Microdissection of FFPE Tissue Areas and Subsequent Whole Genome Amplification by Ampli1™

  • Zbigniew Tadeusz Czyz
  • Nikolas H. Stoecklein
  • Bernhard PolzerEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1347)

Abstract

Laser microdissection (LMD) and whole genome amplification (WGA) are valuable tools to isolate, purify, and genetically analyze cancer cells from tissue sections. In this chapter, we describe a workflow for microdissecting small regions of interest from cancer tissue, i.e. formalin-fixed paraffin-embedded (FFPE) and cryo-conserved specimens, and subsequent whole genome amplification by a deterministic WGA approach (Ampli1™ WGA).

Key words

Laser microdissection (LMD) Whole genome amplification (WGA) Formalin-fixed paraffin-embedded (FFPE) tissue 

References

  1. 1.
    Klein CA, Blankenstein TJ, Schmidt-Kittler O, Petronio M, Polzer B, Stoecklein NH, Riethmuller G (2002) Genetic heterogeneity of single disseminated tumour cells in minimal residual cancer. Lancet 360(9334):683–689. doi: 10.1016/S0140-6736(02)09838-0 CrossRefPubMedGoogle Scholar
  2. 2.
    Klein CA, Schmidt-Kittler O, Schardt JA, Pantel K, Speicher MR, Riethmuller G (1999) Comparative genomic hybridization, loss of heterozygosity, and DNA sequence analysis of single cells. Proc Natl Acad Sci U S A 96(8):4494–4499PubMedCentralCrossRefPubMedGoogle Scholar
  3. 3.
    Arneson N, Moreno J, Iakovlev V, Ghazani A, Warren K, McCready D, Jurisica I, Done SJ (2012) Comparison of whole genome amplification methods for analysis of DNA extracted from microdissected early breast lesions in formalin-fixed paraffin-embedded tissue. ISRN Oncol 2012:710692. doi: 10.5402/2012/710692 PubMedCentralPubMedGoogle Scholar
  4. 4.
    Czyz ZT, Hoffmann M, Schlimok G, Polzer B, Klein CA (2014) Reliable single cell array CGH for clinical samples. PLoS One 9(1):e85907. doi: 10.1371/journal.pone.0085907 PubMedCentralCrossRefPubMedGoogle Scholar
  5. 5.
    Stoecklein NH, Erbersdobler A, Schmidt-Kittler O, Diebold J, Schardt JA, Izbicki JR, Klein CA (2002) SCOMP is superior to degenerated oligonucleotide primed-polymerase chain reaction for global amplification of minute amounts of DNA from microdissected archival tissue samples. Am J Pathol 161(1):43–51PubMedCentralCrossRefPubMedGoogle Scholar
  6. 6.
    Binder V, Bartenhagen C, Okpanyi V, Gombert M, Moehlendick B, Behrens B, Klein HU, Rieder H, Ida Krell PF, Dugas M, Stoecklein NH, Borkhardt A (2014) A new workflow for whole-genome sequencing of single human cells. Hum Mutat 35(10):1260–1270. doi: 10.1002/humu.22625 CrossRefPubMedGoogle Scholar
  7. 7.
    Fuhrmann C, Schmidt-Kittler O, Stoecklein NH, Petat-Dutter K, Vay C, Bockler K, Reinhardt R, Ragg T, Klein CA (2008) High-resolution array comparative genomic hybridization of single micrometastatic tumor cells. Nucleic Acids Res 36(7):e39. doi: 10.1093/nar/gkn101 PubMedCentralCrossRefPubMedGoogle Scholar
  8. 8.
    Guzvic M, Braun B, Ganzer R, Burger M, Nerlich M, Winkler S, Werner-Klein M, Czyz ZT, Polzer B, Klein CA (2014) Combined genome and transcriptome analysis of single disseminated cancer cells from bone marrow of prostate cancer patients reveals unexpected transcriptomes. Cancer Res 74(24):7383–7394. doi: 10.1158/0008-5472.CAN-14-0934 CrossRefPubMedGoogle Scholar
  9. 9.
    Husemann Y, Geigl JB, Schubert F, Musiani P, Meyer M, Burghart E, Forni G, Eils R, Fehm T, Riethmuller G, Klein CA (2008) Systemic spread is an early step in breast cancer. Cancer Cell 13(1):58–68CrossRefPubMedGoogle Scholar
  10. 10.
    Mohlendick B, Bartenhagen C, Behrens B, Honisch E, Raba K, Knoefel WT, Stoecklein NH (2013) A robust method to analyze copy number alterations of less than 100 kb in single cells using oligonucleotide array CGH. PLoS One 8(6):e67031. doi: 10.1371/journal.pone.0067031 PubMedCentralCrossRefPubMedGoogle Scholar
  11. 11.
    Neves RP, Raba K, Schmidt O, Honisch E, Meier-Stiegen F, Behrens B, Mohlendick B, Fehm T, Neubauer H, Klein CA, Polzer B, Sproll C, Fischer JC, Niederacher D, Stoecklein NH (2014) Genomic high-resolution profiling of single CKpos/CD45neg flow-sorting purified circulating tumor cells from patients with metastatic breast cancer. Clin Chem 60(10):1290–1297. doi: 10.1373/clinchem.2014.222331 PubMedGoogle Scholar
  12. 12.
    Polzer B, Medoro G, Pasch S, Fontana F, Zorzino L, Pestka A, Andergassen U, Meier-Stiegen F, Czyz ZT, Alberter B, Treitschke S, Schamberger T, Sergio M, Bregola G, Doffini A, Gianni S, Calanca A, Signorini G, Bolognesi C, Hartmann A, Fasching PA, Sandri MT, Rack B, Fehm T, Giorgini G, Manaresi N, Klein CA (2014) Molecular profiling of single circulating tumor cells with diagnostic intention. EMBO Mol Med 6(11):1371–1386. doi: 10.15252/emmm.201404033 PubMedCentralCrossRefPubMedGoogle Scholar
  13. 13.
    Schardt JA, Meyer M, Hartmann CH, Schubert F, Schmidt-Kittler O, Fuhrmann C, Polzer B, Petronio M, Eils R, Klein CA (2005) Genomic analysis of single cytokeratin-positive cells from bone marrow reveals early mutational events in breast cancer. Cancer Cell 8(3):227–239. doi: 10.1016/j.ccr.2005.08.003 CrossRefPubMedGoogle Scholar
  14. 14.
    Schmidt-Kittler O, Ragg T, Daskalakis A, Granzow M, Ahr A, Blankenstein TJ, Kaufmann M, Diebold J, Arnholdt H, Muller P, Bischoff J, Harich D, Schlimok G, Riethmuller G, Eils R, Klein CA (2003) From latent disseminated cells to overt metastasis: genetic analysis of systemic breast cancer progression. Proc Natl Acad Sci U S A 100(13):7737–7742. doi: 10.1073/pnas.1331931100 PubMedCentralCrossRefPubMedGoogle Scholar
  15. 15.
    Stoecklein NH, Hosch SB, Bezler M, Stern F, Hartmann CH, Vay C, Siegmund A, Scheunemann P, Schurr P, Knoefel WT, Verde PE, Reichelt U, Erbersdobler A, Grau R, Ullrich A, Izbicki JR, Klein CA (2008) Direct genetic analysis of single disseminated cancer cells for prediction of outcome and therapy selection in esophageal cancer. Cancer Cell 13(5):441–453. doi: 10.1016/j.ccr.2008.04.005 CrossRefPubMedGoogle Scholar
  16. 16.
    Ulmer A, Dietz K, Hodak I, Polzer B, Scheitler S, Yildiz M, Czyz Z, Lehnert P, Fehm T, Hafner C, Schanz S, Rocken M, Garbe C, Breuninger H, Fierlbeck G, Klein CA (2014) Quantitative measurement of melanoma spread in sentinel lymph nodes and survival. PLoS Med 11(2):e1001604. doi: 10.1371/journal.pmed.1001604 PubMedCentralCrossRefPubMedGoogle Scholar
  17. 17.
    Ulmer A, Schmidt-Kittler O, Fischer J, Ellwanger U, Rassner G, Riethmuller G, Fierlbeck G, Klein CA (2004) Immunomagnetic enrichment, genomic characterization, and prognostic impact of circulating melanoma cells. Clin Cancer Res 10(2):531–537CrossRefPubMedGoogle Scholar
  18. 18.
    Weckermann D, Polzer B, Ragg T, Blana A, Schlimok G, Arnholdt H, Bertz S, Harzmann R, Klein CA (2009) Perioperative activation of disseminated tumor cells in bone marrow of patients with prostate cancer. J Clin Oncol 27(10):1549–1556. doi: 10.1200/JCO.2008.17.0563 CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Zbigniew Tadeusz Czyz
    • 2
  • Nikolas H. Stoecklein
    • 1
  • Bernhard Polzer
    • 2
    Email author
  1. 1.Department of Visceral, General, and Pediatric SurgeryUniversity Hospital and Medical Faculty of the Heinrich-Heine University DüsseldorfDüsseldorfGermany
  2. 2.Project Group, Personalized Tumor TherapyFraunhofer Institute for Toxicology and Experimental Medicine (ITEM)RegensburgGermany

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