Laser-capture microdissection: applications in routine molecular dermatopathology.
Advances in molecular pathology with the introduction of the Southern blot technique and the polymerase chain reaction (PCR) have emerged as important tools, which are frequently used in routine dermato-histopathology. Applications for PCR-based diagnostics are particularly helpful for the determination of clonality in cutaneous lymphocytic infiltrates and for detection of infectious agents, such as herpes simplex virus (HSV), varicella zoster virus (VZV), Borrelia burgdorferi, Mycobacteria, Leishmania, and Treponema pallidum. As biopsies are always composed of different cells, the cells of interest are often only a minor population. As a consequence, their specific DNA is diluted by the majority of contaminating cells. Another problem is the time- and labor-intensive DNA extraction, because usually only formalin-fixed, paraffin-embedded tissue is available, which makes molecular diagnostics a time and labor consuming, and consequently a cost-intensive procedure. To overcome these shortcomings and to eventually shorten the time to generate a result, we introduce a laser-capture microdissection (LCM)-based method for the detection of infectious agents and clonality. Only the cells of interest for the particular indication are microdissected (e.g. epidermal cells for HSV and VZV and lymphocytes for clonality analysis) and subjected to PCR amplification. Due to an accelerated DNA-extraction procedure which generates DNA in 5 h (compared to 3-4 days using conventional DNA extraction), we are able to generate a result within one working day.
Yazdi
Amir S
AS
Department of Dermatology, Ludwig- Maximilians-University, Munich, Germany.
Puchta
Ursula
U
Flaig
Michael J
MJ
Sander
Christian A
CA
D000818:Animals; D015139:Blotting, Southern / Q000379:methods; D002999:Clone Cells; D004247:DNA / Q000302:isolation & purification; D003880:Dermatology / Q000379:methods; D006801:Humans; D007239:Infections / Q000382:microbiology*; D007834:Lasers; D042282:Microdissection / Q000379:methods; D010338:Pathology, Clinical; D016133:Polymerase Chain Reaction / Q000379:methods; D012867:Skin / Q000382:microbiology
D004247:DNA