Laser-assisted microdissection of membrane-mounted paraffin sections for polymerase chain reaction analysis: identification of cell populations using immunohistochemistry and in situ hybridization.
J Mol Diagn
; 3(3): 105-10, 2001 Aug.
Article
en En
| MEDLINE
| ID: mdl-11486049
Laser microbeam microdissection (LMM) is an increasingly important method for obtaining pure cell samples for genetic and proteomic analysis. Immunohistochemistry (IHC) and in situ hybridization (ISH) are useful techniques for targeting specific cell populations for microdissection but are difficult to apply with the tissue support membranes often used during LMM. Using detection of cytokeratins and Epstein-Barr virus gene products in head and neck carcinoma as a model, we describe optimized protocols for membrane and section preparation and for low temperature antigen retrieval that allow IHC and ISH to be used reliably on membrane mounted paraffin tissue sections. Visualization of cellular targets was markedly improved by staining and this could be further improved using a variety of optical media before microdissection. Tissue fragments thus stained were suitable for subsequent polymerase chain reaction analysis of extracted DNA using standard techniques. These IHC and ISH procedures are generally applicable and will be useful for detecting a wide range of antigens and nucleic acids in paraffin sections in conjunction with LMM.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Inmunohistoquímica
/
Reacción en Cadena de la Polimerasa
/
Hibridación in Situ
/
Rayos Láser
Tipo de estudio:
Diagnostic_studies
Límite:
Humans
Idioma:
En
Revista:
J Mol Diagn
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2001
Tipo del documento:
Article
País de afiliación:
Dinamarca
Pais de publicación:
Estados Unidos