Analysis of antigen receptor gene rearrangements in ethanol and formaldehyde-fixed, paraffin-embedded specimens.
Molecular hybridization analysis of DNA prepared from frozen specimens obtained from patients with lymphoproliferative disorders has aided in the determination of the lineage and clonality of the neoplastic cells in many cases. We investigated whether high molecular weight DNA suitable for nucleic acid hybridization studies could also be prepared from fixed, paraffin-embedded material. After selecting nine representative cases, we extracted DNA from frozen sections by using standard methods, and from ethanol-fixed tissue in paraffin blocks. The yields of DNA from ethanol-fixed blocks were similar to yields from frozen tissue. DNA from frozen and ethanol-fixed tissues was subjected to Southern blot hybridization and probed for rearrangements of immunoglobulin heavy, and kappa, and lambda light chain genes, as well as for the T cell receptor beta-chain gene. In each of the cases, comparable results were obtained, regardless of the source of DNA. DNA extracted from ethanol-fixed blocks stored for 2 years gave identical results. We also prepared DNA from formaldehyde-fixed, paraffin-embedded tissue obtained from six of the patients. Five of the specimens yielded spoolable DNA (average recovered, 313 micrograms), but the DNA was degraded more than that obtained from the frozen or ethanol-fixed specimens. Formaldehyde-treated DNA gave variable results in Southern blot hybridization studies, and less than half of the results were interpretable. We conclude that ethanol-fixed, paraffin-embedded tissues provide an excellent source of DNA for nucleic-acid hybridization studies, and that they are easily handled and stored.
Wu
A M
AM
Department of Molecular Biochemistry, Beckman Research Institute of the City of Hope, Duarte, California.
Ben-Ezra
J
J
Winberg
C
C
Colombero
A M
AM
Rappaport
H
H
D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification; D015329:Gene Rearrangement, T-Lymphocyte; D005803:Genes, Immunoglobulin; D006801:Humans; D008198:Lymph Nodes / Q000276:immunology* / Q000473:pathology; D008223:Lymphoma / Q000235:genetics / Q000276:immunology* / Q000473:pathology; D008232:Lymphoproliferative Disorders / Q000235:genetics / Q000276:immunology* / Q000473:pathology; D009693:Nucleic Acid Hybridization; D015183:Restriction Mapping
D004273:DNA, Neoplasm; D004247:DNA