Microwave irradiation in label-detection for diagnostic DNA-in situ hybridization.
A DNA-in situ hybridization protocol was adapted for application to sections of routinely processed paraffin embedded material. This protocol was developed previously for detecting DNA-virus infected cells in whole cell preparations and employs biotinylated DNA as probe. Three different biotin detection methods were optimized and applied. The first uses streptavidin and a biotinylated complex of alkaline phosphatase, the second consists of an immunogold-silver staining, and the third of a peroxidase technique using a silver amplification. The alkaline phosphatase method was the most rapid, and as sensitive as the immunogold-silver staining. The peroxidase method was the most sensitive. Microwave irradiation was applied to the different incubation steps of these three detection methods. Short incubations with microwave irradiation gave results comparable to those obtained with conventional incubations, when streptavidin, antibiotin, complexed alkaline phosphatase, or gold labelled goat antirabbit were used. It was thus shown that microwave irradiation creates the possibility of a very rapid label-detection for nonradioactive DNA-in situ hybridization.
Department of Pathology, Academic Medical Center, Amsterdam, The Netherlands.
W J
WJ
Van den Brink
H J
HJ
Zijlmans
L P
LP
Kok
P
P
Bolhuis
H H
HH
Volkers
M E
ME
Boon
H J
HJ
Houthoff
D000469:Alkaline Phosphatase; D000818:Animals; D001426:Bacterial Proteins; D001710:Biotin / Q000032:analysis; D003218:Condylomata Acuminata / Q000473:pathology; D004247:DNA / Q000032:analysis* / Q000528:radiation effects; D019286:Endopeptidase K; D006041:Goats / Q000276:immunology; D006868:Hydrolysis; D007124:Immunoenzyme Techniques; D007150:Immunohistochemistry; D008872:Microwaves; D009693:Nucleic Acid Hybridization; D027383:Papillomaviridae / Q000648:ultrastructure; D010434:Pepsin A / Q000627:therapeutic use; D011817:Rabbits / Q000276:immunology; D012697:Serine Endopeptidases; D013194:Staining and Labeling; D019809:Streptavidin
D001426:Bacterial Proteins; D001710:Biotin; D004247:DNA; D019809:Streptavidin; D000469:Alkaline Phosphatase; D012697:Serine Endopeptidases; D019286:Endopeptidase K; D010434:Pepsin A