Unit Code 87862:
Neuroblastoma, N-myc Amplification, FISH
Method Description
Formalin-fixed, paraffin-embedded tissues are cut at 5 m and
mounted on positively-charged glass slides. Four slides are
prepared, with 1 slide stained with hematoxylin-and-eosin (H&E).
The selection of tissue and the identification of target areas on an
H&E stained slide is performed by a pathologist. Using the H&E
as a reference, target areas are etched with a diamond tipped
etcher on the back of the unstained slide to be assayed. Anomalies
involving the N-myc oncogene region at 2p24.1 are detected using
an enumeration N-MYC FISH probe (Vysis, Inc., Downers Grove, IL).
The enumeration probe set consists of a chromosome 2 centromere
(D2Z1) probe labeled in SpectrumOrange (Vysis, Inc., Downers Grove,
IL) (appears red) and an N-MYC probe labeled in SpectrumGreen.
The probe set is applied to the appropriate target areas, denatured,
and hybridized overnight. Two scorers analyze 30 interphase nuclei
each (60 total) and a ratio is calculated from the total number of green
signals as compared to the total number of red signals. Specimens
will be considered within normal limits if they have a N-MYC-to-D2Z1
(green-to-red) ratio of 1.00 to 2.00, which indicates that there are an
equal number of copies of the N-myc oncogene and the centromere
2. Specimens are considered amplified if they have a green-to-red
ratio > or =4.00. (Ketterling RP: Unpublished Mayo information)
Performing Laboratory Location
Rochester
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