Figure 1.
Overview of ImmunoFish procedure.
In the first step, immunohistochemistry with MIB-1 antibody is performed on FFPE oligodendrogliomas. Digital images of the 10 most labelled areas are taken at high magnification (x 400). Then the slide is washed and used for a second FISH step using 1p36 or 19q13 probes. Digital images of the same 10 areas that were selected based on MIB-1 labelling are taken at the same magnification. Analysis of the two sets of images is done simultaneously on two separate screens. Only cells with MIB-1 labelling are taken into account for FISH analysis.
Figure 2.
ImFISH technique allows a simultaneous analysis of nuclear staining with MIB-1 antibody by conventional immunohistochemistry (A) and in situ hybridization with chromosomal 1p and 19q probes (B) on two separate screens. The light haematoxylin counterstaining of the immunohistochemistry step allows an easy identification of the majority of the cells analyzed: oligodendrocytes (thick arrows), astrocytes (thin arrows), neurons (short arrows) and endothelial cells (dotted arrows). Only MIB-1 labeled nuclei with an oligodendroglial morphology are analysed by FISH (framed nuclei on A and B).
Table 1.
Histological grade, MIB1 labelling Index (LI) and 1p/19q ImmunoFISH and FISH status, by ratio and combination methods.
Table 2.
Sensitivity and specificity of ImFISH according to the FISH.
Table 3.
Comparison between FISH and ImFISH results.