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. 2014 Jun 20;9(6):e100342.
doi: 10.1371/journal.pone.0100342. eCollection 2014.

ImmunoFISH is a reliable technique for the assessment of 1p and 19q status in oligodendrogliomas

Céline Duval  1 , Marie de Tayrac  2 , François Sanschagrin  1 , Karine Michaud  3 , Peter Vincent Gould  1 , Stéphan Saikali  1
Affiliations

ImmunoFISH is a reliable technique for the assessment of 1p and 19q status in oligodendrogliomas

Céline Duval et al. PLoS One. .

Abstract

Objective: To develop a new ImmunoFISH technique for the study of oligodendrogliomas by combining a standard immunohistochemical stain using MIB-1 antibody with a standard FISH technique using commercial 1p36 and 19q13 chromosomal probes.

Methods: Validation was performed by two observers on a series of 36 pre-selected oligodendrogliomas and compared to the results previously determined by FISH alone.

Results: The ImFISH technique is easy to perform and to analyze and is no more time-consuming than the usual FISH technique. Our results show that the inter-observer reliability of ImFISH is high (κ = 0.86 and 0.95 respectively for 1p and 19q). Compared to FISH, the ImFISH exhibits a very high sensitivity (∼100%) and specificity (∼90%) for 1p and/or 19q deleted cases. The sensitivity is high for normal cases (∼85%) and imbalanced cases (∼90%) with a specificity ranging between 50 and 85%. Finally, there were no significant differences between FISH and ImFISH results calculated on 60, 40 or 20 cells.

Conclusion: Our study demonstrates the reliability of the ImFISH technique in oligodendrogliomas and emphasizes its advantage in poorly cellular tumoral specimen.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
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
Figure 2. ImFISH interpretation.
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).
See this image and copyright information in PMC

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