20
IASLC ATLAS OF ALK TESTING IN LUNG CANCER
A
B
Figure 4. Specimens unacceptable for analyses because of tissue overdigestion (4A) or tissue underdigestion (4B).
at least one copy each of the 5’ and 3’ signals. Because lung cancers tend to assume a wide range of growth patterns, and because tumor cells may closely intermingle with nontumor tissue elements (e.g., alveolar macrophages and lymphocytes), the accurate identification of tumor cells may be difficult in a dark field. It is advisable to always refer to a serially cut, H & E-stained tissue slide for appropriate morphologic adjustment.
Cell Classification: Signal Patterns
In concept, the genomic areas homologous to the 5' and 3' probes are molecularly very close and these signals are seen as fused, touching, or adjacent in normal cells. In contrast, when the EML4-ALK fusion gene is present, the 5' ALK green signal becomes far A B removed from the 3' ALK red Figure 5. Specimens unacceptable for analyses because of high background signal (by approximately 12.5 noise (5A) and stringy signals (5B). Mb), and the signals are seen as being split. In reality, the 3' and 5' signals may be seen as far apart from or as close to each other in normal host cells because of various degrees of condensation and three-dimensional arrangement of the chromatin. Similarly, because of the proximity of EML4 and ALK, the split can be so narrow that the signals may seem fused in ALK rearrangement (Figure 6). Furthermore, this genomic region seems to be highly unstable, and the homologous regions to one of the probes can be lost, with the corresponding signal being missing. As a result, each tumor cell may display a variety of combinations of co-localized 5'-3' ALK signals and isolated 5' or 3' ALK signals. Despite this diversity in signal profile, cells can be classified in ??H? H????[????\? ] \????\?Y?XX? ?Y?[?[X?\?[???][???\?Y
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