Table 1.

Comparison of immunohistochemistry (IHC), chromogenic in situ hybridization (CISH) and fluorescence in situ hybridization (FISH) techniques.

	IHC	CISH	FISH
Concept of the Method
	assessment of protein expression using antigen-specific antibodies	assessment of chromogenic effect in an enzymatic reaction	assessment of chromosomal aberration using a fluorescent probe
Advantages
preparation	-easy manual preparation [2] -automated preparation possible (e.g., Ventana BenchMark–USA) [7]	-easy manual preparation [8] -automated preparation possible (Ventana BenchMark—USA) [2,7] -permanent result of staining [2,5]	-easy manual preparation [8] -short preparation thanks to fast hybridization buffer -automated preparation possible (e.g., Ventana Medical System, Tucson, USA) [3]
analysis	-low-priced equipment for analysis (light microscope) [2] -automated analysis possible, e.g., ACIS, ChromaVision Medical Systems (San Juan Capistrano, CA) [3], or Applied Spectral Imaging, (Israel) -simple system of result evaluation (0, 1+, 2+, 3+) [9,10]	-low-priced equipment for analysis (light microscope) [2,5] -automated analysis possible (e.g., Applied Spectral Imaging, Israel) -assessment of copy-number alterations and cells morphology possible at the same time [2,5,11] -quantitative interpretation of result [2,11] -use of internal control possible [11]	-possible automatized scoring (e.g., Applied Spectral Imaging, Israel) -quantitative interpretation of result [2,8,10] -established cut-off values for probes [7,9,12,13,14,15]
other	-low-priced [2,16]	-low-priced [3]	-allows distinguishing HER2 amplification from polysomy 17 (pseudoamplification) [9] -allows detecting rearrangement and deletion at the same time (ALK, ROS1) [7] -allows detecting rearrangement with many possible gene partners (ALK, EWSR1) [16] -presence of control cells (nonneoplastic) on the same slide (internal control of preparation) [3] -analysis of concordance of results between independent observers [2]
Disadvantages
preparation	-differences in sensitivity and specificity caused by antibodies used, fixation methods [2,5,7]	-technical errors including fixation and digestion influence the final result [3]	-time-consuming when performed with standard chemicals [3] -gradual fluorescence weakening with time [5]
analysis	-semi-quantitative interpretation of results (subjective assessment) [3,17] -possible discrepancies between results (e.g., for score 2+) due to qualitative interpretation of result based on subjective judgment -necessity of re-evaluation of result of HER2 (2+) using FISH [2] -false positive result of overexpression for 3+ due to polysomy 17 (in breast cancer) -necessity of re-evaluation of positive result of ALK using FISH (sometimes negative result as well) [18]	-no ratio result for amplification [3] -necessity of extra staining to exclude polysomy, e.g., of chromosome 17 [3] -possible problems with interpretation of fusion signals [7]	-specialized equipment (fluorescence microscope with a set of filters) -limited assessment of cell features (size and shape) [2] -possible discrepancies between independent observers in low-level amplification cases, equivocal case (HER2) [3] -possible discrepancies between independent observers in borderline distance between probe parts [10]
other	-	-	-higher costs compared with the other two methods [5,7,17]
	Examples of solid tumors with use of the method
	-breast cancer [3,19] -gastric cancer [20] -lung cancer [16] -glioma [21,22] -ovarian cancer [23,24] -soft tissue sarcomas: EWS, SS, DFSP) [25,26]	-breast cancer [2,3,7] -gastric cancer [7] -lung cancer [7,27] -glioma [28] -soft tissue sarcomas: EWS, SS [29]	-breast cancer [3,9,11] -gastric cancer [30] -lung cancer [7,31] -glioma [22] -ovarian cancer [23,24] -soft tissue sarcomas: EWS, SS, DFSP [14,26,32,33]