What are the major steps of Fluorescence in situ hybridization (FISH) method?

The major steps of FISH are described in numerical order below. 

1. Probe Preparation: Short sequences of single-stranded DNA (known as probes) are prepared to target a specific portion of the gene or genomic region of interest.
2. Probe Labeling: The DNA probes are labeled using various techniques such as random primed labeling, nick translation, or PCR. The two labeling methods are direct labeling and indirect labeling. 
1. Direct Labeling: Probes are directly labeled with nucleotides using a fluorophore
2. Indirect Labeling: Probes are labeled with modified nucleotides using a hapten
3. Denaturation: Both the labeled probe and the target DNA are denatured, separating the double-stranded DNA into single strands.
4. Hybridization: The denatured probe and target DNA are combined, allowing for the annealing of complementary DNA sequences. This results in the binding of the labeled probe to its complementary target sequence.
5. Washing: During the washing step, excess, unbound probes are removed from the sample.
6. Detection: In the case of indirect labeling, an additional step is required for visualization of the non-fluorescent hapten. This typically involves using an enzymatic or immunological detection system to detect the presence of the hapten-labeled probe.