American Society for Clinical Pathology (ASCP) Technologist in Cytogenetics certification (ASCP CG) Practice Test

How can specificity be improved in FISH analysis?

1. By using lower concentrations of probes

2. By using higher hybridization temperatures

3. By reducing the background staining

4. By increasing the postwash time

The correct answer is: By reducing the background staining

Improving specificity in Fluorescence in situ Hybridization (FISH) analysis is crucial for accurate interpretation of results. Using lower concentrations of probes can lead to reduced signal strength and may not adequately bind to the target sequences, resulting in false negatives. Higher hybridization temperatures can enhance the stringency of hybridization, but it may also lead to the loss of specific signals if the temperature is too high, which does not necessarily improve specificity. Increasing the postwash time can help reduce non-specific binding by allowing more time for unbound probes to wash away, but on its own, it does not necessarily enhance the specificity unless combined with other factors. Reducing background staining directly impacts the clarity of the target signals by minimizing noise. A lower background ensures that the true hybridization signal stands out more starkly from the nonspecific signals. This clear distinction between true and nonspecific signals enhances the capacity to identify genuine hybridization events, thus improving the overall specificity of FISH analysis.