Hi Dipshikha,

Sequest HT score of 0 usually indicates that the match between the observed spectrum and the theoretical spectrum is very poor or non-existent. This could be due to noise, poor-quality spectra, or the presence of a peptide sequence not in the database. When setting a cutoff for protein identification and considering the quality of peptide identifications, it's important to balance between the number of unique peptides identified and the confidence in those identifications, which is often represented by a scoring metric like the Sequest HT score.

If you prioritize confidence in identifications, you might set a higher threshold for the Sequest HT score. This reduces false positives but may also reduce the number of identified proteins. If you prioritize sensitivity (identifying as many proteins as possible), you might accept lower scores or even a minimum number of unique peptides without stringent score thresholds. This approach can increase false positives.

Requiring at least one unique peptide can be a minimum criterion, but many researchers prefer two or more unique peptides for higher confidence.

It's generally advisable to exclude identifications with a Sequest HT score of 0, as they typically indicate low confidence. However, if these identifications are supported by additional strong evidence (such as multiple unique peptides), they may still be considered, though cautiously.

Ultimately, the choice of cutoff will depend on your specific research question, the quality of your data, and the acceptable balance between sensitivity and specificity in your results.

Purvi

Thank you so much for the clarification. It really helps a lot in understanding the data.