What are the disadvantages of using mass spectrometry (MS) to detect phosphorylated proteins?

There are various disadvantages of using MS to detect phosphorylated proteins. One limitation is its difficulty in detecting low-abundance phosphorylated proteins and rare phosphosites. The sensitivity of MS is influenced by the dynamic range of the instrument and complexity of the sample. Another limitation is that MS is susceptible to false-positive and false-negative results due to contaminants or non-specific binding of phosphopeptides. False-negatives can occur when phosphorylated peptides are present below the detection limit of the instrument. Another limitation is the large amount of data generated MS-based phosphoproteomics. The technique requires complicated algorithms for data processing, quantification, and identification of proteins. Thus, expertise is required to read the data properly. Preparation of the sample requires several steps: protein extraction, enzymatic digestion, and phosphopeptide enrichment. Any of these steps may cause variability and potential loss of phosphopeptides, which ultimately affect the accuracy of the results.