Label-free quantitation (LFQ) between multiple, individually-analysed samples via chromatogram realignment and peptide intensity normalisation provides an alternative to labelled quantitative approaches.
MS-based quantitation can be absolute or relative. Whilst absolute quantitation requires heavily-labelled versions of the peptide-of-interest as an internal reference standard; relative quantitation can be performed via isobaric labelling or label-free.
To compare multiple samples, relative quantitation of labelled peptides is often the method of choice as all samples are prepared in parallel and simultaneously analysed by mass spectrometry. Nevertheless, label-free quantitation (LFQ) is more appropriate when the number of samples/conditions for analysis increases (e.g., clinical patient cohorts). LFQ can also be used to determine relative quantitation between, e.g., immunoprecipitation experiments.
In the past, LFQ was based on relatively inaccurate methods such as spectral counting, With new methods emerging that accurately re-align and normalise multiple mass spectrometry chromatograms, LFQ that is based on the extraction of area-under-the-curve of eluting peptides is gaining renewed interest.
