Mass Spectrometry Sample Quantitation Support—Getting Started

Relative quantitation compares the amount of an analyte in different samples. Absolute quantitation measures the actual amount of an analyte in a sample using a known standard at different concentrations. Label-free, SILAC, and TMT (Tandem Mass Tag) workflows are used for relative quantitation of protein samples. Heavy (i.e., AQUA) peptides are used as standard for absolute quantitation of target proteins.

Tandem Mass Tag technology is used to individually label different protein samples so they can be combined into a single sample for LC-MS analysis. The major advantages of this workflow are higher sample throughput, less instrument analysis time, high precision of peptide quantitation among replicates, and fewer missing quantified proteins among different samples.

We recommend using high-resolution Orbitrap Tribrid (e.g. Orbitrap Ascend or Orbitrap Eclipse), Orbitrap Exploris (e.g., 240, 480), or Q Exactive Plus series of instruments.

SILAC can be used to multiplex 2 to 3 samples. TMT reagents can be used to multiplex 6 to 11 samples. TMTpro reagents can be used to measure up to 16 samples simultaneously.

Label-free, TMT reagents, and TMTpro reagents can be used with any protein sample type. SILAC and Neucode quantitation can only be used with cell lines and model organisms that can be metabolically labeled.

TMT reagents and TMTpro reagents are different chemicals with different masses. Therefore, we do not recommend mixing the two tags as the combined samples would be significantly more complex resulting in fewer quantified proteins.