The ESI-cleavable TNT mass markers described in this article have several properties that make them very useful for quantitative, multiplex assays, including the following: (i) The TNT portion can be elaborated into very large arrays of tags using only small numbers of starting components. The 20 standard amino acids as well as the large number of isotopic variants of these amino acids that are available provide the possibility of synthesis of numerous marker molecules that are easily resolved by the unique combination of parent and daughter ion massto-charge ratios; (ii) The ESI-cleavability allows direct analysis of solution phase assays without complex workup of the samples unlike MALDI, which requires that samples are spotted onto targets; (iii) The ESI-cleavable linker connecting the DNA and the TNT components cleaves virtually instantaneously during electrospray ionisation that will allow separations such as capillary electrophoresis to be performed in-line with the MS/MS analysis; (iv) In-line separation allows for a further level of multiplexing over and above the large numbers of available tags since the probes can be identified by their elution time as well as by their tag and in future work, we will explore the use of this feature to enable in-line coupling of analytical separations such as capillary electrophoresis; (v) sets of isotopic TNTs can be synthesized that behave identically during separations, hybridizations and labelling reactions enabling accurate measurements of quantities of target nucleic acids without 'dye effects' widening the range of applications for which mass tags can be employed.