The calibration equation is derived from the relationship of kinetic energy to mass and velocity. Rearranging Equation 1 to solve for velocity, v, gives the relationship in Equation 2. Velocity is distance divided by time or d / t. Substituting d / t into Equation 2 and solving for t produces Equation 3.

KE = 0.5mv^{2 }[Eq. 1]

v = [2KE/m]^{1/2} [Eq. 2]

t = m^{1/2} * d/[2KE]^{1/2} [Eq. 3]

Distance and KE are assumed to be constant and can be replaced with A. The relationship between t and m^{1/2} is linear and an intercept is added to produce a simple equation for a straight line (Equation 4).

t = Am^{1/2} + B [Eq. 4]

Solving the calibration equation requires two standards. The equation is solved for the two unkowns A and B using the known m/z ratios and their flight times. The current technology permits mass measurement accuracy of 10 parts-per-million (ppm) or better for peptides in the range of 500-5000 m/z [1]. Mass measurement accuracy of 100 ppm or better can be obtained for proteins in the range of 5000-30000 m/z [2].

*Time-of-Flight Mass Spectrometry – Instruments and Applications in Biological Research* by Robert J. Cotter [3] is an excellent resource for learning the basics of TOF mass spectrometry.

** References: **

[1] Edmondson, R.D., Russell, D.H., *J. Mass Spectrom.*, **1996**, *7*, 995-1001.

[2] Watkins, L.K., Bondarenko, P.V., Cockrill, S.L., Song, S., Barbacci, D.C., Russell, D.H., Macfarlane, R.M, *J. Chrom. A.*, **1999**.

[3] Cotter, R.J., Time-of-Flight Mass Spectrometry – Instruments and Applications in Biological Research; *American Chemical Society: Washington, DC*, **1997**.