The modeling of isotope distributions can be used to evaluate intracellular fluxes and to investigate cellular metabolism. A method of modeling isotope distributions in biochemical networks that addresses some of the shortcomings of conventional methods is presented. Matrix equations representing steady state isotope balances are formulated for each metabolite and solved iteratively via computer. The key feature of this method is the use of atom mapping matrices, which decouple the generation of the steady state equations from the details of the transfer of carbon atoms from reactants to products. The use of atom mapping matrices results in a clear, intuitive description of metabolic networks that is easy to develop, check, and modify. A network representing energy metabolism in a hybridoma cell line was developed and presented as an example of the method. A program that uses the atom mapping matrix method to calculate the isotope distribution in the network as a function of the intracellular fluxes was written. Calculations showed that a single nuclear magnetic resonance (NMR) experiment with C-13-labeled glucose should be able to uniquely determine two important TCA cycle flux ratios. These results demonstrate how in vitro NMR can be used to study cellular metabolism or to validate the flux estimates obtained from other methods.