Escherichia coli K-12 cells were grown in a confined volume using microporous hollow fiber membranes. The local cell concentrations in the reactors were above 400 g dry mass/L, in excess of the predicted limit based on the specific volume of free cells determined by tracer exclusion. Cell mass synthesis and degradation rates in these reactors were measured using radioisotope labeling with (35)S. Net accumulation of cell material persisted at these high cell densities. The rates of substrate uptake and cell growth were predicted from the theory of reaction and diffusion assuming that kinetics of cell metabolism are identical for free-living and immobilized cells. This theory was tested by comparison of overall rates and by the size of the region in which cell growth occurred, measured by autoradiography. A yield coefficient of 4 +/- 1 mol sulfur/mol glucose was measured, in agreement with the value determined for free-living cells in similar conditions. Cell growth occurs in a thin layer (10-30 microm), at a rate similar to the growth rate for free cells. Volume expansion by the cells as a consequence of proliferation induces convection of cell mass out of the growth region into a region of the reactor filled with starving cells, which then accumulate in the reactor. The combination of mass-balance and spatial distribution measurements made possible by the use of radioisotope labeling enables a direct test for mass transfer limitations, the determination of the intrinsic cell kinetics, and noninvasive measurements of cell growth in immobilized cell reactors.