Diffusion restrictions in gelatin are lost by warming to 36°C, in close association with liquification of the gelatin. Diffusion of 2 kD FITC-PEG at 25 and 36°C for 20 h. Six 8-cm pipettes were filled one-half with 10 g% gelatin and one-half with gelatin-free solution containing 10 μM FITC-PEG. Three were incubated at 37°C and three at 25°C for 20 h. Gradients in pipettes incubated at 25°C were very similar to those in Figs. S7 and S8, diffusion being five times slower in the gelatin phase. In pipettes incubated at 37°C FITC-PEG diffused the entire length of the pipettes in 20 h, so that fluorescence at the loaded end decreased by nearly 20% and increased by a similar amount at the unloaded end. To account for the data, the diffusion coefficient in the gelatin phase must be close to 10⁻⁴ cm²/s. Possibly, therefore, solute movements are in some way facilitated in this condition. We conclude that diffusion restrictions in gelatin are dependent on gelatin cross-linkages and not on the molecular crowding per se of proteins in solution. AFU, arbitrary fluorescence units.