High cell density fermentation production of recombinant human granulocyte-macrophage colony stimulating factor (GMCSF) was studied in an Escherichia coli secretory expression system. By using glycerol instead of glucose as a carbon source and feeding in a specific fed-batch mode and by controlling the temperature at about 30 degrees C and maintaining the dissolved oxygen level at 20 similar to 25% saturation, cell density increased from 30 to 91.8 g l(-1), the production of recombinant GM-CSF remained high and the fermentation time was reduced from 45 to 30 h. Recombination GM-CSF was purified by hydrophobic column-molecular sieve-ion exchange column chromatography. The expression of recombinant GM-CSF was not affected by fermentation methods, the production of GM-CSF reached 28.04% (about 1.68 g l(-1)) and 27.51% (about 1.64 g l(-1)) of total somatic proteins for high cell density and normal fermentation, respectively. The corresponding value for specific activity of GM-CSF was 2.6 x 10(7), and 2.4 x 10(7), respectively.