Amniotic fluid obtained from normal full term gestation inhibited the growth of E. coli, Staph. aureus and B. subtilis, while Ps. aeruginosa, Str. faecalis and Str. agalactiae proliferated readily in amniotic fluid. But when amniotic fluid was heated at 100 degrees C for 5 minutes, its antibacterial activity was completely lost. The levels of transferrin and lysozyme in amniotic fluid at term were determined to be 29.1+/-17.6 mg/100 ml (n= 90) and 19.1+/-8.3 micrograms/ml (n= 145), respectively. Antibacterial activity against E. coli was restored by adding transferrin into heat-treated amniotic fluid at a concentration of 250 mg/100 ml or higher, but simultaneous addition of transferrin and sufficient concentration of iron to form a transferrin-iron complex resulted in the loss of antibacterial activities. When lysozyme was added to the amniotic fluid, which had lost its antibacterial activity through exposure to heat, the antibacterial effect on B. subtilis was restored. The growth of Staph. aureus in heat-treated amniotic fluid was inhibited by the concomitant addition of lysozyme and aminobenzyl penicillin.