The susceptibility of mastitis-causing Escherichia coli and Staphylococcus aureus to two commonly used antibiotics, tetracycline and penicillin G was tested in raw milk and in Muller-Hinton medium by introducing a pH indicator, Bromocresol purple, which was shown to be a simple, sensitive and quick method. The minimum inhibitory concentration (MIC) of penicillin G in milk were the same as those in MH, whereas the MIC of tetracycline in milk were 4-32 times those in MH. An irreversible binding between tetracycline and large molecules of milk which might probably due to the hydrophobic interaction was demonstrated by a dialysis test, suggesting the observed impairing effect was due to the action of milk on the tetracycline being tested. Further investigation reveled that much of the reduction of tetracycline’s activity in milk appears to be attributed to the milk casein protein while other heat-sensitive components in milk also play some role.