The preprocessing of milk:
We add milk sample into the centrifuge tube and add the equal amount of tris-saturated phenol into the milk. Then we add four times volume of chloroform into the mixture, and centrifuge the tube at 10000rpm for 5 min. (See Protocol) The result of the preprocessing was showed in the picture above.
The last competitive binding experiment is conducted under the condition of 25 times diluted PBP5-GFP solution and preprocessed milk (containing PBP-GFP) and 405nm excitation wavelength (See more in protocol). The results indicate that the fluorescence intensity measured and penicillin concentration has a correlation factor of 0.8212211, which means the two factors is strong correlation, proving that our method doesn’t just work under the most simplified conditions and has the ability to detect penicillin in milk samples.