A minimally invasive biopsy technique was evaluated for udder tissue collection in dairy cows with Escherichia coli mastitis. Meanwhile, the effect of taking repeated liver and udder biopsies on the systemic and local acute phase response (APR) of the dairy cows was investigated during the disease. The cows were divided into a biopsy group (B) (n = 16) and a no-biopsy group (NB) (n = 16) and were sampled in the acute disease stage and in the recovery stage. The cows’ pre-disease period served as a control period for establishing baseline values for the investigated parameters. A total of 32 Holstein-Friesian cows were inoculated with 20 to 40 colony-forming units (cfu) of E. coli in one front quarter at 0 hour. Liver biopsies were collected at −144, 12, 24 and 192 h, and udder biopsies were collected at 24 and 192 h post E. coli inoculation (PI) using a minimally invasive biopsy technique. Effects of combined biopsying were investigated by recording production traits, clinical response, and measuring inflammatory milk and blood parameters: E. coli, somatic cell count, milk amyloid A (MAA) levels, white blood cell count, polymorphonuclear neutrophilic leukocyte numbers and serum amyloid A levels at several time points. E. coli inoculation changed all production parameters and the clinical and inflammatory response in all cows except one that was not infected. Combined biopsying had no constant or transient effect on the daily feed intake, the clinical responsiveness or the blood parameters, but affected the daily milk yield and some milk parameters transiently, that is, the presence of blood in milk, increased E. coli counts and MAA levels during the acute disease stage. Combined biopsying had no effect on the parameters in the recovery stage apart from the presence of blood in the milk. In conclusion, although, a minimally invasive biopsy technique was used, tissue damages could not be avoided when biopsying and they transiently affected the inflammatory parameters in the mammary gland. Nevertheless, we believe combined biopsying of liver and udder is as an acceptable approach to study the systemic and local APR in dairy cows during E. coli mastitis, if the timing of biopsying and other types of sampling is planned accordingly.