The E. coli AntiMicrobial Satellite (EcAMSat) mission investigated space microgravity effects on the antibiotic resistance of E. coli, a bacterial pathogen responsible for urinary tract infection in humans and animals. The investigation focused on spaceflight effects on bacterial antibiotic resistance and its genetic basis. The fundamental scenario of the experiment protocol started four days after launch of the EcAMSat satellite by allowing an initial growth and then starvation period for E. coli bacteria contained in 48 microfluidic wells. Both wild type (occurring in nature) and mutant strains of E. coli were used. This cycle lasted 48 hours and served to prepare the bacteria for introduction of the antibiotic agent at a time most suitable for meaningful comparisons of antibiotic dose dependence with laboratory data. After the 48-hour growth period, a buffer solution- plus-antibiotic mixture was dispensed into 3 sets of 12-well banks at three different antibiotic concentrations and is left to incubate for 48 hours. A fourth 12-well set was a control and received no antibiotic agent, only buffer solution. After 48 hours a fluid exchange occured and alamarBlue®, a dye used to determine viability of the bacteria by measuring its metabolism, was then introduced to all wells and viability was tracked for at least 48 hours. An optical system tracks viability by color change of the ‘viability dye’, which changes from blue to pink when enzymes generated by cellular metabolic processes act upon it. EcAMSat acquired data from optical measurements in real-time. It is capable of autonomous operation and stores 152 hours of data.