The mechanistic target of rapamycin complex-1 (mTORC-1) is a key nutrient and contraction-sensitive protein that regulates a pathway leading to skeletal muscle growth. Utilizing a multiplex assay, we aimed to examine the phosphorylation status of key mTORC-1-related signalling molecules in response to protein feeding and resistance exercise. Eight healthy men (age, 22.5 ± 3.1 years; mass, 80 ± 9 kg; 1-repetition maximum leg extension, 87 ± 5 kg) performed 4 sets of unilateral leg extensions until volitional failure. Immediately following the final set, all participants consumed a protein-enriched beverage. A single skeletal muscle biopsy was obtained from the vastus lateralis before (Pre) with further bilateral biopsies at 1 h (1 h exercised legs (FEDEX) and 1 h nonexercised legs (FED)) and 3 h (3 h FEDEX and 3 h FED) after drink ingestion. Phosphorylated AktSer473 was significantly elevated from Pre at 1 h FEDEX. Phosphorylated p70S6K1Thr412 was significantly increased above Pre at 1 h FEDEX and 1 h FED and was still significantly elevated at 3 h FEDEX but not 3 h FED. Phosphorylated rpS6Ser235/236 was also significantly increased above Pre at 1 h FEDEX and 1 h FED with 1 h FEDEX greater than 1 h FED. Our data highlight the utility of a multiplex assay to assess anabolic signalling molecules in response to protein feeding and resistance exercise in humans. Importantly, these changes are comparable with those as previously reported using standard immunoblotting and protein activity assays.