Soccer refereeing represents a highly demanding activity, requiring referees to perform complex decision-making processes under challenging physiological conditions (Weston et al. 2012).  Growing interest therefore exists with respect to the physical and physiological demands imposed during match play (Castagna et al. 2007).  However, as large levels of inter-match variability are evident in the activity profiles of referees, the detection of real systematic changes in performance outcomes between matches is difficult (Weston et al. 2011).  Simulation protocols that mimic match play represent alternative approaches that have been used to standardise the internal and external demands imposed on soccer players (Russell et al. 2011).  Nonetheless, a valid and reliable protocol does not currently exist for soccer referees.  The current study therefore sought to explore the validity and reliability of the physiological and perceptual responses elicited during a novel treadmill-based Soccer Referee Simulation (SRS).  Following the collection of baseline measures and habituation procedures, eight male soccer referees (age: 30.1 ± 3.8 years; stature: 178.4 ± 8.8 cm; body mass: 77.1 ± 10.7 kg; V̇O2max: 53.2 ± 4.1 mL·kg·min-1) completed a single trial of the SRS whereby measures of heart rate (HR), oxygen uptake (V̇O2), blood lactate concentrations, and differential ratings of perceived exertion (RPE) were obtained.  Referees’ HR responses were also monitored during a series of competitive matches (5 match observations per referee).  For the reliability aspect of the investigation, eight well-trained males (age: 25.1 ± 4.2 years; stature: 177.6 ± 9.0 cm; body mass: 79.6 ± 12.0 kg; V̇O2max: 50.6 ± 4.8 mL·kg·min-1) were initially habituated, and thereafter completed three separate trials of the SRS during which the reliability of the selected outcome variables were ascertained.  Trials were separated by 3-7 days and performed under standardised conditions (temperature: 19°C; humidity: 40%).  Paired sample t-tests explored any differences in measures of HR between the SRS and match play, with ANOVAs used to detect any systematic differences in physiological and perceptual responses between reliability trials.  Measures of relative and absolute reliability were ascertained through the calculation of intraclass correlation coefficients (ICC) and coefficients of variation (CV), respectively.  Data are presented as mean ± SD.  No differences were evidenced between the SRS and match play in relation to measures of mean HR (SRS: 79.6 ± 6.6%; Match Play: 80.6 ± 6.0%; P=0.444; ES=0.29), peak HR (SRS: 90.0 ± 7.1%; Match Play: 93.8 ± 3.5%; P=0.074; ES=0.74), or HR-based training impulse (SRS: 58.3 ± 20.2 au; Match Play: 61.6 ± 14.1 au; P=0.498; ES=0.25).  Additionally, no systematic differences were detected between reliability trials for any of the measured outcome variables (P≥0.293), whilst good levels of reliability were observed for measures of mean HR (ICC=0.94; CV=3.1%), peak HR (ICC=0.93; CV=2.2%), HR-based training impulse (ICC=0.95; CV=10.0%), mean V̇O2 (ICC=0.95; CV=2.6%); blood lactate concentrations (ICC≥0.89; CV≤11.5%), and differential RPE (ICC≥0.94; CV≤15.1%).  Together, the present findings have demonstrated the SRS to be a valid and reliable protocol that closely replicates the physiological and perceptual responses elicited by soccer referees during match play.