Objective Real-time ultrasound imaging is an established objective outcome measurement with proven reliability. However, it is still largely biased by user-ability. Published research in the area of real-time ultrasound imaging reliability in the cervical region and in particular the deep cervical flexors is quite sparse. The purpose of this investigation was to examine if a novice ultrasound user could agree favourably with an experienced ultrasound sonographer in measuring the cross-sectional area of the longus colli.Methods Ultrasound images were captured from 22 healthy subjects on two different occasions, one week apart, by a novice ultrasound user. They were acquired using a GE Healthcare LOGIQe ultrasound machine, at a depth of 3-4cm with the transducer frequency set to 8MHz, in line with previous research guidelines. Cross-sectional area was then measured on-screen by both a novice and experienced ultrasound user to determine inter-rater reliability. Intra-rater reliability was also analysed using cross-sectional area figures from days one and two.Results Intra-rater reliability for real-time ultrasound imaging for the cross-sectional area of the longus colli was excellent (intra-class correlation 0.90, 95% CI 0.82-0.95). Inter-rater reliability was moderate but in keeping with previous published research (intra-class correlation 0.61, 95% CI 0.37-0.77).Conclusion Difficulties in identifying the borders of the longus colli muscle due to its deep anatomical location and surrounding structures make it difficult for assessors to agree favourably on cross-sectional area measurements, leading to moderate levels of inter-rater reliability and poor agreement. Intra-rater reliability is excellent, and in this instance indicates that a novice user can be just as reliable as a more experienced ultrasound user.