We herein describe a cross-sectional multiple-point laser Doppler velocimetry (CS-MLDV) system for monitoring blood vessels that are sutured and connected during an operation. In order to observe the condition of a blood vessel during an operation, the previously developed linear MLDV (L-MLDV) system can realize velocity distribution imaging of the carotid artery in a living mouse by means of traverse laser light. We subsequently developed a CS-MLDV system, which can measure the instantaneous two-dimensional (2D) flow velocity, by upgrading the optical components and signal processing used in L-MLDV. The validity of the CS-MLDV results was verified through comparison with the results of a computational fluid dynamics (CFD) analysis. The results of the CFD analysis were similar to the experimental results obtained under the same flow field condition. Moreover, an instantaneous 2D velocity distribution can be obtained even for the case of flowing blood. Finally, we carried out in-vivo measurement in a mesenteric vessel of a mouse in order to demonstrate the potential of the CS-MLDV for use in surgery. (C) 2015 Elsevier B.V. All rights reserved.