This document is intended to provide guidance for the design, development and statistical validation of in vivo assays residing in flow schemes of discovery projects. It provides statistical methodology for pre-study, cross-study (lab-to-lab transfers and protocol changes), and in-study (quality control monitoring) validation. Application of the enclosed methods will increase confidence in data from in vivo assays in the critical path and enable better decisions about SAR directions and compound prioritization. Screens using both single dose and multiple dose in vivo assays are discussed, along with acceptance criteria at different stages of validation.