The Vision in Preschoolers (VIP) Study Group conducted a multicenter, multidisciplinary, two-phase study to evaluate the performance of vision screening tests for identifying pre-school children with amblyopia, strabismus, significant refractive error, or unexplained reduced visual acuity (VA). The results of the VIP Study provide evidence-based guidelines for preschool vision screening. The best screening tests administered by eye care professionals were non-cycloplegic retinoscopy, Retinomax Autorefractor, SureSight Vision Screener, and linear, crowded Lea Symbols VA at 10 feet. The best screening tests administered by trained nurses and/or lay screeners were Retinomax, SureSight, and VIP single, crowded Lea Symbols VA screening test system at 5 feet. Eye care professionals can improve detection of strabismus by combining unilateral cover test with a refraction test and trained lay screeners can improve detection of strabismus by combining Stereo Smile II with SureSight. The best performing tests had high testability whether performed by trained eye care professionals, nurses, or lay screeners (≥98%). Although very few children were unable to complete these tests, a child who was “unable” was much more likely to have a vision problem than a child who passed; therefore, children who are unable to complete one of these tests should be referred for further evaluation. When screening using the Retinomax, repeated testing to achieve the manufacturer's suggested confidence number is valuable and improves specificity. Federal initiatives to increase the number of pre-school children receiving vision screening or examination will increase the number of pre-school children identified with amblyopia, strabismus, and/or significant refractive error. Although there is general agreement regarding the importance of early detection of amblyopia, controversy exists regarding the importance of early detection of refractive error. Because of the high prevalence of significant refractive errors and lack of evidenced-based guidelines for correction of refractive error in pre-school children, future research is needed to evaluate the value of correcting refractive errors in preschoolers who do not have amblyopia and/or strabismus.