A microculture neutralization test is described for measuring specific antibody levels to the 35 human adenovirus serotypes in Vero cells. The test is read at 5 days by macroscopic observation after staining the uninfected cells with crystal violet. The test is performed with a minimum of manipulations and gives serum titers comparable with those obtained in tube macrocultures of monkey kidney, human embryonic kidney, and Vero cells. The Vero microculture neutralization test measures inhibition of adenovirus toxicity, although selected human adenoviruses serially subpassaged in Vero cells were shown to successfully adapt and replicate in the absence of detectable helper viruses.