Summary A study of several factors which may affect hemagglutinin (HA) production revealed that a dilute inoculum combined with a relatively late harvest time favored maximum HA titers of ECHO 6 viruses grown in monkey kidney tissue culture. Infectivity to HA ratios of 6 to 7 logs were obtained with viruses which attain high infectivity titers (B phase), while the ratios obtained with the low titer viruses (S phase) were approximately 4 to 5 logs. A survey of ECHO 6 strains revealed that 9 of 24 tested produced HA. HA production could not be correlated with infectivity titers, phase, or number of passages in tissue culture. Cross hemagglutination-inhibition (HI) tests with hyperimmune rabbit antisera revealed a high degree of specificity among the serotypes tested. Differences were noted among several ECHO 6 HA antigens in their ability to detect HI antibody. Nonspecific inhibition of ECHO virus hemagglutination was demonstrated. Adsorption of sera with 0.1% bentonite resulted in almost complete removal of nonspecific inhibitors, whereas this procedure had no apparent effect on specific HI antibody.