Current standard procedures for the isolation and identification of Chlamydia trachomatis are laborious. A more rapid method for the isolation and identification of C. trachomatis has been developed. This method features the use of a multiwell microplate to which trypsinized McCoy cells and specimens are added simultaneously, followed by centrifugation. A simplified one-step iodine-glycerol staining procedure is used to detect the chlamydial inclusions. The requirements of the conventional macro method for preseeding cell monolayers and for the use of cumbersome cover slips in glass vials are eliminated. Studies on 345 clinical specimens showed that the micro method is as sensitive as the conventional method but can be performed more conveniently, rapidly, and economically.