In order to obtain results showing greater repeatability in morphology and sperm function assessment, several laboratorial techniques have been developed continuously. Systems that utilize computerized image analysis, staining methods using fluorescent dies (fluorescent probes) in epifluorescence microscopy or flow cytometry, that allowed a more judicious evaluation of sperm structure integrity arose. Authors describe experiences, over a decade, with the use of these biotechiques, critically analyzing and pointing out future challenges, including the field of sperm molecular biology.