Context .- Biospecimens must have appropriate clinical annotation (data) to ensure optimal quality for both patient care and research. Additional clinical preanalytic variables are the focus of this continuing study. Objective .- To complete the identification of the essential preanalytic variables (data fields) that can, and in some instances should, be attached to every collected biospecimen by adding the additional specific variables for clinical chemistry and microbiology to our original 170 variables. Design .- The College of American Pathologists Diagnostic Intelligence and Health Information Technology Committee sponsored a second Biorepository Working Group to complete the list of preanalytic variables for annotating biospecimens. Members of the second Biorepository Working Group are experts in clinical pathology and microbiology. Additional preanalytic area-specific variables were identified and ranked along with definitions and potential negative impacts if the variable is not attached to the biospecimen. The draft manuscript was reviewed by additional national and international stakeholders. Results .- Four additional required preanalytic variables were identified specifically for clinical chemistry and microbiology biospecimens that can be used as a guide for site-specific implementation into patient care and research biorepository processes. Conclusions .- In our collective experience, selecting which of the many preanalytic variables to attach to any specific set of biospecimens used for patient care and/or research is often difficult. The additional ranked list should be of practical benefit when selecting preanalytic variables for a given biospecimen collection.