The effects of multiple freeze-thaw cycles and various temperatures (-20 degrees C, 4 degrees C, 25 degrees C, 37 degrees C) on the reactivity of human immunodeficiency virus (HIV) antibodies were evaluated using current ELISA, recombinant, and Western blot methodologies. Twenty consecutive freeze-thaw cycles and storage of specimens at -20 degrees C and 4 degrees C for 57 days resulted in no loss of HIV antibody reactivity nor false positive samples. Maintenance of clinical specimens at 25 degrees C and 37 degrees C for 57 days resulted in some loss of HIV antibody reactivity, but all positive and weakly reacting samples remained positive, and negative samples were unaffected.