Protein carbonyls were studied in aging and exercise by immunoblot followed by one- or two-dimensional polyacrylamide gel electrophoresis using antibodies against 2,4-dinitrophenylhydrazones. Proteins of rat kidneys exhibited significant age-related increase in the amount of carbonyl while those of the brain and liver did not. Major carbonylated proteins in the kidney included serum albumin. In nematodes in which protein carbonyls increased with age, one of the carbonylated proteins was identified as vitellogenin, an egg-yolk protein. A possible biological significance of this protein present in abundance even after egg-laying stages is discussed in terms of protection against oxidative stress. Exhaustive exercise induced significant increase in the carbonylation of selected but unidentified proteins in the lung. This oxidative stress might be caused by xanthine oxidase in this tissue and hypoxanthine derived from ATP-depleted muscles. Exercise at high altitude caused higher carbonylation of the skeletal muscle proteins, most notably a protein likely to be actin, than that at sea level but no significant difference was observed in lipid peroxidation. These studies emphasize the value of immunoblot analysis of tissue protein carbonyls in a variety of situations where oxidative stress is likely involved.