The molecular size of microsomal membrane proteins from frozen porcine thyroids before and after solubilization by proteolytic and non-proteolytic techniques has been investigated by means of polyacrylamide-gel electrophoresis in the presence of 1% sodium dodecylsulfate. When thyroid microsomal membrane proteins are solubilized by non-proteolytic methods such as high pH, n-butanol, or deoxycholate, no major change in the electrophoretic pattern compared to untreated microsomes has been observed, thereby suggesting that these non-proteolytic methods are capable of extracting membrane proteins from thyroid microsomes without altering their molecular size. However, treatment of microsomes with protein-solubilizing levels of trypsin (1-5 mug trypsin per mg thyroid protein) results in degradation of all major proteins with a molecular weight greater than 30 000. The high-molecular-weight proteins are particularly susceptible to attack by trypsin. Thus, these experiments indicate that the use of trypsin to solubilize thyroid microsomal membrane proteins, particularly thyroid peroxidase, will result in fragmented proteins and should be avoided if intact membrane proteins are desired.