A METHOD FOR THE DETERMINATION OF TYPE-1 IODOTHYRONINE DEIODINASE ACTIVITY IN LIVER AND KIDNEY USING I-125 LABELED REVERSE TRIIODOTHYRONINEAS A SUBSTRATE

Objectives: A detailed method for the determination of iodothyronine d eiodinase type I (DI-I) activity is described. The objective of the pr esent method development was to consolidate the effective procedures o f previous methods and produce an efficient assay that can be easily r eproduced. Design and methods: This method uses a 5',-I-125-labelled r T3 as substrate and ion-exchange chromatography to separate released i onic iodine. Released I-125- collected in the eluate is counted, and t he results used to calculate DI-I activity. Results: Results were foun d to be linear for tissue homogenates containing 3-11 mg protein . mL( -1). Day-to-day coefficient of variation of liver homogenate was deter mined to be 13%. Conclusions: This method was found to be reliable, re producible, and sample sizes as small as 10 mu L could be readily assa yed. The use of centrifuge filter units to contain the ion-exchange me dium decreased handling of the material, and potential sources of erro r.