38 



125 

 carrier free I (New England Nuclear, carrier free, sodium 



salt, 50mCi/ml) and solid state lactoperoxidase. The lacto- 



peroxidase (Sigma Chemical Co.) was coupled to Sepharose 4B 



activated by cyanogen bromide according to David and Reisfeld 



(1974). BSA was iodinated to a specific activity of lyCi/pg 



and purified by gel filtration on Sephadex G-25 in 0.01M 



phosphate buffer, pH 7. 



Determination of pinocytic activity . For determination 



125 



of pinocytic uptake of I-BSA, AV3 and CHO cultures were 



grown in scintillation vials to near confluence. Each culture 



125 7 



received 50yl of I-BSA containing approximately 10 cpm. 



After 24 hours the media was removed and the monolayers 

 washed six times with 20ml of serum free medium. Trypsin- 

 EDTA (0. 5ml/culture) was used to detach the cells after which 

 the cells were removed to a plastic tube for counting in a 

 gamma counter (Nuclear Chicago) . The culture vessel was 

 washed an additional two times with 1.0ml medium which were 

 added to the cells. Prior to counting, the cells were separ- 

 ated from the media by centrif ugation (10 minutes at 1500rpm) 

 and the cell pellet and supernatant were counted independently. 

 In all cases, negligible activity was found in the superna- 

 tant fraction. Duplicate cultures which received 50^1 of 



125 



I-BSA 5 minutes prior to harvesting were processed as 



above. The activity associated with the cell pellet for 



these cultures was subtracted from the 24 hour culture values 



as a control for nonspecific surface absorption. Pinocytosis 



by EL4 cells was determined by similar methods with the 



