I20 SUBCELLULAR PARTICLES 



hoped that under these conditions, the Golgi material would rise to the appro- 

 priate density interface, while the free ribonucleoprotein particles would remain 

 at or near the bottom of the tube. 



After centrifugation, a very marked band (G) consisting almost entirely of 

 recognizable Golgi material was present at density interface 1.09/1.13. A much 

 less dense band (UG) containing a few Golgi structures but mainly composed 

 of very small particulate material, was present at interface 1.13/ 1. 14. A band at 

 interface 1.14/1.15 could not be separated from the bulk of the material remain- 

 ing in the original position of the extract. 



Only I per cent of the total nitrogen of the homogenate was recovered in the 

 Golgi fraction under these conditions. This figure appears in perspective when 

 it is realized that the entire cytoplasmic particulate fraction of epididymis, as 

 isolated by differential centrifugation, accounts for but 16 per cent of the tissue 

 nitrogen. Another i per cent of the nitrogen appeared in the UG fraction, and 

 an almost immeasurably small quantity in O. Very small amounts of both RNA 

 and DPNH-cyto.r reductase activity were associated with the Golgi fraction. 

 Lipid phosphorus content and /?-glycerophosphatase activity at pH 5 (acid phos- 

 phatase), on the other hand, were very high in this fraction, and to a lesser 

 extent in UG. Alkaline phosphatase activity was still concentrated five-fold in G; 

 but it is seen that the relative specific activity was even greater in the UG 

 fraction. 



When G fractions were isolated by centrifugation for times ranging Ironi 1=5 

 to 70 minutes (fig. 6), the amount of each constituent in the fraction increased 

 progressively. However, it may be noted that the lipid phosphorus content and 

 acid phosphatase activity, as well as their relative concentrations, were appreciable 

 even at the shortest time of centrifugation, whereas alkaline phosphatase activity 

 of appreciable intensity appeared only after 30 minutes. The specific activity of 

 alkaline phosphatase in the UG layer (not shown) at 15 minutes and at all sub- 

 sequent times was higher than the value attained by the G fraction at 70 minutes. 

 The results suggest that acid phosphatase activity was associated with the large 

 Golgi structures (see fig. i), which might be expected to reach equilibrium posi- 

 tion quickly, while alkaline phosphatase was associated primarily with smaller 

 particles that entered the Golgi fraction more slowly. 



The electron microscopic appearance of a Golgi fraction isolated at 30 minutes 

 centrifugation is shown in figures 7 and 8. The material retains many of the 

 structural details peculiar to the Golgi membranes observed within the intact 

 cells: for example, the regular linear array of smooth-surfaced membranes and 

 the expansion of the intramembrane spaces to form large vacuoles. One also ob- 

 serves numerous small vesicles that appear to have been formed by the pinching 

 off or fragmentation of membrane pairs. 



Figure 9 illustrates the contrasting appearance of the UG fraction, the com- 



