A Dialyzable Component of Complement

Abstract

When serum of a normal guinea pig is dialyzed against a normal saline solution with a cellophane tube∗ the hemolytic activity of the solution decreases rapidly. A prolonged dialysis will destroy almost all of the original activity. We have now found that this loss of activity is due to the loss of one of the components of complement which passes through the membrane into the saline solution, and that the original activity may be almost completely restored by the addition of the dialyzable to the non-dialyzable fraction.

A typical experiment of the dialysis is as follows: 10 cc of fresh guinea pig serum containing 80 units of complement per cc were poured into a cellophane tube both ends of which were closed with rubber stoppers and tightened with rubber bands. The sealed tube was then suspended in a graduated cylinder containing 100 cc of a cold normal saline solution. The dialysis was allowed to take place at about 5°C. After 24 hours, the bag was opened and a half cubic-centimeter of serum was pipetted out, and it was found to contain 50 units per cc. The rest of the serum was again dialyzed with another 100 cc portion of a normal saline solution. After 24 hours another half cc of the serum was taken out and its hemolytic activity was found to be 20 units per cc. The rest of the serum was again dialyzed with a fresh volume (100 cc) of the saline solution. Two days later the bag was opened and the dialyzed serum now contained less than 10 units per cc. The total volume of serum after dialysis was about 9 cc. Therefore the decrease of activity was not due to the dilution of serum. A control sample of the same guinea pig serum was stored without dialysis in another cellophane tube and kept at about 5°C. Its complement titer dropped from 80 to 60 only in 4 days.

It was of interest to ascertain whether the loss of activity during dialysis is due to the separation of the components of complement or to some chemical reaction such as oxidation. We have found that the original complementary activity may be restored by combining the dialyzed serum and the dialyzate. This was done as follows: To 0.5 cc of the dialyzed serum diluted 30-fold with 0.35% NaCl were added varying amounts of dialyzate, and enough normal saline solution to make a total volume of 2 cc. Five-tenths cc of hemolysin (2 units/cc) and 0.5 cc of 2% sheep cells were then added. After 30 minutes at 37°C readings were made. The results of a representative experiment are presented in Table I.