128 NEUTEON EFFECTS ON ANIMALS 



irradiation doses in single 6-7-hour exposures on each of 3 successive days, 

 followed by a 55-n dose in 3 hours on the fourth day, cage No. 4 (2) being 

 used for these irradiations. The control animals were placed in any avail- 

 able cages for 5 minutes 6 days a week for a year but received no irradiation. 

 The daily irradiation dose, and total irradiation received by each dog are 

 shown as part of Table I. 



Preparation of Plasma Satnples. To ensure clear, transparent plasma, 

 blood samples were taken from animals from which all food, except water, 

 had been withheld overnight. About 20 ml. of blood were obtained by 

 venipuncture from the jugular or the radial vein, and immediately trans- 

 ferred to tubes containing dr\^ lithium oxalate powder. The plasma ob- 

 tained was stored in sterile vials close to the freezing coils in a refrigerator 

 until used. 



In preparation for electrophoresis, 5 ml. of plasma were added to 10 ml. 

 of a phosphate-saline buffer solution of pH 7.7, of ionic strength 0.2, and 

 of composition 5.76 ml. of 0.2M NaH.POa, 94.24 ml. of 0.2M Na.HPOi 

 and 50 ml. of 3M NaCl in 1 liter of buffer solution. The diluted plasma 

 was then placed in Yisking tubing and the tube securely closed by tying 

 in such manner that a slight pressure was put upon the plasma solution, 

 thus assuring against the entry of any considerable quantity of water, 

 with corresponding dilution of the plasma solution, during dialysis. The 

 diluted plasma was dialyzed at refrigerator temperature for 40-50 hours 

 against approximately 1600 ml. of the buffer solution, with occasional 

 agitation. 



Electrophoresis Measurements. Electrophoresis experiments were made 

 with a standard Tiselius-Ivlett electrophoresis apparatus using a standard 

 11 ml. single section cell. During electrophoresis the current was main- 

 tained at 35 ma., resulting in a field strength of 5 volts/cm. in the cell. 



The scanning photograph obtained in the usual manner (3) of the ascend- 

 ing (positive) arm of the cell after 6 hours of electrophoresis was taken for 

 the determination of the relative amounts of the plasma constituents 

 present, the positive arm usually giving clearer separation of the constitu- 

 ents and being free of anomalous effects due to slight precipitate formation 

 which often occurred in the negative arm. 



The enlarged tracing of the curve was divided into the various com- 

 ponents by drawing an ordinate between the base line and the lowest point 

 between aojacent peaks, following the procedure of Tiselius and Kabat (4) 

 and Longsworth (5). The areas under the various peaks were measured 

 with a planimeter, and the relative composition of the plasma evaluated 

 by determining the percentage of the total area (excluding that of the delta 

 boundary) contributed by each component. 



