64 LABORATORY COURSE IN SERUM STUDY 



Prepare a suspension of typhoid bacilli by adding 10 c.c. of salt 

 solution to an agar slant culture and gently rubbing the growth from 

 the surface of the agar by means of a platinum loop or a capillary glass 

 pipette, being careful not to cut the surface of the agar. Shake the tube 

 gently without wetting the stopper and allow to stand for 10 minutes 

 so that the larger clumps may settle to the bottom. Take a capillary 

 glass pipette, plug it at the suction end with non-absorbent cotton and 

 provide it with a rubber nipple. Draw off 8 or 9 c.c. of the upper por- 

 tion of the suspension, being careful not to disturb the sediment and 

 place in a separate test tube, taking care that the suspension does not 

 flow against the upper portions of the tube. Add 0.2 c.c. of 40 per cent 

 formaldehyde and place in water bath at 56 for thirty minutes. If 

 clumps are still visible the suspension may be filtered through a moist 

 paper. 



B 



Prepare the following dilutions of the immune serum in test tubes : 



1. 1-10 = dilution as distributed 



2. 1-50 = 0.5 c.c. of 1-10 dilution -f 2 c.c. salt solution 



3. 1-250 = 0.5 c.c. of 1-50 dilution + 2 c.c. salt solution 



4. 1-500 =1.0 c.c. of 1-250 dilution + 1 c.c. salt solution 



5. 1-1000 = 1.0 c.c. of 1-500 dilution + 1 c.c. salt solution 



6. 1-2000 =1.0 c.c. of 1-1000 dilution + 1 c.c. salt solution 



Place 0.5 c.c. of each dilution of serum (1-10 to 1-2000) in a series of 

 agglutination tubes and in an additional tube 0.5 c.c. of salt solution for 

 control. 



To each tube add 0.5 c.c. of the killed typhoid suspension. This, 

 of course, doubles the dilution in each case and the tubes should be 

 labeled 1-20, 1-100, 1-500, etc., up to 1-4000. Shake gently till the 

 two fluids mix. The typhoid suspension should be distributed by means 

 of a pipette carefully plugged at the upper end with non-absorbent 

 cotton. This should be used wherever bacteria are handled with a 

 pipette, even when they are thought to have been killed. 



Place the tubes in the water bath, at 37 C., for one hour, and 

 observe carefully by transmitted light. Record the results and 

 place the tubes in the ice box for one half hour and again make a 

 reading. 



