9. Allow five days to elapse, then collect a preliminary sample of blood, say about 2 c.c., from the rabbit's ear. Allow it to clot, separate off the serum and transfer to a sterile test-tube. Place the test-tube in a water-bath at 56° C. for fifteen minutes (to inactivate) and test the serum quantitatively for hæmolytic properties in the following manner: THE TITRATION OF HÆMOLYTIC SERUM. _Apparatus Required:_ Electrical centrifuge. Sterile centrifuge tubes. Water-bath regulated at 56°C. Sterilised pipettes 10 c.c. graduated in tenths. Sterilised pipettes 1 c.c. graduated in tenths. Sterile test-tubes, 16 × 2 cm. Small sterile test-tubes, 9 × 1 cm. Small test-tube rack, or roll of plasticine. Capillary teat pipettes. Stout rubber band or length of small rubber tubing. _Reagents Required and Method of Preparation:_ 1. Normal saline solution. 2. Hæmolytic serum inactivated by preliminary heating to 56° C. for 15 minutes (_vide supra_) in test-tube labelled H. S. 3. Complement. Fresh guinea-pig serum in test-tube labelled C. Kill a normal guinea-pig with chloroform vapour. Open the thorax with all aseptic precautions, and collect as much blood as possible from the heart with a sterile Pasteur pipette. Transfer it to a sterile centrifuge tube and place the tube in the incubator at 37° C. Two hours later separate the clot from the sides of the tube, and centrifugalise thoroughly. Pipette off the clear serum to a clean sterilised test-tube. 4. Erythrocyte solution, in test-tube labelled E. Collect and wash human red blood cells (see page 388, 1-8). Measure the volume of red cells available and prepare a 2 per cent. suspension in normal saline solution. METHOD.-- 1. Take two test-tubes and number them 1 and 2, and pipette into each 9 c.c. of normal saline solution. 2. Add 1 c.c. of hæmolytic rabbit serum to tube No. 1 and mix thoroughly: take up 1 c.c. of the mixture and add it to tube No. 2; mix thoroughly. 3. Set up ten small test-tubes in test-tube rack or in roll of plasticine, and number 1 to 10. 4. Pipette into tube No. 1 0.5 c.c. = 0.5 c.c.} hæmolytic serum } From tube Pipette into tube No. 2 0.1 c.c. = 0.1 c.c. } H. S. hæmolytic serum } Pipette into tube No. 3 0.5 c.c. = 0.05 c.c. } hæmolytic serum } Pipette into tube No. 4 0.3 c.c. = 0.03 c.c. } hæmolytic serum } From Pipette into tube No. 5 0.2 c.c. = 0.02 c.c. } tube 1. hæmolytic serum } pipette into tube No. 6 0.1 c.c. = 0.01 c.c. } hæmolytic serum }