_Materials required_, and their preparation:

Small tube of "washed cells" (red blood discs and
leucocytes); human cells are used in estimating the
opsonising power of the serum of experimental animals.

Small tube of emulsion of bacteria of the species
responsible for the infection of the experimental animal.

Blood pipette containing specific serum.

Blood pipette containing "pooled" serum.

_Washed Cells._--

1. Take a small centrifuge tube and half fill it with sodium citrate
solution. Mark with the grease pencil the upper limit of the fluid.

2. Cleanse the skin of the distal phalanx of the second finger of the
left hand above the root of the nail with lint and ether. Wind the
rubber tubing tightly round the second phalanx; puncture with a sterile
Hagedorn needle through the cleansed area of skin.

3. Take up a sufficiency of the issuing blood (more or less according to
the number of tests to be performed) with a teat pipette, transfer it to
the tube of citrate solution and mix thoroughly. Make a second mark on
the tube at the upper level of the mixed citrate solution and blood.

4. Place the tube in the centrifuge, counterpoise accurately and
centrifugalise until the blood cells are thrown down in a compact mass
occupying approximately the same volume as is included between the two
pencil marks.

The column of fluid in the tube now shows clear supernatant fluid
(citrate solution and blood plasma) separated from the sharp cut upper
surface of the red deposit of corpuscles by a narrow greyish layer of
leucocytes.

5. Remove the supernatant column of citrate solution by means of a teat
pipette, fill normal saline solution into the tube up to the upper
pencil mark, and distribute the blood cells throughout the saline by
means of the teat pipette. Centrifugalise as before.

6. Again remove the supernatant fluid and fill in a fresh supply of
saline solution and centrifugalise once more.

7. Remove the supernatant saline solution as nearly down to the level of
the leucocytes as can be safely done without removing any of the
leucocytes.

8. Next distribute the leucocytes evenly throughout the mass of red
cells by rotating the tube between the palms of the hands--just as is
done with a tube of liquefied medium prior to pouring a plate.

9. Set the tube upright in the plasticine block near to one end.

_Bacterial Emulsion._--

1. Take an 18- to 24-hour culture of the