In order to find the best method to concentrate microfilariae in the test blood, acetic acid, acetone, saponine, formalin, benzarconium hydrochloride, and distilled water were examined for hemolytic action. Also technics for concentrating microfilariae were investigated. As a result, the following method was proved to be the best.
Prepare a solution of:
Filtrated methylene blue (0.5%) 5cc
Acetone 5cc
Sodium citrate 0.2g
Water 90cc
1) Put 9 cc of the soltion in a graduated centrifuge test tube.
2) Add 1 cc of the blood drawn from the radial vein of a dog.
3) Invert the test tube several times to mix the contents, the open end being covered with a finger.(This procedure preserves the contents for a long period at room temperature.)
4) Centrifuge 1500 rpm for 10 minutes.
5) Remove the supernatant fluid with a ru-bber-bulb pipette, leaving the same quantity of fluid, usually 0.05cc, as the sediment.
6) Pick up 0.1 cc of the sediment plus the remaining fluid with a pipette.
7) Examine microscopically the mixture mo-unted on a 24×32 mm cover glass.
Microscopic observation:
As shown in the picture on p., the sediment consists of leucocytes and microfilariae. The microfilariae are in an extended position and are stained with methylene blue.
The authors'experience has confirmed that this method will detect any small number of microfilariae so long as they exist in a dog, and that it is superior to any other method previously used.