The rickettsiae content of infected chick embryo yolk-sacs may be determined by subinoculation of serial tenfold dilutions of the homo­genized material into the yolk-sacs of 7-day-old cruck embryos, 12 or more per dilution. Following inoculation, eggs are incubated at 35°C. and candled daily. All eggs dying during the first five days are dis­carded without further examination and are considered as nonspecific deaths since the organisms would not have reached a detectable level with.in this short time. From the 6th day through the 11th day after inoculation, the yolk-sacs of embryos dying each day are removed, smears are stained and examined microscopically for the presence of rickettsiae. The yolk-sacs from the remaining live embryos are similarly examined on the 11th and final day of incubation. From the data thus obtained, the egg infective dose, or the ID50 of the material can be calculated.
(The number of eggs per dilution will vary owing to the nonspecific early deaths mentioned above, although there were originally 12 eggs per dilution.) The results of such a titration are shown in the following example:
Dilution of yolk sac material: 10^-3, 10^-4, 10^-5, 10^-6, 10^-7, 10^-8
Number of yolk sac positive: 10, 08, 09, 05, 02, 00
Number of yolk sac negative: 00, 01, 02, 07, 08, 09

a: For the above data calculate, using the Reed-Muench Method, the LD5o value.
b: What does this 1D6o represent?