With respect to Dr. Sanford's comment, it is evident that more information should be obtained relative to the death rate of the cells and to the cell turnover in the population. At the present time, there is not sufficient information to give an accurate answer to this question.

However, the equations proposed for this system and the data collected to resolve the equations were based on the number of viable cells in the population.

With respect to the infection of normal bone marrow cells, it must be recognized that the cell elements in this tissue are of many kinds, and it is difficult to evaluate the changes within the first 20 days. At the present time, however, it is evident that the demonstrable response of the bone marrow cultures to the virus is the growth of myeloblasts which do not differ in morphology from the blood myeloblasts, established in culture, of the diseased chicken. The myeloblasts of bone marrow infected in culture differ from normal myeloblasts in the presence of virus-specific structures. Later Dr. Bonar will discuss the ultrastructure of the myeloblast and will describe the present knowledge of the characteristics of the virus-infected myeloblast. In our experience these ultrastructures have been evident fairly constantly during the culture periods and the release of the virus. Studies are now being made during the period prior to the 20 days after which infection of the myeloblast is obvious. Electron micrographic studies show that infection can be recognized about 12 days after exposure of the bone marrow to virus.

Dp. Eagle (National Institutes of Health): From the point of view of cellular physiology and without reference to the question of virus yield per cell, I am puzzled by the concentration of cells at which Dr. Beaudreau obtains logarithmic growth.

In all systems of human and animal cells which we have studied, logarithmic growth was observed at a range of cell concentration almost 1 percent of what you have used. That would be something like 2 to 5 times 10^5 rather than 2 to 5 times 10^7 cells per ml. This is a volume-to-volume ratio of approximately 0.1 percent. At this level, the cells constitute approximately 1 part in 1000 of the fluid volume.

You have said that the myeloblast is a small cell, which could account for part of the discrepancy, but I wonder whether it accounts for all of it. Do you know the number of cells per ml. of packed cells, which, in other animal cells, is about 2.5 X 10^8 per ml.?

Dp. Beaudreau: Yes, 1.0 ml. of packed myeloblasts contains about 2 X 10^9 cells.

Dp. Eagle: From these values, there is a tenfold ratio between the myeloblasts and mammalian cells. You are therefore getting growth at a cell population density which, in terms of cell mass, is 10 times greater than that yielding optimal growth for the mammalian cell.

For this reason Dr. Sanford's question is quite relevant. Is the growth really logarithmic? If not, and only a small fraction of the cells are growing, and if there is an important degree of cell death, your values relative to virus yield per viable cell may be much smaller than they should be.

What was the criterion of the viable cell in these very concentrated suspensions?

Dr. Beard: The best estimate of the viable cell population is that at any one time the number of dead cells in these preparations hardly exceeded 5 percent.

The counts of viable cells were made with trypan blue. It is known that this does not give accurate results, but no better way is thus far available. Thus, within the limits of the methods employed, the values given are related to viable and not dead cells.

With respect to the biologic characteristics of these cells, it has not been possible to distinguish between the myeloblasts in the bone marrow and the myeloblasts in tissue cultures, except that the myeloblasts in culture all contain virus or viroplasts.

Dr. Beaudreau: In answer to Dr. Eagle's comments, one characteristic of the myeloblast, discussed in a previous publication with Dr. Becker, is the indication that the carbohydrate utilization by this cell is very efficient. No lactate accumulates in the medium during culture, since all is actually utilized by the cell. This would give a 10-to 15-fold advantage with respect to carbohydrate utilization and energy supply for cell synthesis. This advantage, together with the cell mass and the fact that the lactate concentration does not increase, may possibly account for the difference in growth levels between the myeloblasts and mammalian cells.