RESULTS AND DISCUSSION
Growth and viability profiles of the different platforms are shown in Figure 1a and 1b. The rolling tubes (A and B) and T-flask
(C) platforms demonstrated equivalent growth rates and cell densities. No difference in growth rate was observed between the
roller tubes being sampled successively throughout the study (A) and roller tubes sampled once, a different rolling tube each
time on a different day (B). Six-well plates (D) demonstrated very little growth during the study, which apparently resulted
from high osmolality, as explained below. Viability was very high (>96%) in all culture platforms throughout the length of
Figure 2. On each of the four days of the study, samples were taken from: A) the same 50-mL rolling tubes (volume reduced
along study), B) a single 50 mL rolling tube sacrificed for sampling (constant volume along study), C) T-flasks, and D) 6-well
plates. Error bars represent the average of two samples (duplicates). The same cell pool was used to seed the cultures on
day 0, therefore the values are the same across all platforms. A and B values for day 1 are the same because the tube sampling
regimens were shared at the first sampling point.
Nutrient and metabolite profiles are shown in Figure 2, and included: glutamine, glutamate, glucose, lactate, pH, pO2, pCO2, and osmolality. Rolling tubes of 50 mL (with either the same or different tubes being sampled each day) and T-flasks demonstrated
similar profiles for all analytes tested. Glutamine (2a) and glucose (2c) were consumed during the culture in all platforms
but the consumption rate was markedly lower for the 6-well plates. Likewise, lactate (2d) production was higher for the 6-well
plates. Glutamate (2b) also was produced during the batch culture in all platforms, but the levels varied between experimental
cases. Again, pH (2e) and gas (2f, 2g) levels were similar between the rolling tube and T-flask platforms but were relatively
lower in the last two days for the 6-well plates. Osmolality (2h) was similar between the 50-mL rolling tubes (with either
the same or different tubes being sampled each day). Slightly higher osmolality was observed with the T-flasks. However, 6-well
plates demonstrated increasingly higher osmolality levels along the culture period. We have determined that these differences
resulted from higher evaporation. Condensation was noticeable on the lids of the flask and plate culture systems. Media volume
reduction was most significant in the 6-well plate culture platform (data not shown). The data demonstrate that media volume
loss is minimal for the rolling tube platform. Because the culture contacts the entire surface of the rolling tubes, condensation
is drawn back into the culture, whereas small amounts of condensation will accumulate over time on the surfaces of the T-flask
that do not contact media.
Figure 3. Potency (a) and specific productivity (b) results on days three and four, based on a product specific assay. Samples
were collected from 50 mL rolling tubes (A), T-75 flasks (B), and 6 well plates (C). Data were normalized to T-75 flasks taken
As shown in Figure 3, potency and product-specific productivity were similar for both 50 mL rolling tube and T-flask platforms.
In contrast, potency was lower for the 6-well plate although specific productivity was apparently higher.
Performance of the rolling-tube platform is robust across a wide range of volumes. The data demonstrate that the rolling tubes
performed similarly whether or not volume was gradually reduced as samples were drawn out over the course of the analysis
(compare A and B in Figures 1 and 2). In group A, 1.3 mL of media was drawn from the (duplicate) 50-mL rolling tubes per day
on days 1 and 2, plus an additional 1.5-mL samples for submission to potency assay on days 3 and 4. From 14 mL of initial
culture, there was approximately 8.6 mL or 61% original volume left in group A tubes by day 4. Thus, decreasing volume has
no apparent impact on growth and viability (Figure 1), nutrients, or gas transfer (Figure 2).