Comparison of proteomic profiles of serum, plasma, and modified media supplements used for cell culture and expansion

Abstract

Background: The culture and expansion of human cells for clinical use requires the presence of human serum or plasma in culture media. Although these supplements have been extensively characterized in their chemical composition, only recently it has been possible to provide by high throughput protein analysis, a comprehensive profile of the soluble factors contributing to cell survival. This study analyzed and compared the presence of 100 proteins including chemokines, cytokines and soluble factors in six different types of media supplements: serum, plasma, recalcified plasma, heat inactivated serum, heat inactivated plasma and heat inactivated recalcified plasma.

Methods: Serum, plasma, recalcified plasma, and heat inactivated supplements were prepared from ten healthy subjects. The levels of 100 soluble factors were measured in each sample using a multiplexed ELISA assay and compared by Eisen hierarchical clustering analysis.

Results: A comparison of serum and plasma levels of soluble factors found that 2 were greater in plasma but 18 factors were greater in serum including 11 chemokines. The levels of only four factors differed between recalcified plasma and plasma. Heat inactivation had the greatest effect on soluble factors. Supervised Eisen hierarchical clustering indicated that the differences between heat inactivated supplements and those that were not were greater than the differences within these two groups. The levels of 36 factors differed between heat inactivated plasma and plasma. Thirty one of these factors had a lower concentration in heat inactivated plasma including 12 chemokines, 4 growth factors, 4 matrix metalloproteases, and 3 adhesion molecules. Heat inactivated decalcified plasma is often used in place of heat inactivated serum and the levels of 19 soluble factors differed between these two supplements.

Conclusion: Our report provides a comprehensive protein profile of serum, plasma recalcified plasma, and heat inactivated supplements. This profile represents a qualitative and quantitative database that can aid in the selection of the appropriate blood derived supplement for human cell cultures with special requirements.

Figure 1

Unsupervised Hierarchical clustering (Eisen Kendall's) of 80 soluble factor levels among a total of 40 samples including 10 serum (S), 10 plasma (P), 10 recalcified plasma (CP), and 10 heat inactivated plasma (P-HI) samples prepared from whole blood from 10 healthy subjects (01 through 10). The levels of the soluble factors were measured by multiplexed ELISA.

Figure 2

Factors that distinguish heat inactivated plasma from plasma, serum, and recalcified plasma. Hierarchical clustering analysis was applied to 80 soluble factors measured in a total of 40 serum, plasma, and plasma derived components (as per Figure 1). The ten heat inactivated plasma components separated into a single group with one recalcified plasma component. A cluster of 15 soluble factors best distinguished the heat inactivated group from the other 29 components.

Figure 3

Supervised hierarchical clustering (Eisen Kendall's) of 80 soluble factors among 60 samples from 10 subjects. The samples included 10 sera (S), 10 plasma (P), 10 recalified plasma (CP), 10 heat inactivated sera (S-HI), 10 heat inactivated plasma (P-HI), and 10 heat inactivated recalified plasma (CP-HI). The serum, plasma, and recalcified plasma were prepared, stored at -80°C and tested. The heat inactivated samples were prepared, stored at -80°C, thawed, heat inactivated, stored again at -80°C and tested. Red bar encompasses regular serum, plasma and recalcified plasma. Black bar heat inactivated samples.