Only 60% of the MethoCult GF+ cultures obtained from CAFC of LTC-IC gave rise to HPP-Q (High Proliferative Potential-Quiescent) colonies, while all cultures had BFU-E, GM and GEMM colonies by day 14, as shown in Figure 2.
LTC-IC counts from CD34+ selected cord blood cells (range: 1261-2906, mean: 1966[+ or -]808) Samples LTC-IC/CD34+ cells 1 2705 2 2906 3 2580 4 1649 5 1781 6 1280 1649 8 2499 9 1350 10 1261 Mean Value 1966 [+ or -] 808
Although the number of articles on cord blood LTC-IC are somewhat limited, our LTC-IC/CD34+ cell numbers were lower compared to some of the published studies (13), (14), but results of several studies were consistent with ours (15).
LTC-IC assay with limiting dilution is relatively easier than the other LTC-IC assays, since the preprepared murine bone marrow stromal cell line plates can be used in five weeks, and counting CFUs is easier than counting CAFCs.
The role of LTC-IC in long-term engraftment is unknown; however, they are the most primitive progenitors that can be detected in an in vitro assay.
A recent study reported that type of stromal feeder layer used in LTC-IC LDA affects the determination of LTC-IC frequencies in uncultured cells and also has a significant effect on cultures.
The temperature of the LTC-IC was kept at 35[degrees]C in our study.
Even though there has been some progress in the standardization of the LTC-IC culture methods, a more standardized approach will help us to achieve better results for ex vivo expansion of hematopoietic cells.
We also suggest that more information about the in vivo scenario can be gained with further insight into the use of human cord blood stromal cells or HUVEC (human umblical vein endothelial cells) as feeder layers and with exploration of in vitro behavior of LTC-IC for CFC production.