Umbilical cord derived stem cell culture: Multiple-harvest explant method

Jeanne Adiwinata, Isabella Kurnia Liem, Ecie Budiyanti, Iqbal Fasha, Lauda Feroniasanti, Taufik Jamaan, Kanadi Sumapradja

Research output: Contribution to journalArticlepeer-review

15 Citations (Scopus)

Abstract

Published methods of umbilical cord explant culture require removing the explant after the MSCs are confluent. However, we observed that the explants were still attached after the MSCs were detached using TrypLE Select. Therefore, the objective of this study was to do explant cultures in xeno-free media, and reculture the explants after the first harvest to test whether the explant could grow further to yield more stem cells. In this study, umbilical cord explant culture were done in 10% platelet rich plasma (PRP) containing alpha minimal essential medium (αMEM) or Dulbecco's modified Eagle's medium (DMEM), or 10% human AB or cord blood serum (CBS) containing αMEM. Three explants were cultured in each well of a twelve well plate in triplicate. The cells were harvested using TrypLE Select, and cell yield was counted after cell growth achieved 80-90% confluence. The remaining attached explants were re-cultured in the respective media for several times after harvesting, until the explants were detached or until there was no more cell growth from the explants. Cell counts for every harvest were noted. Cumulative cell yield from each well for each medium was computed and tabulated. Our results showed that the time needed for cell growth was variable between explants. Therefore, in one well, the three explants might grow with different speed, and in many instances, harvests were done before optimal growth in all explants occurred. The maximal number of harvest was four times, which occurred in 10% human AB serum supplemented αMEM (two times) followed by two harvest in 10% autologous CBS supplemented αMEM, and yielded a cumulative amount of 144,000 viable cells in 37 days. In conclusion, we have developed a multi-harvest explant method to culture umbilical cord stem cells in various xeno-free media that yield far more numerous viable cells compared to standard explant method.

Original languageEnglish
Pages (from-to)1202-1208
Number of pages7
JournalInternational Journal of PharmTech Research
Volume6
Issue number4
Publication statusPublished - 1 Jan 2014

Keywords

  • Explant culture
  • Multiple harvests
  • Stem cell
  • Thrombocyte concentrate
  • Umbilical cord

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