Simplified method of the growth of human tumor infiltrating lymphocytes in gas-permeable flasks to numbers needed for patient treatment

Jianjian Jin, Marianna Sabatino, Robert Somerville, John R. Wilson, Mark E. Dudley, David F. Stroncek, Steven A. Rosenberg

Research output: Contribution to journalArticlepeer-review

65 Scopus citations


Adoptive cell therapy of metastatic melanoma with autologous tumor infiltrating lymphocytes (TIL) is clinically effective, but TIL production can be challenging. Here we describe a simplified method for initial TIL culture and rapid expansion in gas-permeable flasks. TIL were initially cultured from tumor digests and fragments in 40 mL capacity flasks with a 10 cm gas-permeable silicone bottom, G-Rex10. A TIL rapid expansion protocol (REP) was developed using 500 mL capacity flasks with a 100 cm 2 gas-permeable silicone bottom, G-Rex100. TIL growth was successfully initiated in G-Rex10 flasks from tumor digests from 13 of 14 patients and from tumor fragments in all 11 tumor samples tested. TIL could then be expanded to 8-10× 10 cells in a 2-step REP that began by seeding 5× 10 TIL into a G-Rex100 flask, followed by expansion at day 7 into 3 G-Rex100 flasks. To obtain the 30-60× 10 cells used for patient treatment, we seeded 6 G-Rex100 flasks with 5× 10 cells and expanded into 18 G-Rex100 flasks. Large-scale TIL REP in gas-permeable flasks requires approximately 9-10 L of media, about 3-4 times less than other methods. In conclusion, TIL initiation and REP in gas-permeable G-Rex flasks require fewer total vessels, less media, less incubator space, and less labor than initiation and REP in 24-well plates, tissue culture flasks, and bags. TIL culture in G-Rex flasks will facilitate the production of TIL at the numbers required for patient treatment at most cell processing laboratories.

Original languageEnglish (US)
Pages (from-to)283-292
Number of pages10
JournalJournal of Immunotherapy
Issue number3
StatePublished - Apr 2012
Externally publishedYes


  • G-Rex flasks
  • adoptive cell therapy
  • cancer
  • melanoma
  • tumor infiltrating lymphocytes

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology
  • Pharmacology
  • Cancer Research


Dive into the research topics of 'Simplified method of the growth of human tumor infiltrating lymphocytes in gas-permeable flasks to numbers needed for patient treatment'. Together they form a unique fingerprint.

Cite this