Development of an automated closed system for generation of human lymphokine-activated killer (LAK) cells for use in adoptive immunotherapy

Linda Mesler Muul, Kathryn Nason-Burchenal, Charles S. Carter, Herbert Cullis, Dale Slavin, Cornelia Hyatt, Elaine P. Director, Susan F. Leitman, Harvey G. Klein, Steven A. Rosenberg

Research output: Contribution to journalArticlepeer-review


Immunotherapy utilizing the adoptive transfer of lymphokine-activated killer (LAK) cells in conjuction with recombinant interleukin-2 (IL-2) can mediate tumor regression in some patients with advanced cancer. The activation of large numbers of LAK cells was performed in roller bottles in a research laboratory setting and required meticulous aseptic technique, at least one skilled technician per patient and one laminar flow hood per patient. To reduce the complexity and expense of LAK cell generation for human immunotherapy trials we have developed a closed-system automated procedure using a continuous flow blood cell separator. PBL were obtained by standard apheresis techniques. Platelets and plasma were elutriated using countercentrifugal flow of saline in the cell separator machine. The washed PBL were underlaid with Ficoll-Hypaque (FH) in the original separation bag. Lymphocytes were then flushed into a collection bag where they were concentrated and washed with 2 liters of saline. Mean recovery from the automated FH technique was 54.6 ± 4.3% compared to 62.3 ± 4.0% using manual methods in 50 ml tubes (P > 0.05). Cells were diluted in the collection bag with RPMI 1640 ± 2% human AB serum and could be dispensed in an automated fashion to polyolefin bags via a sample port with 1000-1500 U/ml IL-2. After 3-4 days of culture in 5% CO2 at 37°C, activated cells from the bags were harvested and washed in a closed system using the continuous flow cell separator. Cell yield from the harvest was 79.2 ± 5.4% in the automated system compared to 64.9 ± 5.0% in the standard procedure using manual harvest of roller bottles (P < 0.01). Lytic capacity of the cells against fresh human tumor in a 4 h 51Cr release assay was equivalent in cells processed either by the automated or the conventional manual method. The advantages of a closed system include decreased potential for microbial contamination and reduced labor and capital equipment costs. This technique may be easily adapted for use with other cell collection and culture systems.

Original languageEnglish (US)
Pages (from-to)171-181
Number of pages11
JournalJournal of Immunological Methods
Issue number2
StatePublished - Aug 3 1987


  • Adoptive immunotherapy
  • Cell culture
  • Interleukin-2, recombinant
  • Leukapheresis
  • Lymphokine-activated killer cell

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology


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