Loucy

Hematopoietic cell lines PEER, K-562, MONO-MAC-6, HL-60, OCI-AML3, JURKAT, DAUDI, KASUMI-1, LOUCY, MV-4-11, and THP-1 were available in house, DEL and JVM-2 were purchased at the DSMZ repository (Braunschweig, Germany), and KARPAS-422 at Sigma-Aldrich (Saint Louis, Missouri, USA). Cell lines were grown in RPMI medium (Invitrogen, Waltham, MA, USA) supplemented with 10% or 20% Fetal Calf Serum (FCS, ThermoFisher Scientific, Waltham, MA, USA), according to supplier instructions, together with 100 U/mL Penicillin/Streptomycin (10,000 U/mL, Invitrogen) and 100 µg/mL l-glutamine (200 mM, Invitrogen). For THP-1, medium was additionally supplied with 0.05 mM β-mercaptoethanol. Cell lines were incubated at 37 °C in 5% CO₂ incubators.
Mononuclear cell preparations derived from spleen and PB from 3 JMML patients were primary cultured in StemSpan SFEM II medium (Stemcell Technologies, Vancouver, Canada) supplemented with recombinant human IL3 (0.01 µg/mL; PeproTech, London, UK), FLT3L (0.01 µg/mL; PeproTech, London, UK), TPO (0.01 µg/mL; PeproTech, London, UK) and SCF (0.025 µg/mL; PeproTech, London, UK) and incubated at 37 °C in 5% CO₂ incubators.

Leukemia cell lines, LOUCY, MEGAL, and OCI-AML3 were purchased from DSMZ (Braunschweig, Germany). K562, HL60, and HEL cells were obtained from JCRB cell bank (Ibaraki, Osaka). These cell lines were authenticated using STR typing by DSMZ or JCRB cell bank, and also the mycoplasma status was checked by them. LOUCY, MEGAL, and HL60 cells were cultured in RPMI1640 medium (Sigma) supplemented with 20% FBS (Sigma). K562 and HEL cells were cultured in RPMI1640 medium supplemented with 10% FBS. OCI-AML3 was cultured in alpha-MEM medium (WAKO) supplemented with 20% FBS. EB3 ES cells (Niwa et al., 2002 (link); Ogawa et al., 2004 (link)), E14tg2a ES cells (Hooper et al., 1987 (link)) and their derivatives, and HeLa cells were cultured as described previously (Oka et al., 2016 (link)).