L glutamine

RBL-MRGPRX2 cells were generated as previously described [24 (link)]. RBL-MRGPRX2 cells and their parental RBL-2H3 (herein referred to as RBL) counterparts were maintained at 37 °C, in a humidified incubator with 5% CO₂, in adherent cell cultures in low-glucose DMEM (cat # 01-050-1A, Biological Industries, Beit-Haemek, Israel) supplemented with 10% FBS (cat # 12657, GIBCO, Grand Island, NY, USA), 2 mM L-Glutamine (cat # 03-020-1A, Biological Industries), 100 μg/mL streptomycin and 100 U/mL penicillin and 12.5 U/mL nystatin (Biological Industries, Beit-Haemek, Israel), except for RBL-MRGPRX2 cells that were additionally supplemented with 1 mg/mL of G418 (cat # A1720, Sigma Aldrich, St Louis, MO, USA). LAD-2 cells (a kind gift from Dr. A.S. Kirshenbaum and Dr. D. Metcalfe, Laboratory of Allergic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA) were cultured in StemPro-34 (cat # 10640-019, GIBCO) supplemented with 1× StemPro-34 Nutrient, 2 mM L-Glutamine (cat # 03-020-1A, Biological Industries), 100 U/mL penicillin and 100 μg/mL streptomycin (cat # 03-032-1B, Biological Industries), and 100 ng/mL hSCF (cat # 300-07, Peprotech, Rocky Hill, NJ, USA).

RBL cells stably expressing N-terminally tagged hemagglutinin (HA) human MRGPRX2 (RBL-MRGPRX2) were previously described [12 (link)]. Cells were maintained as adherent cell cultures in low glucose DMEM (cat no. 01-050-1A, Biological Industries, Beit-Haemek, Israel) supplemented with 10% FBS (cat no. 12657, GIBCO, Grand Island, NY, USA), 2 mM L-Glutamine (cat no. 03-020-1A, Biological Industries), 100 μg/mL streptomycin, and 100 U/mL penicillin, 12.5 U/mL nystatin (cat no. 03-032-1B, Biological Industries), and 1 mg/mL of G418 (cat no. A1720, Sigma Aldrich, St. Louis, MO, USA), at 37 °C in a humidified incubator with 5% CO₂. LAD-2 cells (a kind gift from Dr. A.S. Kirshenbaum and Dr. D. Metcalfe, Laboratory of Allergic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA) were cultured in StemPro-34 (cat no. 10640-019, GIBCO) supplemented with 1× StemPro-34 Nutrient, 2 mM L-Glutamine (cat no. 03-020-1A, Biological Industries), 100 U/mL penicillin and 100 μg/mL streptomycin (cat no. 03-032-1B, Biological Industries), and 100 ng/mL hSCF (cat no. 300-07, Peprotech, Rocky Hill, NJ, USA).

Human metastatic melanoma cell lines 526mel and 624mel were obtained from Dr. Steve Rosenberg (National Cancer Institute, Bethesda, MD, USA). 009mel and 014mel are primary cultures derived from surgically removed metastatic melanoma specimens of patients 009 and 14, respectively. TIL14 bulk culture was established from a specimen of metastatic melanoma of patient 14 [47 (link)], obtained according to Israel Ministry of Health approval no. 3518/2004. Melanoma cultures were grown in RPMI-1640 medium (Biological Industries, Beit Ha-Emek, Israel) supplemented with 10% FBS, 100 μg/ml Pen/Strep, 2mM L-glutamine, 25mM Hepes and 1mM sodium-pyruvate (Biological Industries, Beit Ha-Emek, Israel). The A375 human metastatic melanoma cell line (ATCC, Manassas, VA, USA) was cultured in DMEM medium (Biological Industries, Beit Ha-Emek, Israel) supplemented with 10% FBS, 100 μg/ml Pen/Strep, 2mM L-glutamine, 1mM sodium-pyruvate and non-essential amino-acids (Biological Industries, Beit Ha-Emek, Israel). TIL14 bulk cultures were grown as previously described [12 (link)].

RBL cells stably expressing N-terminally tagged hemagglutinin (HA) human MRGPRX2 (RBL-MRGPRX2) were generated as previously described (20 (link)). Cells were maintained at 37°C in a humidified incubator with 5% CO₂ in adherent cell cultures in low glucose DMEM (cat no. 01-050-1A, Biological Industries, Beit-Haemek, Israel) supplemented with 10% FBS (cat no. 12657, GIBCO, Grand Island, NY), 2 mM L-Glutamine (cat no. 03-020-1A, Biological Industries), 100 μg/mL streptomycin and 100 U/mL penicillin, 12.5 U/mL nystatin (Biological Industries) and 1 mg/mL of G418 (cat no. A1720, Sigma Aldrich). LAD-2 cells (a kind gift from Dr. A.S. Kirshenbaum and Dr. D. Metcalfe, Laboratory of Allergic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD) were cultured in StemPro-34 (cat no. 10640-019, GIBCO) supplemented with 1x StemPro-34 Nutrient, 2 mM L-Glutamine (cat no. 03-020-1A, Biological Industries), 100 U/mL penicillin and 100 μg/mL streptomycin (cat no. 03-032-1B, Biological Industries), and 100 ng/mL hSCF (cat no. 300-07, Peprotech, Rocky Hill, NJ).

HEK293T and JIMT-1 cells were grown in DMEM (Gibco, MA, USA) containing 10% serum, pen-strep, HEPES, L-glutamine, non-essential amino acids, and sodium pyruvate (all from Biological Industries, Beit Haemek, Israel). NK-92 cell were grown α-mem (Gibco, MA, USA) containing 10%FBS, 10% horse serum, pen-strep, HEPES, L-glutamine, non-essential amino acids and sodium pyruvate (all from Biological Industries), Myo inositol (Sigma, MO, USA), folic acid and β-mercapto-ethanol (Gibco, MA, USA). All tissue cultured cells were kept in a humidified 5% CO₂ incubator.

Caco-2 cell line was cultured in MEM-Eagle Earle’s Salts medium (Biological Industries, Israel) supplemented with 20% (v/v) Fetal Bovine Serum (FBS) (Biological Industries, Israel), 1% (v/v) Penicilin-Sreptomycin (Biological Industries, Israel), 1% (v/v) L-Glutamine (Biological Industries, Israel), 1% (v/v) non-essential amino acids (Biological Industries, Israel). HT-29 cell line was maintained in DMEM High Glucose Medium (Biological Industries, Israel) supplemented with 1% (v/v) Penicilin-Sreptomycin (Biological Industries, Israel) and 1% (v/v) L-Glutamine (Biological Industries, Israel). PCR-based method was used to confirm mycoplasma negativity in the cell lines.

HT-29 colorectal cancer cell line and its 3D spheroid derivative (3D-HT-29) were maintained in Dulbecco’s Modified Eagle Medium (Biological Industries, Israel) supplemented with 10% Fetal Bovine Serum (Gibco, USA), 1% (v/v) Penicilin-Streptomycin (Biological Industries, Israel) and 1% (v/v) L-Glutamine (Biological Industries). HCT-116 colorectal cell line and its 3D spheroid derivative (3D-HCT-116) were maintained in Roswell Park Memorial Institute‐1640 medium (Biological Industries, Israel) with 10% Fetal Bovine Serum, 1% (v/v) Penicilin-Streptomycin and 1% (v/v) L-Glutamine. Cells were incubated at 37°C in a humidified atmosphere of 5% CO₂. The mycoplasma negativity in cell lines was routinely confirmed by a PCR-based method [40 (link)].