Rpmi 1640 medium

The endometrial epithelial cells (EECs) were preserved in our laboratory. The extensively employed EC cells, HEC-1A and Ishikawa, were obtained from the Meisen Chinese Tissue Culture Collections (Zhejiang, China) and preserved in our laboratory. McCoy’s 5A medium (Shanghai BasalMedia Technologies Co., Ltd., Shanghai, China) mixed with 10% fetal bovine serum (FBS) was used to cultivate HEC-1A cells. RPMI 1640 medium (Shanghai BasalMedia Technologies Co., Ltd.) containing 10% FBS was used to cultivate Ishikawa cells and EECs. The cell culture medium was supplemented with streptomycin at a concentration of 100 g/mL and penicillin at a concentration of 100 U/mL (Beyotime, Beijing, China). Cells were grown at 37°C in a 5% CO₂ environment.

CHO-K1, CHO-CLDN18.1-GFP (CLDN18.1 and GFP co-expressing) and CHO-CLDN18.2-GFP (CLDN18.2 and GFP co-expressing) cells were cultured in CD02 medium (Quacell) at 37°C in a 5% CO₂ incubator with constant agitation at 120 rpm. ADCC bioassay effector cells were maintained in 90% RPMI1640 medium (BasalMedia) with L-glutamine and 10% fetal bovine serum (FBS, ExcellBio), 100 µg/mL hygromycin (Invivogen), 250 µg/mL G-418 sulfate solution (Invivogen), sodium pyruvate (1 mM, Gibco) and MEM non-essential amino acids (0.1 mM, Gibco), at 37°C in a humid incubator with 5% CO₂. SNU-620 (human gastric cancer cell line with endogenous CLDN18.2 expression) and NUGC4-CLDN18.2 (human gastric cancer cell line with exogenous CLDN18.2 expression) cells were grown in RPMI1640 medium supplemented with 10% FBS. MIA PaCa-2-CLDN18.2 cells (human pancreatic cancer cell line with exogenous CLDN18.2 expression) were grown in DMEM high glucose medium supplemented with 10% FBS.

Human ovarian cancer cell lines SKOV3 and A2780 were purchased from the American Type Culture Collection (ATCC, United States). Ovarian cancer cell lines were cultured in McCoys’5 A or RPMI-1640 medium (BasalMedia, L110KJ), supplemented with 10% fetal bovine serum (Everyday Green, 13,011–8,611). All cells were maintained at 37°C in a 5% CO₂ incubator. PTX (MedChemExpress, United States) and Cur (Sigma-Aldrich, United States) were dissolved in DMSO (Sigma-Aldrich, D2660) and diluted to appropriate concentrations in culture medium.

HEK293 (ATCC, CRL-3216) cells, Huh7 (Institute of Basic Medical Sciences CAMS, 3111C0001CCC000679) cells and Vero (ATCC, CCL-81) cells were cultured at 37 °C in Dulbecco’s Modified Eagle medium (DMEM, Hyclone SH30243.01) supplemented with 10% fetal bovine serum (FBS). CHO-K1 (ATCC, CCL-61) cells were cultured at 37 °C in Dynamis Medium (GIBCO A2661501). Raji (ATCC, CCL-86) cells, THP-1 (ATCC, TIB-202) cells, K562 (ATCC, CCL-243) cells, and SC (ATCC, CRL-9855) cells were cultured at 37 °C in RPMI 1640 Medium (BasalMedia L220KJ) with 10% FBS. SARS-CoV-2 pseudovirus was prepared and provided by the Institute for Biological Product Control, National Institutes for Food and Drug Control (NIFDC)⁴² .

Vaccinated mice were sacrificed, and the mesenteric lymph nodes (MLNs) and spleens were harvested. By pulverizing the tissue through a 40 μm cell strainer, lymph nodes and spleens were produced as single-cell suspensions. ACK buffer (HyClone) was used to lyse red blood cells, which were then centrifuged and suspended in RPMI 1640 medium (Basal Media) supplemented with 10% FBS (BI, Israel), and 1% streptomycin/penicillin. Single-cell suspensions were directly stained for flow cytometric analysis. Before intracellular cytokine staining, cells were stimulated in BFA and cell stimulation cocktail (Invitrogen) for 6 h. Next, the cells were stained for extracellular markers, fixed and permeabilized with Intracellular Fixation/Permeabilization Buffer (Invitrogen). Rat anti-mouse CD3 (clone 17A2), CD4 (clone GK1.5), IFN-γ (clone XMG1.2), IL-4 (clone 11B11), and IL-17 A (clone TC11-18H10.1) antibodies were purchased from BioLegend (San Diego, CA, USA). Flow cytometry was performed on a FACS Celesta flow cytometer (BD Biosciences, USA).

The human GC cell lines SGC7901, BGC823 and HGC-27 cells were obtained from the Army Medical University (Chongqing, China). SGC7901 and BGC823 cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) with 10% foetal bovine serum (FBS) (Lonsera, Uruguay). HGC-27 cells were cultured in RPMI 1640 medium (Basal Media, Shanghai, China) supplemented with 10% FBS. All cells were cultured at 37 °C in a humidified incubator with 5% CO₂ (standard incubation conditions).