Nacl 0.9

Dulbecco’s modified Eagle medium (DMEM), fetal bovine serum (FBS), phosphate buffered saline (PBS) and penicillin-streptomycin (10,000 U and 10,000 mg/ml) were obtained from Invitrogen (Bleiswijk, Netherlands). The WST-1 reagent was purchased from Roche Applied Science (Vilvoorde, Belgium). NaCl 0,9% was obtained from B. Braun Medical (Diegem, Belgium), Triton X-100 from Flucka (Buchs, Switzerland), the cathepsin B inhibitor CA-074-Me from Bachem (Switzerland). Methanol, dimethyl sulfoxide (DMSO), Tris buffered saline, Tween 20, lipopolysaccharide (LPS), cytochalasin D, ATP and chloroform were purchased from Sigma-Aldrich, 2-mercaptoethanol, Laemmli Sample Buffer from Bio-Rad (Hercules, USA) and hydrofluoric acid (HF) from Merck (Darmstadt, Germany).

Dulbecco’s modified Eagle medium (DMEM) glutamax, fetal bovine serum (FBS) and penicillin-streptomycin were purchased from Life Technologies. NaCl 0.9 % was obtained from B. Braun Medical (Diegem, Belgium), Triton X-100 from Flucka (Buchs, Switzerland). The fluorescent dyes, Hoechst 33342, TMRM, Lysotracker green and TOPRO-3 were purchased from Life Technologies. All the reagents used for liposome synthesis (methanol, chloroform, L-α-phosphatidylcholine, and calcein) were purchased from Sigma-Aldrich (Milan, Italy). Ultrapure Milli-Q water (Millipore) was used throughout.

Cryopreserved vaccine doses were thawed using a thawing solution of RPMI complemented with 1% albumin (Octapharma, Lachen, Switzerland), centrifuged at RT and resuspended in NaCl 0.9% (B. Braun) before being drawn into two 1-mL syringes.

Throughout the experiments, deionized Milli-Q (Millipore, Molsheim, France) water with a resistivity of 18.2 MΩ·cm was used. All of the chemicals were of analytical grade, if not stated otherwise. Copper(II) sulfate, neocuproine (2,9-Dimethyl-1,10-phenanthroline) hydrochloric acid salt, 4-(2-Hydroxyethyl)piperazine-1-ethanesulfonic acid sodium salt (HEPES sodium salt), ascorbic acid, bovine serum albumin (BSA), and hydrochloric acid were purchased from Sigma Aldrich Ltd. (Budapest, Hungary); NaCl 0.9% was obtained from B. Braun (Budapest, Hungary); chloroform and methanol were purchased from Reanal (Budapest, Hungary). Synthetic high-purity 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC), hydrogenated soybean phosphatidylcholine (HSPC), and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)-2000] (ammonium salt) (DSPE-PEG 2000) were obtained from Avanti Polar Lipids (Alabaster, AL, USA) or Sigma Aldrich. The chemicals were used without further purification. Disposable PD-10 and PD MidiTrap G-25 desalting columns were obtained from GE Healthcare Life Sciences (Budapest, Hungary). Concentrated nitric acid (65% w/w) and H₂O₂ (30% w/w) of Suprapur quality needed for the sample preparation of cell lines for Total-Reflection X-ray Fluorescence (TXRF) measurement were supplied by Merck (Darmstadt, Germany).

The 16^th day of gestation (i.e., 5 days before delivery), pregnant females were orally administered daily with Lactobacillus rhamnosus VES001 (L.rhamnosus; 2.10⁸ CFU (Vésale Pharma, Eghezée, Belgium)/100 µL of NaCl 0,9% (B.Braun, Melsungen, Deutschland). The treatment was established after a dose-response assessment and corresponds to the most common dose (22 (link), 24 (link)). It is maintained until postnatal day (PND)3 at the rate of one daily gavage.