Dowex 50wx8 200

LPS was analyzed with matrix-assisted laser desorption/ionization time-of flight (MALDI-TOF) mass spectrometry (MS) using a Waters SYNAPT G2-Si HDMS instrument (Waters Corporation, Milford, MA, USA) equipped with a 1 kHz Nd:YAG laser system. Acquisition of the data was performed using MassLynx software version 4.1 SCN916 (Waters Corporation, Wilmslow, United Kingdom). Mass spectra were assigned with a multi-point external calibration using red phosphorous (Sigma-Aldrich, St. Louis, MO, USA) and recorded in the negative ion mode. An LPS sample (at a concentration of 10 µg/µL) was suspended in a water/methanol (1:1, v/v) solution containing 5 mM EDTA and then dissolved by ultrasonication. After desalting with some grains of cation exchange beads (Dowex 50WX8-200; Sigma-Aldrich, St. Louis, MO, USA), one microliter of the sample was transferred onto a well plate covered with a thin matrix film and allowed to dry at room temperature. The matrix solution was prepared from 2’,4’,6’-trihydroxyacetophenone (THAP) (200 mg/mL in methanol) mixed with nitrocellulose (15 mg/mL) suspended in 2-propanol/acetone (1:1, v/v) in proportion of 4:1 (v/v), according to the published method [71 (link)].

The LPS and oligosaccharide samples were analyzed with matrix-assisted laser desorption/ionization time-of flight (MALDI-TOF) mass spectrometry (MS) using a Waters SYNAPT G2-Si HDMS instrument (Waters Corporation, Milford, MA, USA) equipped with a 1 kHz Nd:YAG laser system. Acquisition of the data was performed using MassLynx software version 4.1 SCN916 (Waters Corporation, Wilmslow, UK). Mass spectra were assigned with a multi-point external calibration using red phosphorous (Sigma) and recorded in the negative ion mode. Phenol-soluble LPS and oligosaccharide samples (both at a concentration of 15 µg/µL) were suspended in a water/methanol (1:1, v/v) solution (containing 2 mM EDTA for the LPS sample) and dissolved by ultrasonication. After desalting with the use of cation exchange beads (Dowex 50WX8-200; Sigma), one microliter of each sample was transferred onto a well plate covered with a thin matrix film and allowed to dry at room temperature. The matrix solution was prepared from 2′,4′,6′-trihydroxyacetophenone (THAP) (200 mg/mL in methanol) mixed with nitrocellulose (15 mg/mL) suspended in 2-propanol/acetone (1:1, v/v) in a proportion of 4:1 (v/v), according to the published method [65 (link),66 (link)].

Sodium Hyaluronate (MW = 100 kDa) was provided by Shandong Freda Biochem Co., Ltd. (Shandong, China). Dowex^® 50WX8-200, tetrabutylammonium hydroxide (TBA-OH), benzyl bromide, TritonX-100 and hyaluronidase were purchased from Sigma-Aldrich (St. Louis, MO, USA). Anhydrous dimethyl sulfoxide (DMSO), ethyl acetate, and Hexafluoroisopropanol (HFIP) were obtained from Aladdin Reagent Company (Shanghai, China). Doxorubicin hydrochloride (Dox•HCl, >99%) and cell counting kit-8 (CCK-8) were brought from Dalian Meilun Biology Technology Co., Ltd. (Dalian, China). Fluorescein diacetate (FDA), propidium iodide (PI), 4′,6-diamidino-2-phenylindole (DAPI), and TRITC-phalloidin were purchased from Beijing Solarbio Science & Technology Co., Ltd. (Beijing, China). DMEM medium, RPMI-1640 and fetal bovine serum were provided by Gibco (Cleveland, TN, USA). Penicillin-streptomycin and trypsin were purchased from HyClone (Logan, UT, USA). All other chemicals were used without further purification.