Sodium periodate

To quantify Fcα/μR cell surface protein expression, pMG or N9 cells were cultured on poly-ornithine (Sigma-Aldrich, St. Louis, MO) coated plates in MSFM and cells were dislodged without trypsin using 2 mM EDTA/PBS. Cells were fixed for 10 min at room temperature in a phosphate buffer containing 2% paraformaldehyde/0.6% methanol (Fisher Scientific, Kent, WA), 2.5 mM sodium periodate (Fisher) and 15 mM L-lysine (Sigma-Aldrich) and then blocked with 1 mg/ml ice-cold mouse sera (BD Biosciences) plus 50 μg/ml unlabeled streptavidin (BD Biosciences) in PBS. Following a wash step, primary antibody incubations were carried out with either 50 μg/ml rat anti-Fcα/μR monoclonal antibody (clone TX-7) (27 (link)) or isotype control (BD Biosciences). For detection we used biotinylated-mouse anti-rat IgG 2b (BD Biosciences) followed by 2 μg/ml PE-Cy7-streptavidin (BD Biosciences). Staining was quantified on a FACSAria™ cytometer/sorter (BD Biosciences) and analyzed with FlowJo™ software (Tree Star). For our primary cultures, we first selected confirmed pMG by flow cytometry based on their expression of CD11b (96% of total cell population).

All synthetic manipulations were conducted in a fume hood. Catechol-functionalized Chitosan (DP-chitosan) was synthesized by coupling chitosan (low molecular weight, Aldrich cat. # 448869) and 3-(3,4-dihydroxyphenyl)propionic acid (98%, Alfa Aesar cat. #A11638) using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC•HCl, Chem Impex) as an activating agent, N-hydroxysuccinimide (e.g. NHS, Acros) as a coactivator, and 2,6-lutidine (Acros) as a base. The pH was monitored using indicator strips (VWR, intermediate range) with a resolution of 0.5 pH units. 200 proof ethanol (EtOH, molecular biology grade, denatured with 5% isopropanol) and deionized water were used as reaction media. Dowex 1×8 (50–100 mesh, beads, Chloride form) was obtained from BeanTown Chemical, and sodium periodate was obtained from Fisher. All other solvents were ACS grade and were obtained from Fisher and used as received.

(3S)-cis-3,6-Dimethyl-1,4-dioxane-2,5-dione (LA, 98%), DMAP (99%, prilled) and copper (I) bromide (CuBr, 99.999% trace metals basis) were purchased from Sigma-Aldrich. PMDETA (99+%), DMPA (99%) and sodium periodate (99%) were purchased from Acros Organics. Benzyl alcohol was purchased from J. T. Baker. α-Methoxy-ω-azido polyethylene glycol (CH₃O-PEG-N₃, manufacturer suggested MW of PEG: 750 Dalton) was purchased from Rapp Polymere. Cy5.5-N₃ was purchased from Lumiprobe Corporation. PTX (99%) was purchased from AvaChem Scientific. GEM (98+%) was purchased from Ark Pharm. Ruthenium dioxide (99.9%) was purchased from Pfaltz & Bauer. Acetone (Certified ACS grade), acetonitrile (HPLC grade), chloroform (CHCl₃, HPLC grade), dichloromethane (DCM, HPLC grade), ethyl acetate (HPLC grade), diethyl ether (HPLC grade), methanol (MeOH, HPLC grade), and N,N’-dimethylformamide (DMF; HPLC grade) were purchased from Fisher Scientific. DMF and DCM were dried by distillation over CaH₂. ACLA and ALLA were freshly prepared following our previous publications.^{49 , 70 (link)} PTX-SH and GEM-SH were prepared according to the approaches reported in literature.^{62 (link), 68}