N acetylneuraminic acid

L. antri was from the DSMZ collection (#16041) and the genomic DNA from L. sakei 23K was kindly provided by Prof. Monique Zagorec (Unité Flore Lactique et Environnement Carné, INRA, France). N-acetyl-D-mannosamine, N-acetylneuraminic acid and other sugars were from Carbosynth (Berkshire, UK). ManNAc dehydrogenase was from Kikkoman (Japan, Tokio). Other reagents were from Sigma-Aldrich (Madrid, Spain).

M-CSF–derived or GM-CSF–derived MDMs were plated at 0.5 × 10⁶ cells/well in 24-well plates. The cells were preincubated with 300 μl of purified mAbs (10 μg/ml), N-acetylneuraminic acid (10 mM SA; Carbosynth, Compton, West Berkshire, United Kingdom), lactose (50 mM), sialyllactose (GM3, 10 mM), recombinant-scaffolded HIV-1–V1V2 proteins (20 μg/ml), PNGase-treated recombinant HIV-1–V1V2 proteins (20 μg/ml), neuraminidase A–treated recombinant HIV-1–V1V2 proteins (20 μg/ml), or varying concentrations of JR-FL FD gp145 trimer for 30 min at room temperature. Following aspiration and washing, 300 μl of fresh infection medium containing purified HIV-1 was added to each monolayer, and the plate was centrifuged at 2500 rpm for 15 or 30 min at 37°C (for HIV-1 entry experiments) or for 90 min at 37°C (for HIV-1 replication experiments). For HIV-1 entry experiments, unadsorbed virus was removed, and the cells were washed 3 times with PBS to remove residual virus. The cells were subsequently lysed, and HIV-1 entry was determined by qRT-PCR or by qPCR. For HIV-1 replication, unabsorbed virus was removed, and the cells were incubated at 37°C/5% CO₂ in 1 ml of M-CSF or GM-CSF medium containing Polybrene. Cells were harvested and stained, and the percentage of HIV-1–infected cells was determined by flow cytometry.

N-acetylneuraminic acid was purchased from Biosynth-Carbosynth (Compton, UK). Chloroauric acid solution (HAuCl₄) was obtained from Sigma-Aldrich (Poole, UK). Sodium hydroxide (NaOH) was supplied from Loba Chemie (Mumbai, India). Deionized water was generated through Milli Q Millipore (Billerica, MA, USA) and used throughout the study.

ABDV was from Wako Chemicals GmbH (Neuss,
Germany). EGDMA and ARS were from Acros Organics. 4-Vinylphenylboronic
acid (FM3), ammonium hydrogen difluoride (NH₄HF₂), acetic acid, acetic anhydride, phenol, ammonium acetate, formic
acid, dimethylformamide (DMF), dry methanol (MeOH), DMSO-d₆, and methanol-d₄ (CD₃OD) were
from VWR chemicals. All solvents for high-performance liquid chromatography
(HPLC) analysis were of HPLC grade and were purchased from VWR. 2-Aminoethyl
methacrylate hydrochloride (FM2) was received from Polysciences. Amino-functionalized
macroporous silica beads (NH₂@SiO₂) with an
average particle size of 30 μm, a surface area (S) of 45 m²g^–1, an average pore diameter (D_p) of 47.5 nm, and a pore volume (V_p) of 0.81 mL/g were purchased from Fuji Silysia Chemical Ltd.
(Kozojicho, Kasugai Aichi, Japan). Monosaccharides Glc, galactose
(Gal), Fru, and TBA hydroxide (TBA–OH) 1 M in methanol were
obtained from Sigma-Aldrich. D-GlcA was received
from Fluka. N-Acetylneuraminic acid, N-glycolylneuraminic acid (Neu5Gc), GalNAc, ManNac, 2,6′-sialyllactose
sodium salt (6SL), and 2,3′-sialyllactose sodium salt (3SL)
were purchased from Carbosynth Ltd. (UK).
EGDMA was passed through
a column of activated basic alumina to remove the inhibitor and stored
at −20 °C before polymerization. N-3,5-Bis(trifluoromethyl)-phenyl-N′-4-vinylphenylurea (FM1) was synthesized as previously
reported.^{33 (link)}