recombinant maltose  (Novus Biologicals)

Journal: bioRxiv

Article Title: Identification of Human Transferrin Receptor as an Entry Co-receptor for Parvovirus B19 Infection of Human Erythroid Progenitor Cells

doi: 10.64898/2026.04.02.715920

Figure Lengend Snippet: Ferritin heavy chain 1 (FTH1) competes with B19V VP1u for binding to hTfR, thereby inhibiting B19V internalization and replication in CD36 + EPCs. (A&B) BLI analysis of FTH1 inhibition of GST-VP1u RBD binding to hTfR ECD . (A) Experimental scheme. hTfR ECD was immobilized on Ni-NTA BLI biosensors, then introduced into the kinetic buffer with or without FTH1, and subsequently into buffer containing GST-VP1u RBD . Maltose-binding protein (MBP) served as a control for non-specific binding. (B) Dose-dependent inhibition of VP1u binding to hTfR by FTH1. Sensograms show progressively reduced association and dissociation of hTfR ECD with GST-VP1u RBD after the introduction of FTH1 at the indicated concentrations. MBP served as a control at 4 µM. (C) Quantification of FTH1 inhibition of B19V internalization. CD36 + EPCs were incubated with FTH1 or MBP control (at the indicated concentrations) at 4°C, followed by B19V infection (MOI=3,000) at 37°C for 1 h. After treatment with trypsin, the cells were washed with PBS, and viral DNA was extracted for quantification of internalized B19V by relative viral DNA levels normalized to mitochondrial DNA. (D) FTH1 inhibits B19V infection. CD36 + EPCs were pre-incubated with FTH1 or MBP control (at the indicated concentrations) at 4°C followed by B19V infection (MOI=1,000) at 37°C for 1 day. At 1 dpi, the cells were collected for quantification of B19V DNA levels. Data are presented relative to the corresponding control-treated cell group at each concentration. Experiments were performed in triplicate, and data shown are mean ± SD, and were analyzed by one-way ANOVA with Dunnett’s test (****P < 0.0001) (E&F) Cell cycle analysis. An EdU incorporation assay was performed in CD36 + EPCs with the indicated treatments, followed by flow cytometry analysis. (E) Flow cytometry histograms. Representative histograms show the cell cycles of mock, 10 µM FTH1-, and 10 µM MBP-treated CD36 + EPCs. (F) Quantification of the percentage of the indicated cell populations in G2/M, S, or G1 phase. Data shown are the mean ± SD from three independent experiments. Statistical significance was determined by two-way ANOVA with Dunnett’s multiple comparisons test (****P < 0.0001; ns, not significant).

Article Snippet: Recombinant Maltose-binding protein (MBP) (#NBC1-18538) was purchased from Novus Biologicals (Centennial, CO).

Techniques: Binding Assay, Inhibition, Control, Incubation, Infection, Concentration Assay, Cell Cycle Assay, Flow Cytometry