Fig. 6.

The 190-HARE(N2280A) ecto-domain does not bind HA in most assays. A. Ligand blot assay. Soluble (SOL) s190-HARE(N2280A) is a single band by western blot (WB) that is smaller than membrane-bound (MEM) 190-HARE(N2280A). Both bind ¹²⁵I-HA in ligand blot (LB) assays. B. HARE pull-down assay. SA-agarose with bound b-HA was incubated with 1 μg of purified s190-HARE [WT], s190-HARE(ΔLink) [ΔL] or s190-HARE(N2280A) [N-A; two different preparations] C. ELISA-like assay. Binding of the three purified ecto-domains to HA immobilized on plate wells was quantified by A₄₀₅ as described in Materials and methods. D. Surface plasmon resonance assay. The three purified ecto-domains at 12.5 nM (lower lines) and 50 nM (upper lines) were each analyzed for binding to immobilized HA and then, after 60–90 s, for dissociation in buffer without ecto-domain as described in Materials and methods