<p>a) Recombinant SP-D (rSP-D), serum, and EDTA plasma were assayed in twofold dilutions of [Ca<sup>2+</sup>] or [EDTA] (concentrations are displayed in mM). For the inclusion of calcium with recombinant SP-D, Hank's Balanced Salt Solution (HBSS) was used as a buffer instead of PBS. Recombinant SP-D was assayed at 5 ng/mL while serum and plasma were diluted twofold. Each line represents an independent experiment. b) ELISAs were performed with detection reagents diluted in HBSS without calcium chloride (-CaCl<sup>2</sup>) or with 5 mM calcium chloride (+CaCl<sup>2</sup>). Wash buffer also either lacked or included calcium chloride. Recombinant SP-D was assayed at 10 ng/mL and Heparin plasma from four subjects was pooled, and sample values were extrapolated from a single standard curve under the conditions lacking calcium. n = 3 samples per group.</p
