Abstract
BackgroundMonomers of the collectin surfactant associated protein-A (SP-A) are arranged in trimers and higher oligomers. The state of oligomerization differs between individuals and likely affects SP-A's functional properties. SP-A can form aggregates together with other SP-A molecules. Here we report and assess a test system for the aggregate forming properties of SP-A in serum and broncho-alveolar lavage samples.MethodsAnti-SP-A antibodies fixed to latex beads bound SP-A at its N-terminal end and allowed the interaction with other SP-A molecules in a given sample by their C-terminal carbohydrate recognition domain (CRD) to agglutinate the beads to aggregates, which were quantified by light microscopy.ResultsSP-A aggregation was dependent on its concentration, the presence of calcium, and was dose-dependently inhibited by mannose. Unaffected by the presence of SP-D no aggregation was observed in absence of SP-A. The more complex the oligomeric structure of SP-A present in a particular sample, the better was its capability to induce aggregation at a given total concentration of SP-A. SP-A in serum agglutinated independently of the pulmonary disease; in contrast SP-A in lung lavage fluid was clearly inferior in patients with chronic bronchitis and particularly with cystic fibrosis compared to controls.ConclusionsThe functional status of SP-A with respect to its aggregating properties in serum and lavage samples can be easily assessed. SP-A in lung lavage fluid in patients with severe neutrophilic bronchitis was inferior.
Highlights
Monomers of the collectin surfactant associated protein-A (SP-A) are arranged in trimers and higher oligomers
Characteristics of the assay to assess SP-A selfagglutination First the specificity for SP-A, and not for SP-D, and the dependency of the assay on the presence of a complete binding chain consisting of the beads coupled with the anti-goat antibody, and the anti-human-SP-A antibody was shown
Results demonstrated that an increasing concentration of mannose as well as the absence of calcium inhibited the formation of large agglutinates (Figure 2 and 3)
Summary
Monomers of the collectin surfactant associated protein-A (SP-A) are arranged in trimers and higher oligomers. We report and assess a test system for the aggregate forming properties of SP-A in serum and broncho-alveolar lavage samples. SP-A belongs like SP-D to a family of innate host defence proteins termed collectins because of the presence of a collagenous and a lectin-like domain [1]. The SP-A monomer consists of 228 amino acids and has a molecular weight of 26-32 kDa. Four domains can be differentiated in the primary structure [2]. The N-terminus is followed by the collagen-like region which is linked to the globular head by the neck. SP-A accumulates in vivo predominantly as octadecamers composed of six trimeric subunits forming a flower bouquet-like structure [3,4].
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