Secretory IgA enzyme immunoassay

Abstract

An enzyme-linked immunosorbent assay (ELISA) is applied for quantitation of secretory IgA (SIgA) in nasopharyngeal secretions (NPS) and saliva. In principle, both the alpha and secretory component determinants of SIgA are utilized in a two-site sandwich technique. Dilutions of a SIgA solution prepared from colostrum (Col-SIgA) are used for standards. A continuous SIgA standard curve, describing the relationship between the absorbance of the coloured end product and the corresponding Col-SIgA dilution, is computed by means of a fitting procedure based on a model. The variation (SD) of the absorbance was found to be proportional to the absorbance level. Knowing the SD of the absorbance, the total variation of the SIgA estimates could be calculated experimentally by means of Monte Carlo methods. Based on these results, the analytical range was chosen from 2000 micrograms/l to 50 micrograms/l of SIgA. Inhibition experiments revealed that plasma IgA could be added to the NPS in equal amounts to the SIgA content without it interfering with the SIgA measurements. Percentage recovery (median) of known amounts of SIgA added to NPS was 96%. In NPS from children suffering from recurrent upper respiratory tract infections the median SIgA level was 1.44 g/l (interquartile range 0.61-2.43 g/l); in saliva from healthy children the median SIgA content was 77 mg/l (interquartile range 56-182 mg/l).

New method to assess dilution of secretions for immunological and microbiological assays

“…Accurate quantitation of antibodies in secretions is a problem when studying mucosal immunity. Absolute results have been obtained with rather cumbersome methods, e.g., aspirating nasopharyngeal secretions under general anesthesia during otolaryngological operations followed by freezing, weighing, and rinsing out the suction device with a known amount of diluent (14). The secretion can be absorbed onto filter paper (8,10) or dextran (12), with the increase in weight representing the amount of secretion obtained.…”