An Elisa for elastase α₁ -protease inhibitor complexes in human plasma and serum

Abstract

An Elisa for neutrophil elastase (ELA) in complex with a_x-protease inhibitor (PI) (α₁ -antitrypsin) was developed in microtitre plates and compared to the ELISA kit from MERCK (2-h version). Recovery of ELA-PI was good in both assays. The detection limits were 4.4μg1^-1 and 7.7μg1^-1 of the in-house and MERCK assay, respectively, while limits of quantitation were estimated to 7.7μg1^-1 (5.5-9.9 μg1^-1) and 28.9 μg1^-1 (14.6-44.3 μg1^-1) for the two assays. Furthermore, as dilution curves of normal plasma were parallel with the calibration curve in the in-house assay over a wide range of dilutions, it is feasible to assay plasma in dilutions of only 1: 6, resulting in a limit of quantitation of only 1.1 μg1^-1. The total analytical coefficient of variation for samples measured in double determinations was 10.5%-12.5% in the in-house assay and 13.9%-14.6% in the MERCK assay. One-hundred-and-eight plasma samples covering a wide range of ELA-PI concentrations were analysed in both assays. A proportional difference between the two methods was detected, the mean ratio (in-house/MERCK) with 95% confidence limits was 1.115 (1.070-1.160). The cause of the difference was probably due to different calibration of the assays. Until this problem is solved, method specific reference intervals are needed. A reference interval for the in-house method based on plasma samples from 123 healthy adults; median age 36 years (range: 19-65 years) was estimated to 16.5-48.5 μg1^-1.

Key Words:

• detection limit
• granulocytes
• neutrophils
• reference interval