Recently we reported that the levels of type II phospholipase A.I (sPLA2~) in human urémie plasma were 100-fold greater than control when determined by ELISA but only 8-fold greater when determined by measuring enzymic activity. To show that ELISA quantifies protein levels more accurately, sPLAi from urémie plasma was enriched by immunoabsorption. washed, and extracted with 0.36 N H2S04/1.6 M NaC). The extract containing greater than 82% of the initial immunodetectable sPLA? showed a single immunoreactive band by Western blot analysis with a molecular weight of 16 kDa which comigrated with purified sPLA. ELISA was used to assess the levels of sPLA% in patients of varying glomerular filtration rates (GFRs). Plasma levels from hemodialysis patients contained 1025.3ng/ml (median, 52-3220ng/ml, nll) while plasma from patients from varying degrees of renal failure (GFRs ranging from 5-93ng/ml) contained 87.9ng/ml (median, 19.4-150.6ng/rnl, n-26) verses control levels of 9.2ng/ml (median, 4.6-17.5ng/ml, n-13). These data demonstrate that the sandwich ELISA detects 14 kDa type II sPLA as the predominant antigen in urémie plasma, and suggests that factors in plasma can dramatically underestimate sPLAi levels measured using enzymatic methods. Plasma levels of sPLA? are increased in patients with renal failure. Significant elevations are most apparent in patients on maintenance hemodialysis.
|Original language||English (US)|
|State||Published - Dec 1 1996|
ASJC Scopus subject areas
- Molecular Biology