atrial-natriuretic-factor and Hemolysis

Recently, the N-terminal fragment of proatrial natriuretic peptide (N-terminal proANP) has been proposed as a marker of chronic congestive heart failure. In this study, we established a two-step immunoradiometric assay using monoclonal antibodies and synthetic N-terminal proANP (1-67) as a standard. It allows us to measure plasma N-terminal proANP in only 4 h without prior extraction. The detection limit of this assay was 15 pmol/L for a 100 microL sample of plasma. Within-run CVs ranged from 1.7% to 2.9% and between-run CVs ranged from 4.2% to 5.1%. The dilution curves of plasma samples showed good linearity and analytical recovery was 89-104%. The mean (+/-SD) N-terminal proANP in plasma of 33 healthy subjects was 188 (+/-71) pmol/L and 1030 (+/-411) pmol/L in 25 patients with heart failure. Our immunoradiometric assay is rapid and precise enough for routine determination of N-terminal proANP in human plasma.

Topics: Antibodies, Monoclonal; Atrial Natriuretic Factor; Biomarkers; Chromatography, Gel; Cross Reactions; Drug Stability; Edetic Acid; Epitopes; Heart Failure; Hemolysis; Humans; Immunoradiometric Assay; Protein Precursors; Reproducibility of Results; Sensitivity and Specificity

Topics: Adult; Atrial Natriuretic Factor; Cross Reactions; Freezing; Hemolysis; Humans; Immunoradiometric Assay; Indicators and Reagents; Male; Specimen Handling

Topics: Atrial Natriuretic Factor; Blood Preservation; Cold Temperature; Hemolysis; Humans; Peptide Fragments

Parameters influencing the measurement of hANP are designated and discussed on the basis of data from the literature and from our own results. Plasma hANP is dependent on the anatomical location of blood withdrawal, posture, state of hydration and salt load and medications. Interfering substances contained in plasma, management of samples and the type of assay used are of utmost importance. Direct RIA of hANP appears to be particularly sensitive to interferences. Preextraction of hANP from EDTA plasma by octadecylsilica cartridges, followed by RIA seems to be the most trustworthy method for measurement of hANP. Nevertheless, with this method too, we obtain different values for identical plasma samples if we use different antisera. Additionally, lipemia significantly lowers the ascertainable amount of endogenous, but not of exogenously added alpha-hANP. Therefore, it is reasonable to conclude that in preextracted samples too, hANP is not a homogeneous species. We observe a time-dependent increase in plasma hANP, reaching a final stable plateau value (about three times the initial values) after about 1.5 h, if EDTA blood samples are left on ice for prolonged time. Hemolysis interferes significantly with hANP determination. Readings of hANP values are reduced in dependence on plasma haemoglobin concentration. According to our experience, EDTA plasma can be stored for several months in either lyophilized form or deep frozen at -20 degrees C without the risk of reduction of hANP immunoreactivity. The data presented in this paper clearly show, that measurement of hANP is susceptible to faults in several details. Results should therefore be viewed critically.

Topics: Atrial Natriuretic Factor; Edetic Acid; Hemolysis; Humans; Indicators and Reagents; Kinetics; Radioimmunoassay; Radioligand Assay

The present paper describes a radioimmunoassay, in conjunction with an extraction procedure on octadecasilyl silica cartridges, of alpha-atrial natriuretic peptide in human plasma and its application in normal subjects under various postural conditions. The sensitivity of this assay is 1.96 pg/ml. The plasma concentrations of alpha-atrial natriuretic peptide in 18 normal male subjects ranged from 10 to 55 pg/ml. This assay was shown to be reproducible as indicated by the low within- and between-assay coefficients of variation. The plasma alpha-ANP concentration decreased during prolonged storage of the plasma at -20 degrees C or -80 degrees C and its level is also lower in hemolyzed plasma samples. The plasma alpha-ANP concentration is similar in standing and supine position but rose significantly when the subjects were tilted 20 degrees head-down.

Topics: Aldosterone; Atrial Natriuretic Factor; Hemolysis; Humans; Posture; Radioimmunoassay; Reference Values; Renin; Temperature