Comparative analysis of obesity-related cardiometabolic and renal biomarkers in human plasma and serum.

The search for biomarkers associated with obesity-related diseases is ongoing, but it is not clear whether plasma and serum can be used interchangeably in this process. Here we used high-throughput screening to analyze 358 proteins and 76 lipids, selected because of their relevance to obesity-associated diseases, in plasma and serum from age- and sex-matched lean and obese humans. Most of the proteins/lipids had similar concentrations in plasma and serum, but a subset showed significant differences. Notably, a key marker of cardiovascular disease PAI-1 showed a difference in concentration between the obese and lean groups only in plasma. Furthermore, some biomarkers showed poor correlations between plasma and serum, including PCSK9, an important regulator of cholesterol homeostasis. Collectively, our results show that the choice of biofluid may impact study outcome when screening for obesity-related biomarkers and we identify several markers where this will be the case.

The international standard for plasminogen activator inhibitor-1 (PAI-1) activity.

Since the finding that plasminogen activator inhibitor-1 (PAI-1) may influence the initiation and progression of acute myocardial infarction, the assay of PAI-1 in plasma using a variety of commercial kits has become commonplace. The need for a standard to define the activity of PAI-1 prompted an international collaborative study (ICS) to calibrate the functional potency of a lyophilised plasma PAI-1 preparation (92/654). Since PAI-1 inhibits the 2 major plasminogen activators, tissue-type plasminogen activator (t-PA) and urinary-type plasminogen activator (u-PA) in an equimolar manner it was important to establish the potency of the PAI-1 inhibitor in terms of both t-PA and u-PA neutralisation. While the ICS indicated a wide spread of data between the laboratories the mean value of 27.5 t-PA neutralisation units and 7.0 u-PA neutralisation units was confirmed by numerous assays at NIBSC using a tedious but technically reliable titration assay procedure. The plasma PAI-1 proposed standard (92/654) was stable at 20 degrees C for 20 months. The Fibrinolysis Subcommittee of the Scientific and Standardization Committee (SSC) of the International Society on Thrombosis and Haemostasis (ISTH), (meeting in Leuven. Belgium in September 1994) has recommended that the plasma PAI-1 (92/654) should be accepted as the International Standard for PAI-1 and should define a unitage in terms of both t-PA and u-PA neutralisation. Subsequently the Expert Committee on Biological Standardization of the World Health Organization (ECBS-WHO) meeting in Geneva, Switzerland in October 1995 approved plasma PAI-1 (92/654) as the International Standard.

Multicentre evaluation of commercial kit methods: plasminogen activator inhibitor activity.

In order to study the analytical performance of different commercial kits for determination of plasminogen activator inhibitor (PAI) activity we distributed eight selected split samples to 11 European laboratories experienced with haemostasis testing. Three different laboratories were involved in the production of data from each of the commercial kits tested. A considerable variation of PAI activity results reported from the laboratories testing the same commercial kits was observed. The range of reported results could in individual samples exceed the median value indicating an interlaboratory variation of more than 100%. When we harmonized the results reported from different kits in different laboratories by means of an international standard from National Institute for Biological Standards and Control (NIBSC) we still observed that the results produced by some kits deviated systematically from results produced by other kits. Also, the harmonized results were used to estimate the overall coefficient of variation (CV) of PAI activity determined in various laboratories by different kits. We observed an inverse correlation between the PAI activity level and the CV with a CV of about 100% for low PAI activity levels and a CV of about 16% for high PAI activity levels. The high imprecision of the kits in the low concentration range of PAI activity indicates that unspecific factors in plasma may interfere with determination of active PAI. This was confirmed by the evaluation of the results from one of the plasma samples, which was PAI-1 depleted. The laboratories involved in the testing reported for this sample a mean value of 6.1 IU/ml.(ABSTRACT TRUNCATED AT 250 WORDS)

Multicenter evaluation of commercially available methods for the immunological determination of plasminogen activator inhibitor-1 (PAI-1).

In order to evaluate the comparability of data obtained with various available kits for the immunological determination of PAI-1 antigen in plasma and in order to investigate the underlying cause of observed differences, e.g. problems of specificity or of proper calibration of the provided standard, a multicenter study was organised in the framework of the Subcommittee of Fibrinolysis of the Scientific and Standardization Committee. Eight different plasma samples were distributed among 16 laboratories: a pooled normal plasma, NIBSC 87/512, PAI-1 antigen depleted plasma, PAI-1 depleted plasma supplemented with 59 ng/ml active PAI-1 and four different individual plasma samples. A considerable variation in absolute values is observed between the various kits, e.g. in pooled normal plasma a value is found ranging between 7.4 and 28 ng/ml. Harmonization of all data relative to the PAI-1-depleted plasma supplemented with an exact amount of active PAI-1 (59 ng/ml), followed by a statistical analysis using a two way analysis of variance, revealed that 6 out of 7 kits yielded values that were not significantly different with coefficients of variation around 30%. Correlations between the values obtained with these kits yielded slopes between 0.75 and 1.44 with correlation coefficients between 0.973 and 0.999. Values obtained with one kit appeared to be significantly different (even after harmonization) from the other kits (p < 0.001 to p < 0.05). Comparison of PAI-1 antigen with the PAI activity values in the analysed samples suggests that one kit may deal with a problem of a difference in reactivity between active and latent PAI-1.(ABSTRACT TRUNCATED AT 250 WORDS)

A specific immunologic assay for functional plasminogen activator inhibitor 1 in plasma--standardized measurements of the inhibitor and related parameters in patients with venous thromboembolic disease.

A new assay for functional plasminogen activator inhibitor 1 (PAI-1) in plasma was developed. The assay is based on the quantitative conversion of PAI-1 to urokinase-type plasminogen activator (u-PA)-PAI-1 complex the concentration of which is then determined by an ELISA employing monoclonal anti-PAI-1 as catching antibody and monoclonal anti-u-PA as detecting antibody. The assay exhibits high sensitivity, specificity, accuracy, and precision. The level of functional PAI-1, tissue-type plasminogen activator (t-PA) activity and t-PA-PAI-1 complex was measured in normal subjects and in patients with venous thromboembolism in a silent phase. Blood collection procedures and calibration of the respective assays were rigorously standardized. It was found that the patients had a decreased fibrinolytic capacity. This could be ascribed to high plasma levels of PAI-1. The release of t-PA during venous occlusion of an arm for 10 min expressed as the increase in t-PA + t-PA-PAI-1 complex exhibited great variation and no significant difference could be demonstrated between the patients with a thrombotic tendency and the normal subjects.