Assessment of immunoprecipitation with subsequent immunoassays for the blood-based diagnosis of Alzheimer's disease.

The Aß42/40 ratio and the concentration of phosphorylated Tau181 in blood plasma represent attractive biomarkers for Alzheimer's disease. As a means for reducing potential matrix effects, which may interfere with plasma immunoassays, we have previously developed a pre-analytical sample workup by semi-automated immunoprecipitation. Here we test the compatibility of pre-analytical immunoprecipitations with automated Aß1-40, Aß1-42 and phosphorylated Tau181 immunoassays on the Lumipulse platform and compare the diagnostic performance of the respective immunoprecipitation immunoassay approaches with direct plasma measurements. 71 participants were dichotomized according to their Aß42/40 ratios in cerebrospinal fluid into the diagnostic groups amyloid-positive (n = 32) and amyloid-negative (n = 39). The plasma Aß1-42/1-40 ratio and phosphorylated Tau181 levels were determined on the Lumipulse G600II platform (Fujirebio) by direct measurements in EDTA-plasma or after Aß- or Tau-immunoprecipitation, respectively. Pre-analytical immunoprecipitation of Aß turned out to be compatible with the Lumipulse Aß assays and resulted in a numerical, yet statistically not significant increase in the area under the ROC curve for plasma Aß1-42/1-40. Additionally, we observed a significant increase in the standardised effect size (Cohen's D). Pre-analytical immunoprecipitation of Tau resulted in increased differences between the diagnostic groups in terms of median and mean phosphorylated Tau 181 levels. Furthermore, we observed a greater Cohen's d (p < 0.001) and a larger area under the ROC curve (p = 0.038) after Tau-IP. Our preliminary findings in a small, preselected sample indicate that pre-analytical immunoprecipitation may have the potential to improve the diagnostic performance of plasma biomarker immunoassays for Aß1-42/1-40 and phosphorylated Tau181 to predict brain amyloid deposition.

A novel approach reveals that HLA class 1 single antigen bead-signatures provide a means of high-accuracy pre-transplant risk assessment of acute cellular rejection in renal transplantation.

BACKGROUND: Acute cellular rejection (ACR) is associated with complications after kidney transplantation, such as graft dysfunction and graft loss. Early risk assessment is therefore critical for the improvement of transplantation outcomes. In this work, we retrospectively analyzed a pre-transplant HLA antigen bead assay data set that was acquired by the e:KID consortium as part of a systems medicine approach. RESULTS: The data set included single antigen bead (SAB) reactivity profiles of 52 low-risk graft recipients (negative complement dependent cytotoxicity crossmatch, PRA < 30%) who showed detectable pre-transplant anti-HLA 1 antibodies. To assess whether the reactivity profiles provide a means for ACR risk assessment, we established a novel approach which differs from standard approaches in two aspects: the use of quantitative continuous data and the use of a multiparameter classification method. Remarkably, it achieved significant prediction of the 38 graft recipients who experienced ACR with a balanced accuracy of 82.7% (sensitivity = 76.5%, specificity = 88.9%). CONCLUSIONS: The resultant classifier achieved one of the highest prediction accuracies in the literature for pre-transplant risk assessment of ACR. Importantly, it can facilitate risk assessment in non-sensitized patients who lack donor-specific antibodies. As the classifier is based on continuous data and includes weak signals, our results emphasize that not only strong but also weak binding interactions of antibodies and HLA 1 antigens contain predictive information. TRIAL REGISTRATION: ClinicalTrials.gov NCT00724022 . Retrospectively registered July 2008.

A two-step immunoassay for the simultaneous assessment of Aß38, Aß40 and Aß42 in human blood plasma supports the Aß42/Aß40 ratio as a promising biomarker candidate of Alzheimer's disease.

BACKGROUND: The quantification of amyloid-beta (Aß) peptides in blood plasma as potential biomarkers of Alzheimer's disease (AD) is hampered by very low Aß concentrations and the presence of matrix components that may interfere with the measurements. METHODS: We developed a two-step immunoassay for the simultaneous measurement of the relative levels of Aß38, Aß40 and Aß42 in human EDTA plasma. The assay was employed for the study of 23 patients with dementia of the Alzheimer's type (AD-D) and 17 patients with dementia due to other reasons (OD). We examined relationships with the clinical diagnosis, cerebral Aß load as quantified by amyloid-positron emission tomography, apolipoprotein E genotype, Aß levels and Tau protein in cerebrospinal fluid. RESULTS: Preconcentration of plasma Aß peptides by immunoprecipitation substantially facilitated their immunological measurements. The Aß42/Aß40 and Aß42/Aß38 ratios were statistically significantly lower in the AD-D patients than in the OD group. The areas under the receiver operating characteristic curves reached 0.87 for the Aß42/Aß40 ratio and 0.80 for the Aß42/Aß38 ratio. CONCLUSIONS: The measurement of plasma Aß peptides with an immunological assay can be improved by preconcentration via immunoprecipitation with an antibody against the Aß amino-terminus and elution of the captured peptides by heating in a mild detergent-containing buffer. Our findings support the Aß42/Aß40 ratio in blood plasma as a promising AD biomarker candidate which correlates significantly with the validated core biomarkers of AD. Further studies will be needed for technical advancement of the assay and validation of the biomarker findings.