Effect of Time to Detection on the Measured Concentrations of Blood Proteins Associated with Alzheimer's Disease.

Background: For assays using immunomagnetic reduction, a reagent composed of antibody-functionalized magnetic nanoparticles is dispersed in phosphate-buffered saline solution. The real-time signals of alternating-current (ac) magnetic susceptibility, χac, of the reagent are subsequently recorded after mixing the reagent with a biofluid sample. After mixing the reagent and sample, the reduction in χac of the mixture is calculated and used to quantify the concentration of the target biomarker in the sample. The reduction does not occur immediately but rather occurs at some time after mixing. This observation implies that the time elapsed before recording the real-time signals of χac of a reagent-sample mixture needs to be investigated to ensure that the signals are fully recorded. In this work, the effect of time to detection on the measured concentrations of proteins in human plasma after mixing the reagent and sample is examined. Methods: The proteins analyzed are related to Alzheimer's disease: amyloid ß 1-40, amyloid ß 1-42, and Tau protein. The investigated times to detection after the mixing the reagent and sample are 0, 20, 30, 40, and 120 min. Results: The results show that the recording of real-time signals of χac should be conducted within 20 min after mixing the reagent and sample.

Investigation of the Number of Tests Required for Assaying Plasma Biomarkers Associated with Alzheimer's Disease Using Immunomagnetic Reduction.

INTRODUCTION: Concentrations of plasma biomarkers associated with Alzheimer's disease have been reported to be as low as several tens of picograms/milliliter (pg/ml). However, in assays measuring these biomarkers, it is likely that repeated measurements are necessary to obtain reliable values. METHODS: We performed assays as a single test or as duplicate, quadruplicate, fivefold and tenfold repeated tests, on samples spiked with different concentrations of amyloid ß 1-40 (Aß1-40; 1-1000 pg/ml), Aß1-42 (1-30,000 pg/ml) and total Tau protein (T-Tau; 0.1-1000 pg/ml), with the aim to to calculate the coefficients of variation (CVs). RESULTS: The results demonstrated common changes in the CVs with changes in the number of tests for a given sample: the CVs decreased with increases in the number of tests from one to ten. All CV values were distributed within the range of 0.35 to 15.5%; as such, the CV values were all lower than the acceptable value of 20%. CONCLUSION: Based on this study, a single assay of Aß1-40, Aß1-42 and T-Tau, respectively, provides reliable results in terms of the measurement of that plasma biomarker.

Effect of Times to Blood Processing on the Stability of Blood Proteins Associated with Dementia.

BACKGROUND: The stability of proteins in the collecting tubes after blood draw is critical to the measured concentrations of the proteins. Although the guidelines issued by the Clinical and Laboratory Standards Institute (CLSI) suggest centrifugation should take place within 2 h of drawing blood, it is very difficult to follow these guidelines in hospitals or clinics. It is necessary to study the effect of times to blood processing on the stability of the proteins of interest. METHODS: In this work, the plasma proteins of interest were those relevant to dementia, such as amyloid ß 1-40 (Aß1-40), Aß1-42, Tau protein (Tau), and α-synuclein. The times to blood processing after blood draw ranged from 0.5 to 8 h. The storage temperatures of blood were room temperature (approx. 25°C) and 30°C. After storage, blood samples were centrifuged at room temperature to obtain plasma samples. Ultrasensitive immunomagnetic reduction was applied to assay these proteins in the plasma. RESULTS: The levels of plasma Aß1-40, Tau, and α-synuclein did not significantly change until 8 h after blood draw when stored at room temperature. Plasma Aß1-42 levels did not change significantly after 8 h of storage at room temperature before blood processing. Higher storage temperatures, such as 30°C, for blood samples accelerated the significant variations in the measured concentrations of Aß1-40, Tau, and α-synuclein in plasma. CONCLUSION: According to these results, for clinical practice, it is suggested that blood samples be stored at room temperature for no longer than 4.5 h after blood draw until centrifugation for the assay of dementia biomarkers in plasma.

A Novel Detection Platform for Shrimp White Spot Syndrome Virus Using an ICP11-Dependent Immunomagnetic Reduction (IMR) Assay.

Shrimp white spot disease (WSD), which is caused by white spot syndrome virus (WSSV), is one of the world's most serious shrimp diseases. Our objective in this study was to use an immunomagnetic reduction (IMR) assay to develop a highly sensitive, automatic WSSV detection platform targeted against ICP11 (the most highly expressed WSSV protein). After characterizing the magnetic reagents (Fe3O4 magnetic nanoparticles coated with anti ICP11), the detection limit for ICP11 protein using IMR was approximately 2 x 10(-3) ng/ml, and the linear dynamic range of the assay was 0.1~1 x 10(6) ng/ml. In assays of ICP11 protein in pleopod protein lysates from healthy and WSSV-infected shrimp, IMR signals were successfully detected from shrimp with low WSSV genome copy numbers. We concluded that this IMR assay targeting ICP11 has potential for detecting the WSSV.

Development of antibody functionalized magnetic nanoparticles for the immunoassay of carcinoembryonic antigen: a feasibility study for clinical use.

BACKGROUND: Magnetic nanoparticles functionalized antibodies are used for in-vitro assays on bio-markers. This work demonstrates the synthesis of high-quality magnetic nanoparticles coated with antibodies against carcinoembryonic antigen (CEA). Various characterizations, such as particle size, particle suspension, bio-activity and the stability of bio-magnetic nanoparticles suspended in liquid, are studied. The properties for the assay of CEA molecules in serum are also studied. The assay method used is so-called immunomagnetic reduction. RESULTS: The results show that the effects of common materials in serum that interfere with detected signals are not significant. The low-detection limit is 0.21 ng/ml, which is well below the clinical threshold of 2.5 ng/ml. CONCLUSIONS: The dynamic range for the assay of CEA molecules in serum is 500 ng/ml. By assaying serum CEA molecules from 24 normal controls and 30 colorectal-cancer patients, the threshold for the serum-CEA concentration to diagnose colorectal cancer is 4.05 ng/ml, which results in a clinical sensitivity of 0.90 and specificity of 0.87.