Method for determining free copper

A technology for the determination of free copper in vitro, applied in the field of determination of "free" copper in serum, can solve the problems of protein absorbance peak attenuation, low concentration accuracy, high cost, etc.

Inactive Publication Date: 2016-05-04
CANOX4DRUG
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Problems solved by technology

[0004] Previously, a method for determining the amount of ceruloplasmin starting from the protein oxidase activity of a commercial standard (HumanSerumCeruloplasmin, Sigma-Aldrich) has been described, but spectral analysis showed that the absorbance peak of the protein decayed, reducing the ability to use enzymatic detection to quantify the protein. Confidence in the amount of protein necessary to estimate the value of free copper
[0005] Quantification of copper and ceruloplasmin based on enzymatic methods described in the prior art has several disadvantages such as high cost, variable purity of commercially available ceruloplasmin, general recommendation to report serum enzymes in International Units (UI), and measured Concentration accuracy is low

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example 1

[0068] For the separation of blood proteins, solid phase extraction (SPE) chromatography was developed. Ultra-high molecular weight polyethylene (UHMPE) resin (Sigma-Aldrich lot #434264-1KG) was used as the solid phase, capable of interacting with and retaining serum proteins. In order to prevent the release of protein (ceruloplasmin)-bound copper, instead of pure water, a physiological solution (0.9% NaCl) was used as the mobile phase, pumped by a peristaltic pump to maintain a constant elution flow (flow rate: 400 μl / min) . A 1-ml column is packed with 500 mg of resin ( Figure 6 ) and are moderated using two different strategies:

[0069] - 500 mg of resin, loaded into a column, conditioned with 6 ml of physiological solution.

[0070] - 500 mg of resin was suspended in about 3 ml of methanol and then used to pack the column. Then 6 ml of distilled water was eluted through the column to completely remove the methanol, followed by 6 ml of physiological solution. Then, i...

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Abstract

The present invention relates to a new method for the determination of 'free' copper concentration in serum, i.e. the portion of serum copper not structurally bound to ceruloplasmin. The present invention also refers to a method with a high degree of sensitivity and precision for the determination of free copper in serum samples of patients with Alzheimer's disease.

Description

technical field [0001] The present invention relates to a novel method for the determination of "free" copper (ie, the portion of serum copper not structurally bound to ceruloplasmin) in serum. The present invention also relates to a highly sensitive and accurate method for the determination of free copper in serum samples of patients suffering from Alzheimer's disease. Background technique [0002] Determination of serum copper is of paramount importance in a large number of diseases, for example in Alzheimer's disease (AD). Alzheimer's disease is a neurological disorder characterized by memory loss and progressive dementia. The cause of the disease is closely related to the accumulation of β-amyloid (Aβ) protein and Tau peptide in the brain. Furthermore, the ε4 allele of the apolipoprotein E (APOE) gene has been shown to increase the risk of Alzheimer's disease. New details have recently emerged about the "amyloid cascade," widely recognized as the most popular hypothes...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64G01N21/31
CPCG01N21/31G01N21/6428G01N2021/6439G01N2800/2821C07D405/12G01N31/22G01N33/52G01N33/84G01N2021/7786
Inventor 尼古拉·安东尼·克拉布弗弗朗西斯科·贝拉尔迪马尔塞洛·莱奥波尔多罗伯托·佩罗内罗莎娜·斯科尤缇
Owner CANOX4DRUG
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