Aggregate Detection via Incubation and Hydrophobic Binding
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Solution Overview
Problem
Current methods for detecting aggregate forms of aggregate-forming polypeptides in biosamples, such as those associated with neurodegenerative diseases, face challenges due to the small amount or size of the antigen, making diagnosis difficult, especially when the amount in the body is not proportional to the sample, and existing techniques like sandwich ELISA are not effective.
Innovation Solution
A method involving spiking a biosample with a monomeric or multimeric form of the aggregate-forming polypeptide or its hydrophobic deleted derivative, followed by incubation to form aggregates, and using a binder-label to detect the aggregate form, maximizing the diagnostic signal difference between patients and normals through differences in clearing systems and hydrophobic interactions.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If sandwich ELISA is used to detect aggregate-forming polypeptides, then the detection method is simple and widely applicable, but it cannot effectively detect small amounts or small sizes of antigens
Solution Approach 1:
The method performs preliminary aggregation of the target polypeptide by incubating the biosample under conditions that promote aggregate formation (e.g., adding aggregation-prone peptides, adjusting pH, temperature, or ionic strength) before the actual detection step. This preliminary action concentrates the target antigen into larger aggregate structures that are more detectable by ELISA, thereby improving measurement precision without sacrificing method simplicity
Solution Approach 2:
The invention introduces intermediary substances such as aggregation-prone peptides, chaperones, or specific buffers that mediate the formation of detectable aggregate structures from monomeric polypeptides. These intermediaries facilitate the conversion of undetectable small antigens into detectable large aggregates, enhancing detection sensitivity while maintaining the simplicity of the ELISA approach
2Loss of information
If the amount of antigen in the body is not proportional to the amount in the sample, then the diagnostic information is limited, but increasing sample volume is not always feasible
Solution Approach 1:
The method changes physical-chemical parameters such as pH, ionic strength, temperature, or adding specific chemicals to the sample to induce aggregate formation. This parameter change concentrates the target polypeptide into aggregate forms that can be detected at lower concentrations, effectively extracting more diagnostic information from the same sample volume without needing to increase the amount of biological material
3Loss of time
If aggregate forms are directly detected without pre-incubation, then the detection is faster, but the diagnostic signal difference between patients and normals is reduced
Solution Approach 1:
The method incorporates a preliminary incubation step where the biosample is pre-treated to promote aggregate formation before detection. This preliminary action creates a more pronounced difference in aggregate levels between patient and normal samples, enhancing diagnostic signal differentiation. The incubation time is optimized to be as short as possible while still achieving sufficient aggregate formation, thus minimizing time loss
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This method enhances the diagnostic signal for diseases like Alzheimer's and Parkinson's by increasing the signal generated from biosamples of patients compared to normals, allowing for more accurate differentiation and diagnosis.
Implementation Method 1
the method maximizing a differentiation in the diagnostic signal between a patient and a normal subject using a difference in the clearing system suppressing the formation of an aggregate form of a polypeptide or a difference in hydrophobic interaction
Data Source
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AI summary
The present invention relates to a method for detecting an aggregate form of an aggregate-forming polypeptide in a biosample, comprising the steps of: (a) spiking, in a biosample to be analyzed, (i) a monomeric or multimeric form of an aggregate-forming polypeptide, (ii) a hydrophobic deleted derivative of the aggregate-forming polypeptide, or (iii) a monomeric or multimeric form of the aggregate-forming polypeptide and a hydrophobic deleted derivative of the aggregate-forming polypeptide; (b) additionally forming the aggregate form of the aggregate-forming polypeptide by incubating the product of step (a); (c) making the product of step (b) come into contact with a binder-label in which a signal-generating label is coupled to a binder binding to the aggregate form of the aggregate-forming polypeptide; and (d) detecting a signal to be generated from the binder-label bound to the aggregate form of the aggregate-forming polypeptide.