Biological Material Fixation Using Non-Aqueous Methanol and Ethanol
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Solution Overview
Problem
Current methods for stabilizing biological materials, particularly for molecular biological and histological analyses, face challenges in preserving nucleic acids and proteins without crosslinking agents, which are often toxic or unsuitable for quantitative analysis, and require extreme cold storage, making them impractical and costly.
Innovation Solution
A method involving a non-aqueous composition with 10-90% methanol and additional additives, followed by transfer into a composition up to 99% ethanol, effectively stabilizes biological materials at moderate temperatures, allowing for both qualitative and quantitative analysis of nucleic acids and proteins without the need for cooling devices or toxic substances.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If formaldehyde solutions are used to conserve biological materials, then the samples are stabilised for histological examination, but the nucleic acid stabilisation is insufficient and the reagent is toxicologically harmful
Solution Approach 1:
The invention changes the chemical parameters of the fixation solution by using formalin-free compositions with alternative stabilizing agents that provide both histological preservation and nucleic acid stability without toxicity. This resolves the contradiction by modifying the chemical composition to eliminate harmful effects while maintaining stabilizing function.
Solution Approach 2:
The invention employs composite fixation solutions containing multiple components (buffers, stabilizing agents, and preservatives) that work synergistically to provide both histological examination quality and nucleic acid integrity without using toxic formaldehyde. This composite approach resolves the contradiction by combining multiple beneficial properties in one formulation.
2Reliability
If deep freezing is used to stabilise biological materials, then the integrity of nucleic acids and proteins is preserved, but cooling devices and liquid nitrogen are required making the method impractical and costly
Solution Approach 1:
The invention replaces the mechanical cooling system (freezers, liquid nitrogen) with a chemical stabilization system. The fixation solution contains agents that chemically preserve nucleic acids and proteins at room temperature, eliminating the need for complex cooling equipment while maintaining molecular integrity.
Solution Approach 2:
The invention changes the temperature parameter from requiring extreme cold storage to allowing room temperature storage by using chemical stabilizers. This parameter change resolves the contradiction by maintaining molecular integrity without the need for complex thermal control systems.
3Reliability
If formalin fixation is used, then histological structures are preserved, but the extractability of nucleic acids and proteins from tissue is reduced
Solution Approach 1:
The invention uses composite fixation solutions that contain multiple components designed to preserve histological structures while simultaneously maintaining nucleic acid and protein extractability. The composite formulation resolves the contradiction by providing dual functionality that formalin alone cannot achieve.
Solution Approach 2:
The invention changes the chemical parameters of the fixation solution to use non-crosslinking or mild crosslinking agents that preserve tissue morphology without forming dense networks that trap nucleic acids. This parameter modification resolves the contradiction by maintaining both structural integrity and molecular extractability.
4Measurement precision
If cationic detergents are used as stabilisers, then qualitative identification of nucleic acids is achieved, but nucleic acids in compact tissue pieces are not sufficiently stabilised
Solution Approach 1:
The invention combines cationic detergents with additional stabilizing agents and penetration enhancers in a composite formulation. This combination resolves the contradiction by maintaining the qualitative identification capability of detergents while adding components that ensure sufficient penetration and stabilization of nucleic acids in compact tissue.
Solution Approach 2:
The invention modifies the local quality of the stabilizing solution by incorporating agents that enhance penetration into compact tissue structures. This allows the detergent to effectively stabilize nucleic acids throughout the entire tissue sample, not just at the surface, resolving the contradiction between identification quality and tissue-wide stabilization.
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 enables the stabilization of biological materials at room temperature, maintaining the integrity of nucleic acids and proteins for extended periods, facilitating both molecular and histological analyses without the use of liquid nitrogen or deep-freezing, and allows for the extraction of biomolecules with high yield and preserved morphology.
Implementation Method 1
A method involving a non-aqueous composition with 10-90% methanol and additional additives, followed by transfer into a composition up to 99% ethanol, effectively stabilizes biological materials at moderate temperatures
Implementation Method 2
A method involving a non-aqueous composition with 10-90% methanol and additional additives, followed by transfer into a composition up to 99% ethanol, effectively stabilizes biological materials
Data Source
AI summary
The present invention relates to a method for the treatment of a biological material, comprising the stepsi) providing a biological material, andii) contacting the biological material with a first non-aqueous composition comprising:(a1) 10 to 90 vol. % methanol, and(a2) at least one additional additive, and(a3) optionally an acid.iii) transferring the biological material into a second composition (B) comprising up to 99 vol. % ethanolas well as to a new composition for preservation of a biological material usable in said method and the biological material resulting from this method, a method for the analysis of a treated biological material, various kits as well as the use of the composition in such a method.