Whole Cell Isolation from Frozen Tissue via CPA-Free Flash Freezing
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Solution Overview
Problem
Current methods for isolating whole cells from frozen tissue samples often disrupt cellular integrity due to the use of cryoprotective agents or slow freezing techniques, leading to inaccurate biochemical analysis and difficulty in studying cells in their native environment, especially for cancer research where protein expression patterns are crucial.
Innovation Solution
A method involving cutting frozen tissue samples into pieces using a tissue sample cutter, forming an extraction mixture, and using a separation funnel to isolate whole cells without cryoprotective agents, allowing for rapid and efficient isolation of cells directly from stroma material, maintaining cellular structures for subsequent biochemical analysis.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional cryopreservation methods using cryoprotective agents are used, then cellular integrity is preserved during freezing, but cellular integrity and biochemical accuracy deteriorate due to cytotoxicity and deleterious side effects
Solution Approach 1:
The invention removes cryoprotective agents from the freezing process entirely, extracting the harmful element while maintaining the beneficial outcome of cellular integrity preservation through CPA-free flash freezing protocols
Solution Approach 2:
The invention changes the freezing parameters by implementing ultra-rapid cooling rates that prevent ice crystal formation without requiring cryoprotective agents, thereby eliminating cytotoxicity while preserving cellular structures
2Reliability
If flash freezing techniques are used, then cellular structure and biochemical identity are preserved, but cellular structure and function deteriorate due to marked changes from conventional freeze/thaw protocols
Solution Approach 1:
The invention performs preliminary optimization of the freeze/thaw protocol by establishing specific parameters (cooling rates, thawing conditions, incubation times) that prevent cellular damage before the actual isolation process begins
Solution Approach 2:
The invention implements dynamic control of the freezing and thawing processes, using controlled rate freezing followed by optimized gradual thawing protocols that adapt to tissue characteristics, preventing the marked structural changes seen in conventional static protocols
3Productivity
If living cells are extracted and grown in culture media, then cell availability for research is improved, but cellular characteristics deteriorate due to changes from stroma absence and culture conditions
Solution Approach 1:
The invention creates accurate copies of the native cellular state by isolating whole cells directly from frozen tissues with preserved protein expression patterns, providing a faithful representation of in vivo characteristics without requiring long-term culture expansion
Solution Approach 2:
The invention segments the research approach by obtaining cells at different stages (freshly isolated vs. cultured) and analyzing them separately, allowing comparison of cellular characteristics before and after culture-induced changes
4Ease of operation
If detergent treatment and homogenization are used, then cellular component analysis is enabled, but cellular integrity deteriorates due to cell disruption
Solution Approach 1:
The invention extracts only the necessary cellular components (isolating whole cells or specific proteins) rather than disrupting the entire cell structure, enabling targeted analysis while preserving cellular integrity for studies requiring intact cells
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 rapid and efficient isolation of whole cells from frozen tissues, preserving cellular structures and functions, allowing for accurate biochemical characterization of malignant cells, including protein expression patterns, which is beneficial for cancer research and oncology.
Implementation Method 1
cutting frozen tissue samples into pieces using a tissue sample cutter
Implementation Method 2
using a separation funnel to isolate whole cells without cryoprotective agents, allowing for rapid and efficient isolation of cells directly from stroma material
Data Source
AI summary
The present disclosure relates to methods for isolating whole cells from frozen tissue samples. The present disclosure further provides kits for obtaining isolated whole cells from a frozen tissue sample. One benefit to the methods and kits can be the rapid isolation of whole cells from frozen tissues upon thawing. Another benefit to the methods and kits disclosed herein can be the rapid and efficient extraction of isolated whole cells from stroma material in frozen tissues. Another benefit to the methods and kits disclosed herein can be the isolation of whole cells from frozen tissues suitable for subsequent biochemical analysis.