Sperm Sorting Using Split Enhancing Dye to Reduce Noise
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Solution Overview
Problem
Current sperm sorting methods face challenges in differentiating small variations in DNA content between X-chromosome and Y-chromosome bearing sperm due to noise from membrane compromised sperm and non-specific fluorescence, leading to reduced sorting efficiency and sperm damage from elevated temperatures and pH during staining.
Innovation Solution
The method involves staining sperm with a DNA selective fluorescent dye and a dead quenching dye to exclude membrane compromised sperm, followed by the use of a split enhancing dye to reduce noise and improve signal distinction, allowing for more precise sorting at lower temperatures and pH levels.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If a dead quenching dye is used to differentiate membrane compromised sperm, then sorting purity is improved, but over-quenching of desired signal occurs reducing measurement precision
Solution Approach 1:
The patent changes the chemical parameters of the staining system by introducing a split enhancing dye with different spectral properties than the dead quenching dye. This allows differentiation between quenched dead sperm and viable sperm with intact membranes, resolving the over-quenching problem while maintaining sorting purity
Solution Approach 2:
The split enhancing dye acts as an intermediary that modifies the interaction between the dead quenching dye and sperm cells. It provides an additional optical channel that mediates the detection signal, allowing the system to distinguish between specific quenching of dead sperm and non-specific quenching of viable sperm
2Stability of the object's composition
If Hoechst 33342 staining is performed with sufficient incubation time and concentration, then DNA staining uniformity is improved, but sperm damage increases due to elevated temperature and pH
Solution Approach 1:
The patent changes the physical parameters of the staining conditions by using alternative dyes and optimized incubation parameters that achieve sufficient staining uniformity at lower temperatures and pH levels, thereby reducing thermal and chemical stress on sperm cells
Solution Approach 2:
The patent employs dyes that provide adequate staining signal with shorter incubation times, effectively treating the staining process as a rapid transient operation that minimizes exposure to harmful conditions while achieving the required staining quality
3Measurement precision
If small variations in DNA content are measured to differentiate X and Y chromosome sperm, then sorting precision is improved, but noise from membrane compromised sperm reduces measurement precision
Solution Approach 1:
The patent segments the sperm population into distinct optical categories by using multiple dyes with different spectral characteristics. This creates separate detection channels that can independently identify viable sperm, dead sperm, and differentiate them based on DNA content, effectively isolating the signal of interest from noise
Solution Approach 2:
The split enhancing dye serves as an intermediary that enhances the optical contrast between viable and dead sperm populations. It mediates the detection process by providing additional spectral information that helps filter out noise from membrane compromised sperm while preserving the subtle DNA content variations needed for sex sorting
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 approach enhances sorting purity and speed by reducing noise and minimizing sperm damage, resulting in improved sex sorting of sperm with increased viability and reduced quenching of the overall signal.
Implementation Method 1
staining at least a portion of the population of sperm with a DNA selective fluorescent dye which fluoresces when excited
Implementation Method 2
staining said portion of the population of sperm with a dead quenching dye for selectively quenching fluorescence emitted by the DNA selective fluorescent dye within membrane compromised sperm
Data Source
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AI summary
A method of sex sorting sperm is disclosed. The sperm may be stained with a DNA selective fluorescent dye, which fluoresces when excited, a dead quenching dye, which selectively quenches fluorescence emitted by the DNA selective fluorescent dye within the membrane of compromised sperm, and a split enhancing dye. The stained sperm may then sorted into one or more gender enriched subpopulations of viable sperm.