Cryo-EM Sample Qualification via Dynamic Light Scattering
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Solution Overview
Problem
Current methods for qualifying samples for cryo-electron microscopy, such as transmission electron microscopy, are invasive, unreliable, and costly, leading to issues like sample aggregation and clumping due to drying and interaction with sample carriers, which affects the accuracy of sample qualification and is time-consuming.
Innovation Solution
Implementing dynamic light scattering (DLS) directly on cryo-electron microscopy sample carriers to assess particle size distribution without altering the sample, using a DLS arrangement with a sample holder that protects the sample from evaporation and minimizes interaction with the grid, allowing for non-invasive, reliable, and efficient sample qualification.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If transmission electron microscopy is used for sample qualification, then sample suitability can be assessed, but the sample must be completely dried out which leads to oversaturation and aggregation
Solution Approach 1:
The patent replaces the mechanical drying process required in TEM with optical measurement (dynamic light scattering) that can be performed in liquid medium. The DLS technique uses laser light scattering to measure particle size distribution without requiring sample drying, thus maintaining the sample in its native hydrated state and preventing aggregation caused by water removal.
2Measurement precision
If transmission electron microscopy is used for sample qualification, then particle size can be measured, but heavy metal salts used as contrast agents can bind to macromolecules and cause rejection effects
Solution Approach 1:
The patent substitutes the TEM measurement method that requires heavy metal contrast agents with dynamic light scattering measurement. DLS measures particle size based on light scattering properties of the particles themselves in their native state, eliminating the need for heavy metal salts like uranyl acetate that can bind to proteins and cause de-aggregation or false positive results.
3Reliability
If transmission electron microscopy is used for sample qualification, then sample integrity can be assessed, but the process is time-consuming and has low sample throughput
Solution Approach 1:
The patent replaces the complex TEM imaging process with dynamic light scattering measurement. DLS provides rapid particle size distribution data without requiring sample preparation, imaging, and analysis steps associated with TEM. This substitution dramatically increases sample throughput while maintaining reliable assessment of sample integrity through accurate particle size measurement.
4Measurement precision
If transmission electron microscopy is used for sample qualification, then sample suitability can be determined, but the procedure is invasive and does not occur under the same conditions as cryo-EM recording
Solution Approach 1:
The patent replaces TEM with dynamic light scattering that can measure samples in liquid medium under conditions compatible with subsequent cryo-EM recording. DLS allows qualification to be performed on the same sample in its native hydrated state without invasive preparation steps, ensuring that the measurement conditions are compatible with the freezing and imaging conditions used in cryo-EM.
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 provides a reliable, cost-effective, and efficient method for sample qualification, ensuring accurate particle size determination and preventing aggregation, with higher sample throughput and reduced technical effort compared to traditional methods.
Implementation Method 1
DLS is an analysis method that is carried out on dissolved or suspended samples, usually proteins, by means of scattered light from a laser
Implementation Method 2
the fluctuations of the scattered light are quantitatively evaluated, wherein the particle size and its distribution are calculated by mathematical methods from these interference phenomena
Data Source
AI summary
A qualification process for a sample to be examined by means of cryo-electron microscopy. The, sample (12) is applied to a sample carrier (10) provided for cryo-electron microscopy and subsequently the sample (12) arranged on the sample carrier is examined by means of dynamic light scattering. The particle size distribution within the sample (12) is determined by means of the dynamic light scattering. Further, a sample holder designed to carry out the qualification process.