APR Nov/Dec 2022 - 67

« SPECTROSCOPY
What is Single-Cell Analysis and
Why is it Important?
As the title suggests, single-cell analysis is the determination of trace
elements in individual cells. Understanding trace element presence
and concentration at the cellular level is imperative as there is often
great heterogeneity, even within the same cellular population and
in ideal conditions. Without understanding the spectrum of a trace
element's presence and its mass within individual cells, the nuances
of its impact on a cell, a population or even a bioculture cannot be
fully elucidated.
Traditional trace-element analysis methods involve digesting large
numbers of cells and analyzing the metal content within. Although
effective at measuring total trace element mass, these methods do
not account for cell-to-cell variation and cannot identify the number
of cells containing the element or the concentration contained within.
This means that valuable insights contained within the cells are lost.
Effective single-cell analysis is needed to accurately quantify the
average mass per cell and identify the element distribution across cell
cohorts (number concentration). These measurements are established
through two main calculations (Figure 1).
protect the cells, transport efficiency and detection sensitivity can be
improved. Transport efficiencies (for transferring cells from the sample
to the plasma source of the ICP-MS) of greater than 70% can now be
routinely achieved and high detection sensitivity, (measured through a
standardization curve and accounting for element ionization, focusing
and detection within the ICP-MS instrument), is achievable for a wide
range of elements.
The latest software can also help increase efficiency, improve
throughput and lower laboratory costs by automating the userdetermined
inputs (shown in green). Algorithms built into the software
can accurately measure transport and detection efficiencies and use
analytical parameters to suggest optimized volumetric flow rates and
dwell times to improve these rates.
Figure 1. Calculations for analyte mass and number concentration
used in single-cell trace element analysis.
Due to recent advances in ICP-MS technology, two key factors in
these calculations can now be observed: signal intensity and the
number of cell-derived events (shown in red), but the technology
and its software also bring other important accuracy benefits. By
using optimized workflows and tailored equipment that helps to
scICP-MS: Reaching New Levels
of Sensitivity and Speed for
Single-Cell Analysis
So-called single-cell ICP-MS (scICP-MS) is a technique that utilizes
the single-cell mode of the latest ICP-MS technology and can analyze
multiple trace elements (in sequential order) from a single sample.
The sample is nebulized to form a stream of single cells for individual
analysis. The resulting cells are then ionized, and ion optics focus
the beam into a quadrupole mass analyzer where ions, separated
according to their mass-charge ratio (m/z), are detected.
The latest scICP-MS technology provides very low detection limits
in low sample volumes, increasing sensitivity and allowing even low
levels of trace elements to be accurately identified. Added to this,
simple sample preparation methods and high throughput workflows
mean that a greater number of samples can be analyzed, even at the
single-cell level. Part of this efficiency comes from the technology's
ability to control interference and reduce background noise, meaning
that analysis of complex cultures requires minimal preparation steps.
However, to achieve high levels of efficiency and throughput
while maintaining accuracy, scICP-MS technology must be used in
combination with complementary technology and software.
Firstly, specialized nebulizers must be used to decrease the flow rate.
This protects the cells from damage and ensures a stream of single
cells for individual analysis with high transport efficiency.
Secondly, specialized software helps to drive key elements of the
workflow to maximize detection accuracy and efficiency. Decreased
dwell times enable detection of the very short, transient signals
that are emitted from single cells and sequential analysis programs
help to reduce settling times, maximize duty cycles to fully measure
m/z, and split sample time equally among analytes (if multi-analyte
measurement is needed).
Finally, data packages can automate evaluation parameters for more
accurate calculations. Known cell suspension standards can be used
to calculate transport efficiency and detection sensitivity can be
calculated by calibrating against ionic standards. Once the data is
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APR Nov/Dec 2022

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