Acoustic Focusing Technology Overview
The Attune® Acoustic Focusing Flow Cytometer is the first cytometer that uses ultrasonic waves (over 2 MHz, similar to those used in medical imaging), rather than hydrodynamic forces, to position cells into a single, focused line along the central axis of a capillary.
What is Acoustic Focusing Cytometry?
Acoustic focusing cytometry is a technology that uses ultrasonic waves (over 2 MHz, similar to those used in medical imaging), rather than hydrodynamic forces, to position cells into a single, focused line along the central axis of a capillary (Video 1). Acoustic focusing is largely independent of the sample input rate, enabling cells to be tightly focused at the point of laser interrogation regardless of the sample-to-sheath ratio. This, in turn, allows slower cell velocities to collect more photons for high-precision analysis at unprecedented volumetric sample throughput.
|Video 1. Acoustic focusing in action. This video demonstrates how samples are aligned when the acoustic focusing is on or off.|
The Attune® cytometer accomplishes all this without high velocity or high volumetric sheath fluid, which can damage cells. In addition, volumetric syringe pumps enable absolute cell counting without beads—minimizing cost and sample preparation time.
Hydrodynamic Focusing vs. Acoustic Focusing
Manipulating cells in a conventional flow cytometer is accomplished using hydrodynamic forces. A suspension of cells (the sample stream) is injected into the center of a rapidly flowing sheath fluid, and the forces of the surrounding sheath fluid confine the sample stream to a narrow “core” that carries cells through the path of a laser that excites the associated fluorophores and creates a scatter pattern.
Keeping cells within a confined focal point is important for consistent excitation of the associated fluorophores as they pass through the tightly focused laser beam. Cytometers that use hydrodynamic focusing maintain the same sample speed at all flow rates, causing cells to lose focus as the sample core widens to accommodate the increase in flow rate (Figure 2).
|Figure 2. A wider sample core results in a broader distribution of cells as they transit through the laser, meaning fewer cells are accurately aligned with the laser focal point. To obtain optimal data from a conventional flow cytometer, with the lowest variability in signal detection, the instrument must be run at the lowest sample rate, which is typically 10–20 μL/min. Higher sample rates result in greater variability and less precise measurements. Acoustic focusing avoids this compromise in data and sample rates by uncoupling cell alignment from sheath flow.|
Due to many limitations of hydrodynamic cytometers, users often have to sacrifice:
- Throughput for sensitivity
- Features for ease of use
- Performance for price
- Power for footprint
Acoustic focusing technology keeps cells within a confined focal point, so these tradeoffs are not required.