multifunctional single beam acoustic tweezer for non-invasive cell/organism manipulation and tissue imaging - mylar film roll
Precise manipulation of contact with individual particles, cells and raw objects has aroused great interest in the fields of biphysics and biomedical engineering.
Like optical tweezers, acoustic tweezers are thought to be able to manipulate particles and even cells.
Despite concerted efforts, the development of tools
Contact operation, no alternative to complex multi-function tweezers has been found yet.
Here we report a simple, low
Multi-Function single beam Acoustic tweeter (SBAT)
Being able to manipulate a single millimetre instead
Spherical cells under the Thunder regime, even single millimeter-level organisms under the Mie regime, and imaging tissues.
Our experiments show that SBAT with ultra-low f-number (
Focal length/aperture size)
Single red blood cells and single red blood cells can be manipulated. 6u2009mm-
The eggs of Zebrafish were fertilized in diameter.
In addition, in-vitro rat aortic images were successfully collected at dynamic focus, where the lumen and outer surface of the aorta can be clearly seen.
Super cheap f-
Number, SBAT provides a combination of large acoustic radiation forces and narrow beam widths, resulting in strong capture and high capture
Resolution imaging capability.
These properties enable non-performing using a single acoustic deviceinvasive multi-
Simultaneous functions of biomedical and biomedical applications.
SBATs are from a Krimholtz, Leedom and Matthaei (KLM)model.
Optimized thickness of lithium salt (LiNbO)
Single crystal is ~ 45 u2009 m, the aperture size is 1.
6 mhz mm with a center frequency of 60 mhz.
The first acoustic matching layer is a/4-
Thick silver epoxy resin made of silver particle mixture (Adrich Chem. Co. Milwaukee, WI)
501 epoxy resin and insulation (
US security Technology Corporation (Roslan, New Jersey)
, Cast on the opposite side of the chromium/Gold sputtering wafer.
A conductive silver epoxy (E-
Von Roll Isola, Solder 3022New Haven, CT)
As a backing layer, cast on the front of the wafer.
The acoustic stack with matching and support layers is pressed-
Focus on the focal length of 1.
0mm to get ~ A of 0. 6. A ~10u2009m-
The thick parylene layer is steam-
Deposited on the front of the SBAT as the second matching layer and protective layer.
Assemble the SBAT in the SMA connector for further experiments.
An acoustic treble experimental device was constructed using SBAT in the solution Chamber (see ).
The solution is Alsever's solution (A3551, Sigma-Aldrich, MO)
Studies on red blood cells when the solution provides water to the Zebrafish egg supplier (
Caroline Biological Supply Company)
Study on the eggs of Zebrafish fish.
In the red blood cell experiment, extract from healthy blood donors ~ 4 µL blood, washed three times with 1 µml phosphate
Buffer salt water (PBS).
In the solution of Alsever, the washed cells were diluted to 10/l.
During the experiment, the object is suspended in the solution.
Randomly target an object in the field of view of the microscope.
SBAT installed in a three
Shaft motor stage (LMG26 T50 MM;
OptoSigma, Santa Ana, California)
Controlled by a custom LabVIEW program, the focal length of the SBAT above the acoustic transparent mylar film is perpendicular to the beam axis.
SBAT is driven in sine burst mode under optimal driving conditions for combination of frequency, voltage, duty cycle and pulse repetition frequency.
Capture motion of an object is recorded at a capture frame rate of 10 frames/sec via a CMOS camera (ORCA-Flash2.
8 Hamamatsu, Japan)
With a microscope (IX-
71, Olympus, Japan).
To demonstrate the ability of acoustic manipulation, the SBAT of the trapped object is randomly moved by the maneuvering stage.
Take fresh blood from healthy donors and wash it three times with phosphate
Buffer salt water (PBS).
In each cell viability test, membrane-permeable live-
Calcein AM, a cell marker dye (In-vitrogen Corp.
Grand Island, New York, USA)
, As a reserve solution of 1mm, prepared in PX stored at room temperature.
10 CUCM of Calcein-
Work Solutions (
Is loaded into the cell.
Fluorescence imaging of cells before and after SBAT exposure with the highest driving power of sound manipulation (
Voltage = u2009 32 u2009 V, heavy-duty cycle u2009 0 =.
2%, pulse repeat frequency = u2009 1 kHz, exposure time limit 30 kbps s =).
Comparing changes in cell activity at a specific time interval (
0, 10, 20 and 30 minutes).
For statistical analysis, the mean value and standard deviation of the fluorescence level were obtained at each time interval, with sample size of n = ÷ 10.
Rat aorta of about 0.
9 cuccm diameter was harvested by surgeons at the University of Southern California medical school and stored in Eurocollins solution at 4 °c before the experiment.
Obtain images using an ultrasound biological microscope (UBM)setup.
SBAT connects a 3D electric linear stage (
Newport Corporation (CA)for scanning.
Water coupling for tissue samples.
The excitation pulse and echo signal are generated and received by the commercial pulse generator receiver (
JSR ultrasound, New York).
The signal is 2 ghz High-
Speed data acquisition card (Gage, IL). A Matlab (Mathwork, MA)-
Software based on control and imaging processing is developed, where the echo signal passes through the 4 th order Butterworth Band-
The information passed through the filter and covered was detected using the hillport transform.
In the merge
In the image, the triangle weighting function is used, where the weighting factor at the focus is 1, and the weighting factor at the focus is linearly reduced from 100 µm to 0.
For images with the closest/farthest focus, use a ladder weighting function.
Then, by applying the corresponding weighting function to the post-computing composite image
Filter the image.
All experiments on live mammals were conducted in accordance with the program approved by the University of Southern California's Institutional Animal Care and Use Committee.