Applications

	Compound Resolubilization in Screening By sonicating precipitated samples just prior to liquid handling, those compounds which are capable of being dissolved will be driven back into solution, at least to the limit of their thermodynamic solubility.
	Compound Solubilization in Solubility Testing The SonicMan is an ideal adjunct to shake flask methods, as it provides a much faster method than shaking for driving compounds to the limit of their thermodynamic solubility.
	Small Volume Mixing Sonication provides a rapid mixing method which is independent of surface tension and wall effects in small wells.
	Bacterial and Mammalian Cell Lysis Sonication is a completely physicomechanical method of lysing cells, which can be performed in any buffer or medium. It can be fine-tuned for the cell type in question to provide just the right amount of energy to achieve lysis or release of a particular component, potentially resulting in cleaner extracts.
	DNA Shearing Sonication has been shown to break DNA into random small fragments.
	Tissue Homogenization and Disruption Sonication is an established method for homogenizing and disrupting tissues to release various components and can be done in any buffer, allowing cleaner extracts and homogenates to be produced.
	TPotential Applications (in the process of being validated): Sonochemistry Sonoporation and loading of macromolecules PMCA

Sonicman Features

	Stand-alone or integrated bench-top instrument
	Interchangeable 96, 384, and 1536 format disposable pinned lids (custom labware upon request)
	Plate shuttle which allows for direct integration with pick and place robotics
	Touchscreen interface
	Variable power settings between 1 and 1,150 Watts
	Variable sonication time intervals from 0.1 to 20 seconds
	Gasketed pin lids to ensure sample integrity with no well- to-well cross contamination

Disposable 96 & 384 pinned lids

Touchscreen interface

Gasketed lid design ensures airtight seal during sonication to eliminate cross-contamination

Measurement of Sonication Induced Mixing

The SonicMan uses sonic energy to speed mixing in small volumes as well as to remove air bubbles trapped in wells. In low volume 384 or 1536 well plates, sample mixing is achieved in just a few seconds of sonication. The following experiments demonstrate the measurement and visualization of sonication-induced mixing:

Diffusion vs. Sonication

Simple experiment: 50uL of concentrated sucrose colored with red dye was placed in 1.5mL microtubes, allowed to dry to a solid film and then covered with 500uL of DI water (Tube A). Even in the presence of plate shaking for eight hours (Tube C) mixing is not complete. By contrast, the SonicMan achieves complete mixing in 10-20 seconds (Tube B).


Measurement of Mixing

The evenness of mixing can be visualized in a 96-well plate (see series of figures below) using a two-component system, using a pH-sensitive dye (bromothymol blue, BTB) and selecting the starting pH and buffering capacity of the two components. The dye starts out in the blue form, but quantitatively converts to the yellow form on mixing. After 9 seconds of sonication, all but one well are completely mixed, and after 12 seconds, all wells are mixed.

The figure to the left shows the time profile of the absorbance at 615nm (blue dye form) for a 384- well plate with 70 uL per well. 20uL of pH=5 buffer, 50% sucrose, BTB (yellow form) was overlain with 50uL of a pH=10 buffer. Upon sonication, 98% of the wells are fully mixed within 10 seconds from the onset of sonication with all wells being fully mixed within 40 seconds. Differences in final signal are due to errors in pipetting the 50% sucrose solution.

Enhancement of Bioassay Results

Data shows a 384-well plate pre and post-sonication.
384-well plate containing related compounds and tested in a biological assay.

Data provided courtesy of Aventis Pharmaceuticals, Bridgewater, NJ.

Compounds in BLUE show biological activity pre-sonication.


Compounds in RED show significant activity post- sonication with the SonicMan.

Biological Applications

The figures below show the effect of increasing sonication on suspended mammalian and bacterial cells as measured by the protein released into medium. In both cases, the amount of protein increases up to a limit which is consistent with the amount of protein released by a commercial protein release agent. Maximal protein release is accomplished in less than 25 seconds.

Enhancing Solubilization

Dissolving compounds in DMSO prior to liquid handling is common practice in the setup of assays. An unknown but significant fraction of those compounds may not go into solution, or may go into solution only to precipitate out at a later time as a result of water pickup from multiple freeze-thaw cycles or extended time on the bench. Sonicating those samples just prior to liquid handling drives the compounds which are capable of being dissolved back into solution. Typically, such samples need to be shaken overnight to accomplish mixing and solubilization of compounds. The SonicMan shortens that process to a matter of seconds, turning a time consuming process into a reliable automated process.

The figure to the left shows results from five selected 96-well storage racks (80 cmpd/plate) out of a large number which were being prepared for screening. Each sample tube in each rack was individually inspected for precipitation. The racks were then sonicated using SonicMan, and reinspected. The graph shows the percentage of precipitated samples which were clarified by sonication. The improvement in solubilization ranges from around 25% up to 70%. (Data courtesy of Chiron Corp.)

In the experiment described below, three different methods of mixing are compared for 22 biologically active compounds of varying aqueous solubility. The amount of compound in solution in each case was measured using the NanoStream Veloce system. Results for the three methods are comparable for each compound, showing that sonication can achieve solubilization within seconds. (Data courtesy of Nanostream)

System Specification

Modes of Operation

	continuous
	timed
	pulsed
	timed/pulsed

Pulse on and off time is adjustable in 0.1 second increments from 0.1 to 20 seconds. Sonication power

Physical Dimensions Height: 787mm/31” Width: 356mm/14” Depth: 432mm/17” Weight: 125lbs/56.7kg
Required System Components
SonicMan instrument: Part#SCM1000


Plate Shuttle

settings are variable from 1 to 1,150 Watts. This combination of intermittent operation and variable amplification permits application specific, high- intensity sonication while avoiding heat build-up in the processed sample.

Remote Control

Ethernet port and command set

Plate Shuttle

The plate shuttle is in landscape presentation and allows for direct integration with pick and place robotics. Plate shuttle extends 115mm/4.5” (to center of plate) and is 140mm/5.5” above bench top

Sonic Horn

Constructed from titanium with carbide coating
CE Certified

Integration Software Package: Part#SCM1000-IS
Other System Requirements Compressed air: 100psi/7bar Electrical Requirements
208-240 VAC, 50/60 Hz, 10 amps
Environmental Requirements
Operating ambient temperature: 50-95°F/15-35°C Operating relative humidity: 0-90% at 85°F/30°C Altitude: 0-2000 meters
Accessories
Portable air compressor: Part#PAC-01
Disposable Sonic Lids (80/case):
• 96-format pinned lid Part#SL0096
• 384-format pinned lid Part#SL0384
• 1536-format pinned lid Part#SL1536
Step down transformer: Part#SDT-01 (110V)

Physical Dimensions

Height: 787mm/31”
Width: 356mm/14”
Depth: 432mm/17”
Weight: 125lbs/56.7kg

Required System Components

SonicMan instrument: Part#SCM1000
Integration Software Package: Part#SCM1000-IS

Other System Requirements

Compressed air: 100psi/7bar

Electrical Requirements

208-240 VAC, 50/60 Hz, 10 amps

Environmental Requirements

Operating ambient temperature: 50-95°F/15-35°C
Operating relative humidity: 0-90% at 85°F/30°C
Altitude: 0-2000 meters

Accessories

Portable air compressor: Part#PAC-01
Disposable Sonic Lids (80/case):
- 96-format pinned lid Part#SL0096
- 384-format pinned lid Part#SL0384
- 1536-format pinned lid Part#SL1536
Step down transformer: Part#SDT-01 (110V)

Compound Resolubilization in Screening

Resolubilization just prior to screening eliminates one major source of false negatives. It can have a noticeable impact on the quality of hits by facilitating better, more consistent data for structure-based screening data analysis software and SAR models.

The figure at top shows the comparison of IC50s determined before and after sonication of the stock DMSO solution for randomly chosen hits from a screen. Five of the thirteen compounds (38%) showed approximately two orders of magnitude improvement in measured IC50 if the stock compound solution was sonicated prior to aliquotting for the concentration-response experiment.