The PCT MicroTube Adapter Kit, in combination with the PCT SPS, can reliably and reproducibly control the enzymatic digestion of proteins while reducing the time of digestion from hours to minutes with the same or better quality as other, currently available techniques. Find out more at : www.pressurebiosciences.com

1. Pressure BioSciences, Inc.
PCT MicroTubeTM Adapter Kit*
for
Pressure-Enhanced Enzymatic Proteolysis
1 hr Trypsin Digest with PCT 12 hr Trypsin Digest No PCT
Pressure Cycling Technology
(PCT)
Discovery Starts with Sample PreparationTM
*Patent Pending

2. Pressure BioSciences, Inc.
Pressure BioSciences, Inc. (NASDAQ: PBIO) is a life sciences company
focused on the development and commercialization of a novel, enabling
platform technology called Pressure Cycling Technology (“PCT”).
Barocycler
PCT Sample Preparation System
The Pressure Cycling Technology Sample Preparation
System (“PCT SPS”) employs rapid cycles of hydrostatic
pressure, between ambient and ultra high levels, to
precisely control biomolecular interactions. The PCT SPS
can be used to accelerate enzymatic reactions, such as
protein digestion with trypsin and other proteolytic enzymes,
to prepare samples for analysis by mass spectrometry. In
addition, the PCT SPS can be used to disrupt tissues and
cells to extract cellular structures and components such
proteins, lipids, nucleic acids, and small molecules for
further analysis. The PCT SPS is comprised of a Barocycler
(a small, semi-automated bench top instrument used to
generate high, hydrostatic pressure) and specially designed
single-use processing tubes. Temperature can be
controlled during the PCT-based sample preparation
process by an external circulating water bath, which can be
an important addition to the Pressure Cycling Technology
Sample Preparation System.
“PCT offers clear advantages over our standard method of sample prep, including shorter processing time,
higher throughput, and reduced operating costs – and all without sacrificing quality. We found these advantages
so significant that we recently incorporated PCT into our standard mass spec proteomics analysis workflow.”
Dr. Shane Burgess
Director, Life Science and Biotechnology Institute
Mississippi State University
PCT MicroTube Adapter Kit*
Processing time, sample throughput, accuracy, efficiency, and standardization are key elements of the sample preparation
process for mass spectrometry. The PCT MicroTube Adapter Kit, in combination with the PCT SPS, can reliably and
reproducibly control the enzymatic digestion of proteins while reducing the time of digestion from hours to minutes with the same
or better quality as other, currently available techniques. The PCT MicroTube Adapter Kit comes complete with an ergonomically
designed, space-saving Work Station containing PCT MicroTubes and PCT MicroCaps, as well as tools and hardware, to enable
the user to process from one to forty eight samples simultaneously in the PCT SPS.
* Patent Pending
PCT MicroTube Adapter Kit

3. PCT MicroTube*
The PCT MicroTube was designed to meet the critical requirements of modern
proteomic analysis by mass spectrometry. It is made from a fluoropolymer,
fluorinated ethylene propylene (FEP). This polymer was selected because of
its unique features. FEP is highly inert and retains its integrity within an
extremely wide temperature range (-200oC to +100oC). The PCT MicroTube’s
outstanding chemical resistance, non-wetting surface, and negligible protein
and nucleic acid adsorption, help to ensure nearly complete sample recovery,
features that are essential for high pressure-enhanced enzymatic proteolysis.
* Patent Pending
PCT MicroCap*
PCT MicroCaps were also designed to meet the critical
requirements of modern proteomic analysis by mass
spectrometry. They are made from polytetraflouroethylene
PCT M icroCaps (PTFE), a polymer selected for features that are ideal for
high pressure applications. The PCT MicroCap is
Indent for available in three designs, designated 50, 100, or 150.
Picking G el Plug
Each MicroCap design designates an approximate reaction
volume. By selecting a particular MicroCap type, samples
may be digested in reaction volumes of approximately 50,
100 and 150 mL. In addition, all PCT MicroCaps have the
100 µ L added unique feature of being able to be used as gel spot
pickers to excise and transport small protein spots from a
150 µ L gel. The PCT MicroCap holds the gel protein spot safely
within the cap, while it is transferred to the PCT MicroTube,
50 µ L ready for processing by PCT.
* Patent Pending
“Incorporating PCT into our standard mass spec workflow has resulted in several significant advantages,
including an improved number of unique peptides identified and a reduction in undesired protein
modifications…and now with the new PCT MicroTube Adapter Kit, throughput will no longer be a bottleneck.”
Dr. Paul Pevsner
New York University School of Medicine
Suggested PCT Workflow

4. “PCT results in identification of more unique
peptides than our standard sample preparation
technique for mass spectrometry. It appears that
PCT may also be highly successful in digesting and
recovery of very complex samples, an area of great
difficulty at the present time. These advantages,
coupled with the unique characteristics of the
recently designed PCT MicroTubes, offer
researchers working in mass spectrometry-based
proteomics a wonderful new tool for scientific
discovery.”
Dr. Alexander R. Ivanov
Harvard School of Public Health
Figure 1. Quantitative Profiling of Peptide Abundance for In-Solution
Digested Model Proteins. Selected examples illustrate superior
digestion efficiency and peptide recovery using PCT.
Data provided by Dr. Alexander Ivanov, Harvard School of Public Health
Figure 2. PCT significantly increases digestion efficiency and peptide recovery following in-solution digestion.
Digestion was performed using an array of conditions, as indicated. LC-MS was performed on C-18, 10 cm x
150 µm (VC-10-C18WS-150EU) column using a 30 minute 5-60% gradient (A: 0.1% formic acid; B:
acetonitrile, 0.1% formic acid) Peptides were detected on an LTQ-Orbitrap XL mass spectrometer with ETD
ionization source; Alternating ETD/CID spectra were collected and the data were analyzed using Proteome
discoverer software with 1% FDR data filtering to determine sequence coverage and the total number of
peptides. Peak areas for selected peptides were measured using LCquan package for each LC/MS run (9
runs per digest condition). Values were normalized to the average area obtained for each peptide (n=72).
Median values from each set of 9 LC/MS runs for each peptide and each condition are presented.
Data provided by Dr. Roger Biringer, Thermo Fischer Scientific

5. Figure 3. PCT-assisted digestion has been shown to increase efficiency and peptide recovery from
gel-separated proteins. New consumable containers offer features which facilitate gel spot picking
using a combination of disposable gel cutter tool combined with the tube closure.
Data provided by Dr. Michail Alterman, CBER, FDA.
“We believe that this new patent pending sample containment and processing system, combined with the unique
power of PCT, gives the researcher extraordinary ability to gain new knowledge using today’s analytical
identification techniques, such as mass spectrometry. Based on the results from our beta site testers, we believe
that our PCT MicroTube Adapter Kit can benefit a large segment of the proteomics and genomics research
community.”
Dr. Edmund Ting
Sr. VP of Engineering
Pressure BioSciences, Inc
Features and Benefits of the PCT SPS and PCT MicroTubes
• Standardize Protein Digestion for Mass Spectrometry
• Substantially Increase Protein Digestion Efficiency
• Significantly Accelerate In-Solution and In-Gel Protein Digestion
• Promote Accurate & Efficient Peptide Cleavage
• Avoid Undesired Protein Modifications Due to Prolonged Digestion Time
• Process Up to 48 Samples at a Time
• Versatile Range of Sample Sizes (50-150 µL)
• Compatible with a Variety of Proteolytic Enzymes
• Inert, Non-Wetting, Single-Use MicroTube Processing Containers
• Unique MicroCap Design for Protein Spot Transfer from Acrylamide Gels
.

6. References
Papers
Schumacher R.T., et al., Automated Solution for Sample Preparation: Nucleic Acid and Protein Extraction from Cells and Tissues Using
Pressure Cycling Technology (PCT). Am. Laboratory. 2002, 34, 38-43
López-Ferrer D, et al., Application of pressurized solvents for ultra fast proteolysis: Proteomics on the fly. J Proteome Res. 2008;7(8).
Gross V., et al., Tissue Fractionation by Hydrostatic Pressure Cycling Technology:
The Unified Sample Preparation Technique for Systems Biology Studies. J Biomol Tech. 2008, 19(3)
Posters
A Comparison of Methods for Efficient Digestion of Protein Therapeutics
Lorna L. Maheu, Heather M. Connelly, Adam G. Harder, and Steven L. Cockrill
Analytical Sciences, Amgen Inc., Longmont, CO 80503
th
Poster presented at the 56 ASMS Conference, 2008
High Pressure Trypsin Digestion of Proteins for Proteomic Analysis
Shane A. Wyatt; Timothy R. Croley
Commonwealth of Virginia, Richmond, VA
Poster presented at the 56th ASMS Conference, 2008
Application of Pressurized Solvents for Ultra Fast Proteolysis: Proteomics on the Fly
Daniel Lopez- Ferrer; Konstantinos Petritis; Natacha M Lourette; Brian H. Clowers; Kim K. Hixson; Tyler H Heibeck; Eric A. Livesay; Ryan
Kelly; David Prior; Ljiljana Pasa-tolic; David G. Camp; Mikhail Belov; Richard D. Smith Pacific Northwest National Laboratory, Richland, WA
Poster presented at the 56th ASMS Conference, 2008
IMAGING MALDI of Colorectal Carcinoma-Field Defects in Satellite Tissue
Tiffany Remsen, M. Momeni, P. Kessler, F. Francois, A. Stern, S. Anand, and P. Pevsner
New York University School of Medicine
Poster presented Biomarker World Congress, 2009
Searching for efficient and high-throughput alternatives for essential sample preparation techniques in mass spectrometry-based
functional proteomics
Emily Freeman; Yelena Margolin; Alexander R. Ivanov
Harvard School of Public Health, Boston, MA
Poster presented at the ABRF Conference, 2009
Ultra-Rapid Pressure Digestion and Label-Free Quantitative Proteomics of Yersiniae Infected Mice Tissues
Kim K. Hixson1; Daniel Lopez Ferrer1; Matthew Bender2; Patricia L. Worsham3; Karl K. Weitz1; Nate Lawrence4; Amy Rasley5; Therese W.
Clauss ; Ljiljana Pasa-tolic1; Richard D. Smith1; Mary S. Lipton11Pacific Northwest National Lab, Richland, WA; 2NBACC, Washington, DC;
1
3
USAMRIID, Frederick, MD; 4Pressure Biosciences, Inc., South Easton, MA; 5Lawrence Livermore National Lab, Livermore, CA
th
Poster presented at the 57 ASMS Conference, 2009
Improved Protein Coverage and Throughput in Proteomics Using On-Line Multiplexed Enzyme Digestions and Targeted MS/MS
with a Modified LTQ-FTICR
Daniel Lopez Ferrer1; Konstantinos Petritis1; Andrei Liyu1; Yufeng Shen1; Benito Canas2; Kim K. Hixson1; Richard D. Smith1; Mikhail Belov1
Poster presented at the 57th ASMS Conference, 2009
Application of pressurized solvents for ultra fast proteolysis: Proteomics on the fly.
1
PNNL / Battelle Northwest, Richland, WA; 2Universidad Complutense de Madrid, Madrid, Spain
Poster presented at the 57th ASMS Conference, 2009
High-pressure assisted in-gel tryptic digest: qualitative and quantitative aspects
1 1 2 2
Michail Alterman ; Melkamu Getie-Kebtie ; Alexander Lazarev ; Vera S. Gross
1
FDA, CBER, Rockville, MD; 2Pressure Biosciences, Inc, Woburn, MA;
th
Poster presented at the 57 ASMS Conference, 2009
Proteomics under pressure: Rapid extraction and digestion in a single tube
2 1 1 1 2 th
Alexander Lazarev; Emily Freeman ; Vera S. Gross ; Greta Carlson ; Edmund Ting ; Alexander R. Ivanov Poster presented at the 57
ASMS Conference, 2009
For more information please visit Pressure BioSciences website: www.pressurebiosciences.com
For Research Use Only
Not for Use in Diagnostic Procedures
Barocycler®, PULSE ®Tubes, PCT MicroTubesTM, PCT MicroCapsTM , ProteoSolveTM
and The PCT Shredder KitTM
are trademarks of Pressure BioSciences, Inc.
14 Norfolk Avenue
South Easton, MA 02375
PH:508-230-1828 Fax:508-230-1829
www.pressurebiosciences.com