TM-Sprayer™ Tissue MALDI Sample Preparation System Technical Note Optimization of Automated Matrix Spraying for MALDI Imaging The TM-Sprayer™ System is an automated MALDI plate matrix deposition system offering high reproducibility and superior data quality. TM-Sprayer™ is an easy-to-use, versatile spraying system that provides an automated process for the preparation of MALDI plates. The patented technology1 of the TMSprayer™ guarantees a very uniform and consistent coating crucial for reliable imaging. Automated Matrix Deposition Techniques for spray coating have utilized manual methods of matrix deposition, and are subject to variations dependent on the individual preparing the sample. Standardization of the matrix deposition provides a uniform and reproducible methodology. In the TM-Sprayer, a constant flow of heated sheath gas is delivered conjointly with the matrix spray which results in two major benefits: The unique design of the TM-Sprayer™ spray nozzle creates a fine solvent mist that is transported by a heated inert gas and quickly evaporates upon contact with the MALDI plates. Spray characteristics are easily adjustable via our intuitive Operator Interface. Users can select from a large selection of spray pattern designs and plate types to meet the requirements of their application. 1 Applicable patent 5,772,964 • Very rapid evaporation for minimal analyte delocalization. • Highly efficient co-crystallization for superior mass spec signal. Coupled with the benefit of faster sample preparation, the TM-Sprayer™ delivers the automated matrix deposition solution required to keep up with the high speed of the latest MALDI Mass Spectrometers. Large choice of Matrix The design of the matrix flow path provides a great flexibility in solvent choices. The TM-Sprayer™ permits processing of up to 1.0ml/min of matrix solution. Flow rates are controlled through an auxiliary pump. Consistent Matrix Spraying The temperature control technology used for the patented nozzle of the TM-Sprayer™ ensures very stable and reproducible spray characteristics, across a wide range of matrix solvent. Intuitive User Interface • • • Adjustable Method Parameters Nozzle height above target, temperature of matrix/solvent mix, temperature of gas and forward speed of cross table are easily controlled to result in proper application wetness, optimal resolution and extraction performance. High Quality Robotics The precise robotics of the TM-Sprayer™ deliver a smooth and constant-speed linear displacement of the MALDI target along the X-Y axis, which results in a uniform matrix deposition. Hot-spots often associated with manual techniques are eliminated. Easy Method Development The TM-Sprayer can save an unlimited number of methods, which makes it easy to access optimal parameters for each sample type of MALDI plate, matrix chemistry or types of tissue samples. • • Microsoft Windows™ Operator Interface Single view User Interface Graphical Selection of the desired deposition areas Linear offset tracks or an alternating 90° rotation during matrix application Save and Load routine sample programs Large selection of Spray Pattern Design Select from serpentine to vertical or horizontal line patterns. Custom configurable MALDI plate interface and multi-vendor plate holder Create new plate types using the simple plate configuration tool. SPRAY AUTOMATION IMPROVES SAMPLE PREPARATION AND REPRODUCIBILITY Figure 1: Improved reproducibility Figure 2: Improved data quality 1411 Day 496 856 1411 Manual Spray Manual Spray TM Sprayer™ TM Sprayer™ Day Less Protein Delocalization Fewer Hot Spots Sharper images All of the above images were acquired using 150 micron resolution. Samples were mouse brain sections. Manual spraying was performed using a glass Venturi sprayer. Details of automated spraying shown on right panel. Reproducibility studies were performed on different days, while data quality was evaluated using serial sections. High resolution MALDI mass spectrometric image of a mouse brain Method: Normal mouse brain was sectioned at 12 micron thickness and thaw mounted onto a metal MALDI target. The sections were dehydrated using ethanol washing and dried in a desiccated environment. Matrix was applied with the TM Sprayer™ using the following parameters: 142.5º C Sinapinic Acid 5mg/mL Block Temperature: Matrix: 300 mm/min 50% Acetonitrile in 0.1% TFA Stage Speed: 240 μL/min 10 psi Matrix Flow Rate: Air Pressure: 3 mm Line Spacing: 40 mm Nozzle Distance: Figure 3: After drying, the sample is imaged at 75 micron resolution and visualized using BioMap™ (Novartis) 1411 752 1519 6219 4970 671 5969 499 14166 (r) + 15198 (g) 14166 (r) + 4970 (g) 14166 (r) + 6219 (g) 14166 (r) + 4990 (g) Technical Characteristics TM-Sprayer™ nozzle design ensures a reproducible matrix from one sample to the next. Figure 4 Temperatu Temperature Contr Control l N2 Sheath G block heater h t • Sheath Gas – Ambient to 200°C (+/- 2°C), software selected • Collection Unit – X-Y Robotic Stage, fixed nozzle • Gas Supply Sheath gas flow 5-15.5 liter/min • Electrical 120 VAC, 60 Hz or 220 VAC, 50Hz: 2.5A fused • Dimensions/Weight 42x34x31 cm, 14kg Preheat capillary coiled in block it LEAP Technologies offers a complete line of sample preparation platforms for MALDI imaging matrix spray TM-iD™ Vision Enabled Liquid Dispensing for MALDI Spotting. 2D iDx™ Automated Gel Processing System Minimal Maintenance and Easy Service All key components are easily accessible for quick adjustment or maintenance. Figure 5 MALDI Spotter kit for HTS PAL Acknowledgements Protein Discovery, Knoxville, TN Additional Information TM-Sprayer™ is available worldwide from LEAP Technologies, Inc. To request further information please call LEAP Technologies at +1-919-929-8814 Contact: TM-Sprayer™ Specifications • Deposition – Spray deposition in linear or serpentine modes with variables offsets. Spray patterns by alternating perpendicular paths for even coatings • Nozzle Flow – 50 to 1000µl/min Alain Creissen International Sales Manager [email protected] LEAP Technologies PO Box 969 Carrboro, NC 27510 +1-919-929-8814 [email protected] www.leaptec.com