Helios Gene Gun System Instruction Manual Catalog Numbers 165-2431 and 165-2432 For Technical Service Call Your Local Bio-Rad Office or in the U.S. Call 1-800-4BIORAD (1-800-424-6723) Warranty and Regulatory Notices Warranty Statement This warranty may vary outside of the continental United States. Contact your local Bio-Rad office for the exact terms of your warranty. Bio-Rad Laboratories warrants to the customer that the Helios Gene Gun System (catalog number 165-2431 and 165-2432) will be free from defects in material and workmanship, and will meet all performance specifications for the period of one year from the date of shipment. This warranty covers all parts and labor. In the event that the instrument must be returned to the factory for repair under warranty, the instrument must be packed for return in the original packaging. Bio-Rad shall not be liable for any incidental, special or consequential loss, damage, or expense directly or indirectly arising from the use of the Helios Gene Gun System. Bio-Rad makes no warranty whatsoever in regard to products or parts furnished by third parties, such being subject to the warranty of their respective manufacturers. Service under this warranty shall be requested by contacting your nearest Bio-Rad office. The following items are considered customer-installable consumables: tubing, desiccant pellets, and microcarriers. The battery, razor blade, O-rings, barrel liner, cartridge holders, and syringe are replacement parts (see Section 9.2). These consumables and replacement parts are not covered by this warranty and are warranted only to be free from defects in workmanship. This warranty does not extend to any instruments or parts that have been subject to mis- use, neglect, or accident, or that have been modified by anyone other than Bio-Rad, or that have been used in violation of Bio-Rad instructions. The foregoing obligations are in lieu of all other obligations and liabilities including negli- gence and all warranties, of merchantability, fitness for a particular purpose or otherwise, expressed or implied in fact or by law, and state Bio-Rad’s entire and exclusive liability and buyer’s exclusive remedy for any claims or damages in connection with the furnishing of goods or parts, their design, suitability for use, installation or operation. Bio-Rad will in no event be liable for any special, incidental or consequential damages whatsoever, and Bio-Rad’s liability under no circumstances will exceed the contract price for the goods for which liability is claimed. Bio-Rad is not responsible for any injury caused by the use of this instrument for purposes other than those for which it is intended. Regulatory Notices Important: This Bio-Rad instrument is designed and certified to meet EN55011, EN50082-1, and EN61010 requirements, which are internationally accepted electromagnet- ic compliance and electrical safety standards. Certified products are safe to use when operated in accordance with the instruction manual. This instrument should not be modified or altered in any way. Alteration of this instrument will result in the following: Void the manufacturer’s warranty. Void the regulatory certifications. Create a potential safety hazard. Note: This equipment has been tested and found to comply with the limits for a Class A digital device, pursuant to Part 15 of the FCC rules. These limits are designed to provide rea- sonable protection against harmful interference when the equipment is operated in a commercial environment. This equipment generates, uses, and can radiate radio frequency energy and, if not installed and used in accordance with the instruction manual, may cause harmful interference to radio communications. Operation of this equipment in a residential area is likely to cause harmful interference in which case the user will be required to correct the interference at his own expense. Patent License and Usage Particle bombardment technology is covered by several patents which are held by E. I. duPont de Nemours & Co. and Auragen, Inc. Particle bombardment may be used for research purposes for gene delivery. Use of particle bombardment for commercial purposes requires a commercial license from the appropriate patent holder. The Helios Gene Gun is designed for research purposes only and is not intended for human or veterinary use. Licensed only for research use. Table of Contents Page Section 1 General Safety Information....................................................................1 1.1 Helios Gene Gun Safety........................................................................................1 1.2 Pressurized Helium and Nitrogen Safety..............................................................1 1.3 Power Safety..........................................................................................................1 1.4 Ear and Eye Protection..........................................................................................2 Section 2 Introduction to Particle Delivery...........................................................2 2.1 Particle Delivery Technology ...............................................................................2 2.2 The Helios Gene Gun............................................................................................3 2.3 Operating Principle of the Helios Gene Gun System...........................................3 2.4 Requirements for System Operation.....................................................................4 Section 3 Product Description ................................................................................7 3.1 Packing List...........................................................................................................7 3.2 Identification of System Components and Controls.............................................8 Section 4 Setting up the Helios Gene Gun System .............................................12 4.1 Inserting the Battery into the Helios Gene Gun .................................................12 4.2 Connecting the Helios Gene Gun to a Helium Source.......................................13 4.3 Connecting the Tubing Prep Unit to a Nitrogen Source ....................................14 Section 5 Operation of the Helios Gene Gun System.........................................17 5.1 Quick Guide to Operation...................................................................................17 5.2 Preparation of System Components Prior to Bombardment..............................18 5.3 Particle delivery using the Helios Gene Gun......................................................23 5.4 Removing Used Cartridges, Depressurization, and Shut Down ........................28 Section 6 Preparation of Mammalian Cell Targets............................................29 6.1 In vitro Delivery to Adherent Cells ....................................................................29 6.2 In vitro Delivery to Suspension Cultures............................................................30 6.3 in vivo Delivery to Epidermis .............................................................................31 Section 7 Optimization of Gene Gun Parameters ..............................................32 7.1 Overview .............................................................................................................32 7.2 Parameters for in vitro Delivery..........................................................................34 7.3 Parameters for in vivo Delivery ..........................................................................35 Section 8 Troubleshooting .....................................................................................35 8.1 DNA/Microcarrier Preparation...........................................................................35 8.2 Cartridge Preparation..........................................................................................35 8.3 Helios Gene Gun Operation................................................................................36 8.4 In Vitro and in Vivo Targeting ............................................................................36 Section 9 Product Information .............................................................................37 9.1 Helios Gene Gun System ....................................................................................37 9.2 Spare Parts...........................................................................................................38 9.3 Specifications ......................................................................................................38 Section 10 Appendices .............................................................................................39 10.1 Precipitation of RNA onto Microcarriers ...........................................................39 10.2 Replacing the O-rings and Inner Sleeve on the Helios Gene Gun.....................40 10.3 Replacing the O-ring on the Tubing Prep Unit...................................................41 10.4 Replacing Tubing Cutter Razor Blade and Unit Disassembly...........................42 10.5 Cleaning and Sterilizing the Helios Gene Gun...................................................43 10.6 Testing Cartridges for Microcarrier Penetration and Density............................43 10.7 Quantitation of DNA in Cartridges.....................................................................45 10.8 References ...........................................................................................................46 10.9