Bio-rad PDS-1000 / He™ and Hepta™ Systems Manual do Utilizador

Biolistic®PDS-1000/HeParticle Delivery SystemCatalog Numbers165-2257 and 165-2250LEASEto 165-2255LEASEFor Technical Service Call Your Local Bio-Rad Of

Tools3 Hexagonal gap setting tools (1 each- 1/8", 1/4", and 3/8"), used to set gap between the bottom of the rupture disk retaining cap

PDS-1000/He Main unit (bombardment chamber with control panel and gauges; shippedfully assembled)Reinforced PVC vacuum tubing (1/2" ID x 3/4"

Microcarrier launch assembly (shipped fully assembled) consists of the following:Launch Assembly Shelf with Recessed Set ScrewMacrocarrier Cover LidAd

2.2 Identification of Unit Controls and ComponentsThe following is a brief description of the operation controls for the PDS-1000/He unit(Figure 2.9)T

Fig. 2.9. Front view of PDS-1000/He unit.Table 2.2. Front View-Interior of Bombardment ChamberBombardment Chamber Closes chamber with a solid piece of

Table 2.3. Rear ConnectionsRefer to Figure 2.10 for a rear perspective view of the unit.Helium Connection to Gas Tube Connects top of Gas Acceleration

Section 3Installation3.1 Connecting the PDS-1000/He System to a Helium SourceRefer to Section 2.2, Identification of Unit Controls and Components, pri

1. Secure the cylinder to a wall, post, or other anchored fixture so it will not tip or fall dur-ing use.2. Inspect the cylinder valve for dirt, dust,

3.2 Connecting the PDS-1000/He Unit to a Vacuum SourceThe main unit is shipped with the vacuum hose assembly attached to the port on the rearof the un

Fig. 3.2. Components used to connect vacuum source to main unit.3.3 Power Cord / Voltage RegulatorThe power cord must be plugged directly into a 100 V

Warranty and Regulatory NoticesWarranty StatementThis warranty may vary outside of the continental United States. Contact your local Bio-Radoffice for

Section 4Operation of the PDS-1000/He Instrument4.1 Preparation of System Components Prior to BombardmentInstrument Preparation1. Verify that helium t

Three hexagonal gap adjustment tools of 1/8", 1/4", and 3/8" have been provided to repro-ducibly set the gap distance (Figure 2.4). A 1

3. Prepare the Rupture Disk Retaining Cap. After setting the gap between cap and micro-carrier launch assembly, wrap the Rupture Disk Retaining Cap in

Consumable PreparationSeveral consumables are available for use with the PDS-1000/He system (Figure 4.3).Fig. 4.3. Consumables for the PDS-1000/He ins

Fig. 4.4A and 4.4B. Insertion of macrocarrier into macrocarrier holder with plastic insertion tool.3. Stopping screensTransfer selected stopping scree

Coating Washed Microcarriers with DNAThe following procedure is sufficient for six bombardments; if fewer bombardments areneeded, adjust the quantitie

Firing the Device1. Plug in power cord from main unit to electrical outlet.2. Power ON.3. Sterilize chamber walls with 70% ethanol.4. Load sterile rup

4. Coating microcarriers with DNAThe day of the scheduled bombardment, coat the microcarriers with DNA. To obtain thebest results, use the DNA-coated

7. Loading the rupture diskUnscrew the rupture disk retaining cap from the gas acceleration tube from within thebombardment chamber (Figure 4.6) or un

Screw the rupture disk retaining cap onto the gas acceleration tube using a left -to-rightmotion. Never tighten the rupture disk retaining cap without

Table of ContentsSection 1 Biolistic PDS-1000/He Particle Delivery System...11.1 Particle Delivery Techno

Fig. 4.9. Placement of stopping screen inside fixed nest with macrocarrier and cover lid removed. Fig. 4.10. Removal / replacement of macrocarrier cov

9. Target cells/tissue placement in chamberPlace the Target Shelf at the desired level inside the bombardment chamber. Place thesample (usually contai

Estimate rupture disk burst pressure by observing the helium pressure gauge at the top ofthe acceleration tube. A small pop will be heard when the rup

Fig. 4.13. View of spent rupture disk within the retaining cap.4.3 Removal of Residual Helium Pressure—Shut DownAfter completing all bombardment(s), r

Table 5.1. Cell Types, Settings, and ConditionsTarget HeliumCell Growth Cell Vacuum Distance Pressure ParticleType Phase Density Osmoticum (inches Hg)

Suggested starting helium pressure conditions for optimizing various biological systems are: Cell type Rupture disk pressureBacteria 1,100 psiFungi 1,

Fig. 5.1. Target shelf placement and corresponding target distances.5.9 Microcarrier SelectionTypes of MicrocarrierTwo types of microcarriers are avai

Recommended starting particle size/type for bombardment of various cell types isBacteria 0.7 µm (M5) tungstenYeast 0.6 µm goldAlgae 0.6 µm goldPlant c

Some kinds of embryogenic suspension cultures approach this ideal. In fact, it is this type oftissue which has allowed great success in the microcarri

more appropriate to use an extractive assay procedure in which expression level can be measured, for example, as a function of enzyme activity per uni

Section 1Introduction to Particle Delivery1.1 Particle Delivery TechnologyBiolistic particle delivery is a method of transformation that uses helium p

Section 6Troubleshooting 6.1 Rupture Disk Bursts at Incorrect PressureProbable Cause Action1. The helium flow rate is too fast. Note the helium burst

6.3 Excessive Gas UsageProbable Cause Action1. Leaking helium fitting in the system Isolate the components from the He cylinder to the solenoid assemb

6.6 Unit Will Not Pressurize Gas Acceleration TubeProbable Cause Action1. Rupture disk not in position or Insert disk properlynot sealed by retaining

Section 7Product Information7.1 Biolistic Particle Delivery SystemCatalogNumber Product Description165-2250LEASE PDS-1000/He Lease - 1st Year - Academ

CatalogNumber Product Description165-2296 500 Standard Pressure Kit, with 1.6 µ gold + 1,800 psi disks165-2297 500 Standard Pressure Kit, with 1.6 µ

Spare PartNumber Product Description910-9511 Helium Pressure Gauge910-9515 Vacuum Gauge 910-9529 Wrench, Open End, 1/4" x 5/16"Metal Case Ve

Section 8Appendices8.1 Cleaning the PDS-1000/He Device1. ChamberClean the chamber with 70% ethanol. Allow time for drying.2. Gas Acceleration tubeA. U

Aspects unique to the metal chamber design• Metal bombardment chamber.• Top slots of the metal case. These are used only for the target shelf placemen

Table 8.1 Sterilization and Decontamination CompatibilityMaterialAluminum Bunan Delrin®Neoprene Nylon Polycabonate PolyethyleneMethodSterilizationAuto

8.3 SpecificationsGeneral Specifications, PDS-1000/He Biolistic Particle Delivery SystemDimensions 29 (width) x 25.5 (depth) x 47.5 (height) cmConstru

The Biolistic ProcessThe Biolistic PDS-1000/He system uses high pressure helium, released by a rupture disk,and partial vacuum to propel a macrocarrie

8.4 Performing a BombardmentQuick GuideBefore the Bombardment1. Select/adjust bombardment parameters for Gap distance between rupture disk retainingca

Australia, Bio-Rad Laboratories Pty Limited, Block Y Unit 1, Regents Park Industrial Estate, 391 Park Road, Regents Park, NSW 2143 • Phone 02-9914-28

1.3 Important Safety InformationPressurized Helium Safety InformationCaution: Although helium is neither toxic nor flammable, all gases under pressure

The helium pressure regulator (supplied) has a CGA 580, female fitting (standard in theUnited States) for attachment to the user-supplied helium tank.

948, YEp352 DNA, CaCl2, spermidine, culture medium, and plating medium. Enough mate-rial for 60 bombardments is provided.Macrocarrier Holders, set of

6.0 ft PEEK plastic tubing (1/16" OD x .010" ID tubing & fittings), used to connect 3-wayhelium metering (solenoid) valve to helium pres