Operation Manual Encapsulator B-390 11593477 en
Table of contents Table of contents 1 2 3 4 5 About this manual . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 Safety . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 2.1 User qualification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 2.2 Proper use . . . . . . . . . .
Table of contents 6 7 8 9 10 4 Operation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .31 6.1 Starting up the instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31 6.2 Screens and menu functions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .31 6.3 Menu structure of the control unit . . . . . . . . . . . . . . .
1 1 About this manual About this manual This manual describes the Encapsulator B-390. It provides all information required for its safe operation and to maintain it in good working order. It is addressed to laboratory personnel in particular. If the instrument is used in a manner not specified in this manual, the protection provided by the instrument may be impaired.
2 2 Safety Safety This chapter points out the safety concept of the instrument and contains general rules of behavior and warnings from direct and indirect hazards concerning the use of the product. For the user's safety all safety instructions and the safety messages in the individual chapters shall strictly be observed and followed. Therefore, the manual must always be available to all persons performing the tasks described herein. 2.
2 2.4 Safety Safety warnings and safety signals used in this manual DANGER, WARNING, CAUTION and NOTICE are standardized signal words for identifying risk levels, related to personal injury and property damage. All signal words, which are related to personal injury are accompanied by the general safety sign.
2 Safety Explosive gases, explosive environment Harmful to live-forms Device damage Pressurized gas/air Hot Surface Wear laboratory coat Wear protective goggles Wear protective gloves Additional user information Paragraphs starting with NOTE transport helpful information for working with the device/software or its supplementaries. NOTEs are not related to any kind of hazard or damage (see example below). NOTE Useful tips for the easy operation of the instrument/software.
2 2.5 Safety Product safety Safety warnings in this manual (as described in section 2.4) serve to make the user alert and to avoid hazardous situations emanating from residual dangers by giving appropriate counter measures. However, risks to users, property and the environment can arise when the instrument is damaged, used carelessly or improperly. 2.5.1 General hazards The following safety messages show hazards of general kind which may occur when handling the instrument.
2 Safety Notice Risk of instrument damage by wrong mains supply. • • External mains supply must meet the voltage given on the type plate. Check for sufficient grounding. Notice Risk of damaging labratory glasses or utensils by moving syringe pump unit. • 2.5.2 Do not place any laboratory glasses or other utensils on the Encapsulator. Safety measures Always wear personal protective equipment such as protective eye goggles, protective clothing, and gloves when working with the instrument. 2.5.
2 2.6 Safety General safety rules Responsibility of the operator The head of laboratory is responsible for training his personnel. The operator shall inform the manufacturer without delay of any safety-related incidents which might occur during operation of the instrument. Legal regulations, such as local, state and federal laws applying to the instrument must be strictly followed.
3 3 Technical data Technical data This chapter introduces the reader to the instrument and its specifications. It contains the scope of delivery, technical data, requirements and performance data. 3.1 Scope of application and delivery The scope of delivery can only be checked according to the individual delivery note and the listed order numbers. NOTE For additional information about the listed products, see www.buchi.com or contact your local dealer. 3.1.
3 3.1.2 Technical data Standard accessories Table 3-2: Standard accessories Set of 8 single nozzles 11057918 Set of 8 single nozzles with high precision opening of 0.08, 0.12, 0.15, 0.20, 0.30, 0.45, 0.75 and 1.00 mm, made of stainless steel 316L including nozzle rack Pressure bottle 500 mL 11058190 Pressure bottle 1000 mL 11058191 Glass bottles with fittings, tubes and air filter, working pressure up to 1.5 bar, autoclavable 3.1.
3 3.1.4 Technical data Recommended spare parts Table 3-4: Recommended spare parts 14 Product Order no. O-ring set for single nozzle 11057954 O-ring set for concentric nozzle 11057955 Pre-filters for nozzle, diameter 7 mm (10 pcs.) 11057957 Drain filters for reaction vessel, diameter 35 mm (10 pcs.
3 3.2 Technical data Technical data Table 3-5: Technical data Encapsulator B-390 Power consumption max. 150 W Connection voltage 100–240 VAC Mains supply voltage fluctuations up to ±10% of the nominal voltage Frequency 50/60 Hz Fuse 3.15 A Dimensions (W × H × D) 32×29×34 cm Weight 7 kg Nozzle diameter of single (= core) nozzles 0.08, 0.12, 0.15, 0.20, 0.30, 0.45, 0.75 and 1.00 mm Nozzle diameter of shell nozzles 0.20, 0.30, 0.40, 0.50, 0.60, 0.70 and 0.90 mm Droplet size range 0.
4 4 Description of function Description of function This chapter explains the basic working principle of the Encapsulator B-390. It also shows how the instrument is structured and provides a general functional description of its assembly. 4.1 Functional principle The instrument provides the following key functions: Reproducible bead size from one production to the next –– A djustable parameters (nozzle size, liquid flow rate and vibration frequency) determine bead size.
4 Description of function Figure 4-1: Schematic representation of the Encapsulator B-390 a Pressure bottle Bead producing unit Vibration unit Single nozzle Electrode Dispersion control 17 Vibration control LED/stroboscope Polymerization bath Magnetic stirrer (P) Air pressure B-390 Operation Manual, Version C
4 Description of function The main parts of the Encapsulator B-390 are the control unit, the bead producing unit, and the pressure bottle. All parts of the instrument which are in direct contact with the immobilization mixture can be sterilized by autoclaving. The product to be encapsulated (active ingredients, enzymes, chemicals and cells) is mixed with an encapsulating polymer (typically alginate) and the mixture put into the pressure bottle see figure 4-1.
5 5 Putting into operation Putting into operation This chapter describes how the instrument has to be installed. It also gives instructions for the initial startup. NOTE Inspect the instrument for damages during unpacking. If necessary, prepare a status report immediately to inform the postal company, railway company or transportation company. Keep the original packaging for future transportation. 5.1 Installation site Put the instrument on a stable, horizontal surface.
5 5.2 Putting into operation Installing the Encapsulator B-390 Place the instrument on the lab bench with convenient access to an AC electrical outlet and to compressed air. Place the instrument in a way that disconnection of the electric supply plug is possible at all times. Connect the external air/gas supply (=air inlet) and the vibration unit as shown in figure 5-1. 4 3 1 a Air inlet (blue tube 2.6×4.
5 Putting into operation Installation of the air line A 3 m air tube (2.6×4.0 mm) is included with each Encapsulator to connect it to external compressed air or nitrogen. 1. Stick the air tube into the air inlet plug. 2. Attach the other side of the air tube to the external gas supply. 3. Deliver gas to the Encapsulator at 1.5 to 2 bar (23 to 30 psi) when running the instrument. NOTE The integrated pneumatic system (valve and fittings) will tolerate up to 7 bar (100 psi) at the inlet.
5 5.4 Putting into operation Assembling of the bead producing unit The bead producing unit is the central part of the Encapsulator B-390. It is fully autoclavable. Figure 5-3 shows the different parts of the bead producing unit. The assembled bead producing unit is attached with the screw on the carrier plate of the control unit. The vibration unit is placed on the magnet holder without the need of further attachment. 1 2 3 4 5 6 a Magnet Holder * b O-Ring (14×1.
5 5.6 Putting into operation Electrode The electrode is part of the electrostatic dispersion unit. It is attached from below to the heating block. The distance between the electrode and the nozzle tip can be changed as needed. This distance is approximately 3 to 8 mm. It should be set so that the droplets are formed near the upper side of the electrode. When the beads pass through the electrode they pick up the electrostatic charge.
5 Putting into operation Figure 5-7: Position of the electrode below the carrier plate Figure 5-8: Grounding the polymerization bath 24 B-390 Operation Manual, Version C
5 5.7 Putting into operation Pressure bottle The pressure bottle is an autoclavable container used to push the encapsulation mixture by air pressure into the bead producing unit. Figure 5-9 shows the different parts of the pressure bottle. The glass recipient has a guaranteed pressure resistance of 1.5 bar. The liquid flow rate is controlled at two levels: 1. With the air pressure by the pressure regulation system, and 2. with the liquid flow regulating valve situated on top of the control unit.
5 5.7.1 Putting into operation Installation of the pressure bottle 1. Assemble and – if needed - autoclave the pressure bottle. 2. Fill the bottle with the immobilization mixture. 3. Attach the silicone tube of the pressure bottle to the luer lock inlet of the bead producing unit. 4. Pass the silicone tube in the liquid regulating flow valve. Squeeze it so that no liquid can pass. 5. Insert the nipple of the air tube into the quick coupling of the air outlet at the control unit.
5 5.8 Putting into operation Option: Concentric nozzle system The concentric nozzle system (CN system) is an optional kit to the single nozzle unit. It is for the production of capsules in a one-step procedure. The system consists of CN bead producing unit, a set of 7 shell nozzles (0.20, 0.30, 0.40, 0.50, 0.60, 0.70 and 0.90 mm) and one pressure bottle of 1000 mL. The shell liquid is pumped by air pressure using the pressure bottle.
5 Putting into operation Figure 5-13: CN bead producing unit with set of 7 shell nozzles. The following nozzle apertures are standard: 0.20, 0.30, 0.4, 0.50, 0.60, 0.70 and 0.90 mm. 1 2 4 3 7 8 5 9 6 10 Figure 5-14: Single parts of the CN bead producing unit a Screw M3×6 b Luer lock female c Pre-filter grid 50 µm mesh, D= 7 mm d O-Ring 14.0×1.78 e Screw M3×8 28 f Screw M3×25 g O-Ring 3.68×1.78 h CN pulsation body i O-Ring 12.42×1.
5 5.8.1 Putting into operation Mounting of CN nozzles 1 2 Put the O-ring 12.42×1.78 in the grove of the CN bead producing unit. Put the inner nozzle (with attached O-ring) into the hole of the CN bead producing unit. There is no thread. The inner nozzle is centered and fixed by the shell nozzle. Exit of the shell liquid O-ring 12.42×1.78 Figure 5-15: Mounting of the inner nozzle Put carefully the shell nozzle over the inner nozzle. Attach the shell nozzle with two screws (M3×6).
5 Putting into operation Attach the complete CN bead producing unit to the carrier plate with two screws (M3×25). Attach the silicone tube of the core liquid to the core inlet port and the silicone tube of the shell liquid to the shell inlet port. Pass the silicone tubes in the liquid regulating flow valves. Squeeze them so that no liquid can pass. Figure 5-18: Connection of the pressure bottles to the air outlet. A T-piece feeds both pressure bottles. 5.
6 6 Operation Operation This chapter gives examples of typical instrument applications and instructions on how to operate the instrument properly and safely. See also section 2.5 “Product safety” for general warnings. 6.1 Starting up the instrument • • • 6.2 Make sure the Encapsulator B-390 is properly connected to the mains supply. Carry out a final installation check (see section 5.9) before every bead production. Switch on the Encapsulator B-390. The system runs an internal check.
6 Operation The frequency regulation generates the appropriate electric oscillation in the vibration unit. Pressing on the (+) and (–) buttons will change the frequency. Pressing the “on/off” button activates or deactivates frequency. Pressing “Esc” will return you to the start menu and the set value will be kept. Screen 6-2: Frequency regulation Screen 6-3: Electrostatic dispersion unit The electrostatic dispersion unit is used to charge the surface of the beads.
6 Operation The light intensity of the stroboscope lamp and amplitude ( = intensity) of the vibration can be set from 1 to 9. Above a frequency of 1500 Hz the amplitude can be set from 1 to 12. By increasing the amplitude the vibration becomes stronger. Values above 3 are mainly for solutions with viscosity > 100 mPa s. Pressing on the (+) and (-) buttons will immediately change the parameters. Pressing the “Esc”-button will cause a return to the start menu and the set value will be kept.
6 6.4 Operation Manual air pressure control In the control unit the pressure is manually controlled by the pressure regulating valve, integrated in the front panel of the control unit (see figure 6-2). Set the air pressure at a value which is 0.2 to 0.3 bar higher then the maximal air pressure needed during the encapsulation procedure; but not higher then 1 bar. Turning the knob of the pressure regulating valve clockwise increases the pressure, counterclockwise decreases the pressure.
6 6.5 Operation Practicing with the Encapsulator, using water Before working with encapsulation polymers, use water for practicing with the Encapsulator to become familiar with the effects of the controls. 1. Assemble the bead producing unit, screw the 0.30 mm single nozzle to the bead producing unit and attach all on the carrier plate with the screw (M3×25). Fix the electrode. Place the vibration unit on the bead producing unit. 2.
6 Operation Table 6-1: Determination of the working field (with pressure bottle) Nozzle size: Air pressure Clear bead chain without electrostatic tension Clear bead chain with electrostatic tension Lowest frequency Electrostatic Voltage Highest frequency Lowest frequency Highest frequency Nozzle size: Air pressure 36 Clear bead chain without electrostatic tension Clear bead chain with electrostatic tension Lowest frequency Electrostatic Voltage Highest frequency Lowest frequency Highest fr
6 Operation 7. Set the liquid flow rate and the vibration frequency to a value where a clear bead chain is obtained. Activate the electrostatic dispersion unit at 300 V and increase the tension stepwise by 100 V until the one-dimensional liquid jet is transformed into a funnel-like multi-line stream. The higher the electrostatic charge the earlier the bead chain is separated. This prevents the beads from hitting each other in flight, and from hitting each other as they enter the hardening solution.
6 6.6 Operation Practicing with the Encapsulator, using alginate solution After getting comfortable with the bead formation controls, perform test runs with non-sterile alginate solutions. Sodium alginate is the most commonly used polymer, but there are others in use with varying properties. We recommend the low viscosity grade alginate. The alginate concentration strongly influences the viscosity and this in turn influences the pressure drop in the nozzle.
6 6.6.2 Operation Working with alginate solution 1. Attach a 200 µm or 300 µm nozzle to the assembled bead producing unit. Attach all to the carrier plate. Check that the electrode is attached. Put the vibration unit on the bead producing unit. Put a magnetic stirrer below the nozzle and a large beaker on the stirrer. Fill the beaker with 100 mM CaCl2 so that at least 2 cm (approx. ¾”) is filled with the polymerization liquid.
6 Operation NOTE The stronger the circular dispersal of the bead stream the better is the bead homogeneity. This does not only depend on the electrostatic tension, but the liquid flow rate and the vibration frequency are also factors. Ideally, the bead should separate from the liquid jet within the electrostatic field between the nozzle and the end of the electrode. 8.
6 6.7 Operation Theoretical background Equ. 1: When a laminar jet is mechanically disturbed at the frequency ƒ, beads of uniform size are formed1. The optimal wavelength λopt for breakup, according to Weber2 is given by: Equ. 2: where: D = nozzle diameter η = dynamic viscosity [Pa s] ρ = density [kg/m3] (ca. 1000 kg/m3 for alginate solutions) σ = surface tension [N/m] (ca.
6 Operation Figure 6-4: Influence of the liquid jet velocity and the nozzle diameter on flow rate, as calculated by Equation 4. Figure 6-5 shows the correlation between the vibration frequency and the bead diameter for five different flow rates as calculated by equation 4. Lower flow rates, which corresponds to lower pumping rates, produce smaller beads. Higher vibration frequencies produce smaller beads also.
6 Operation Table 6-4: Optimal working conditions for the Encapsulator determined with alginate solution Nozzle diameter [µm] Flow rate * [mL/min] Frequency interval ** Amplitude Air pressure [bar] 1.0 mm 30 to 40 40 to 220 Hz 2 to 6 0.3 to 0.6 750 µm 19 to 25 40 to 300 Hz 2 to 5 0.3 to 0.5 450 µm 9 to 14 150 to 450 Hz 2 to 5 0.3 to 0.5 300 µm 5.5 to 7 400 to 800 Hz 1 to 3 0.3 to 0.5 200 µm 3.5 to 4.5 600 to 1200 Hz 1 to 3 0.4 to 0.6 150 µm 2.3 to 2.
6 Operation Figure 6-6: Amount of beads with a diameter of 0.3 to 0.6 mm formed from 1 mL of immobilization mixture. Figure 6-7: Amount of beads with a diameter of 0.6 to 1.1 mm formed from 1 mL of immobilization mixture.
6 Operation Figure 6-8: Amount of cells per bead made from different cell concentrations for bead diameters of 0.3 to 0.6 mm. Figure 6-9: Amount of cells per bead made from different cell concentrations for bead diameters of 0.6 to 1.1 mm.
7 7 Maintenance and repairs Maintenance and repairs This chapter gives instructions on maintenance work to be performed in order to keep the instrument in good and safe working condition. All maintenance and repair work requiring the opening or removal of the instrument housing must be carried out by trained personnel and only with the tools provided for this purpose.
7 7.4 Maintenance and repairs Cleaning ! Warning Pressure increase in the inlet-system due to clogged nozzles. Bursting of the inlet system. Death or serious poisoning by contact or incorporation of harmful substances at use. • Clean nozzle immediately after use, see following section. Wear laboratory coat Wear protective goggles Wear protective gloves 7.4.
7 Maintenance and repairs Figure 7-1: Cleaning procedure of the nozzle –– –– –– –– Take a syringe containing air on top and water on the bottom. Push the air through the nozzle (left figure). Push the water through the nozzle immediately afterwards (right figure). Examine the nozzle tip under a stereoscopic microscope to make sure the passage is clear and clean. NOTE If lipophilic immobilization solutions were used, then use appropriate solvents for cleaning.
8 8 Troubleshooting 8.1 Malfunctions and their remedy Troubleshooting The table below lists possible operating errors and their cause. As remedy set the parameter stepwise in the opposite direction or fix the missing part. Table 8-1: Possible cause Problem Possible cause Unstable liquid stream The liquid flow rate is too low. The nozzle is not adequately cleaned (frequent cause). The frequency is too high. The amplitude is too high. Unstable bead chain The frequency is too high or too low.
9 9 Shutdown, storage, transport and disposal Shutdown, storage, transport and disposal This chapter instructs how to shut down and to pack the instrument for storage or transport. Specifications for storage and shipping conditions can also be found listed here. 9.1 Storage and transport Switch off the instrument and remove the power cord. Wait until all hot parts (e.g. heating block and carrier plate) have cooled down.
9 9.2 Shutdown, storage, transport and disposal Disposal For instrument disposal in an environmentally friendly manner, a list of materials is given in chapter 3.3. This helps to ensure that the components can be separated and recycled correctly. You have to follow valid regional and local laws concerning disposal.
10 10 Declarations and requirements 10.1 FCC requirements (for USA and Canada) Declarations and requirements English: This equipment has been tested and found to comply with the limits for a Class A digital device, pursuant to both Part 15 of the FCC Rules and the radio interference regulations of the Canadian Department of Communications. These limits are designed to provide reasonable protection against harmful interference when the equipment is operated in a commercial environment.
10 10.2 Declarations and requirements Health and Safety Clearance form Health and Safety Clearance Declaration concerning safety, potential hazards and safe disposal of waste. For the safety and health of our staff, laws and regulations regarding the handling of dangerous goods, occupational health and safety regulations, safety at work laws and regulations regarding safe disposal of waste, e.g.
10 10.
BÜCHI Labortechnik AG CH-9230 Flawil 1/Switzerland T +41 71 394 63 63 F +41 71 394 65 65 www.buchi.
