LU-CB1 Leiden Micro-Incubator User`s Manual
Contents
1. mal wear and tear or accident This warranty extends only to the original customer purchaser IN NO EVENT SHALL HARVARD APPARATUS BE LIABLE FOR INCIDENTAL OR CONSEQUENTIAL DAMAGES Some states do not allow exclusion or limitation of incidental or consequential damages so the above limitation or exclusion may not apply to you THERE ARE NO IMPLIED WARRANTIES OF MERCHANTABILITY OR FITNESS FOR A PARTICULAR USE OR OF ANY OTHER NATURE Some states do not allow this limitation on an implied warranty so the above limitation may not apply to you If a defect arises within the one year warranty period promptly contact Harvard Apparatus Inc 84 October Hill Road Building 7 Holliston Massachusetts 01746 1371 using our toll free number 1 800 272 2775 Goods will not be accepted for return unless an RMA returned materials authorization number has been issued by our customer service department The customer is responsible for shipping charges Please allow a reasonable period of time for completion of repairs replacement and return If the unit is replaced the replacement unit is covered only for the remainder of the original warranty period dating from the purchase of the orig inal device This warranty gives you specific rights and you may also have other rights which vary from state to state Repair Facilities and Parts Harvard Apparatus stocks replacement and repair parts When ordering please describe parts as completely as poss2. the 1 mm sockets ant the CO duct with the cutout of the sup port ring b Insert the micro incubator all the way up starting at the bottom of the support ring c Secure the micro incubator by tightening the screws located in the opposite side of the cutout of the support ring Heating Current 5 VDC 3 A max d Assemble one of the PVC insulator rings depending on the size of the open ing of the stage insert plate to the outside of bottom of the micro incubator This ring will act as an insulator to prevent heat loss by insulating the LU CB1 from the microscope stage Secure the bottom section of the support ring to the stage insert plate or PDMI Alignment Ring by using a sticky compound such as modeling clay Silicon Glue or similar method Connect the 6 pin Din connector output cable Catalog No 65 0100 to the TC 202A Front Panel I O Insert the cables with the 1mm plug through the strength relief holes and con nect the 1mm pin to the sockets of the micro incubator The ring terminal can be used to ground any other instrument related to your experiment Connect the temperature probe to the Bath Sensor in the TC 202A 7 Insert the 35 mm culture dish into the micro incubator 8 Attach the magnetic thermistor holder on the magnetic base metal plate on top 1 of the support ring Insert the temperature sensor probe through the coil and into the bath The temperature will be displayed in the LCD of the
3. the lowest possible temperature gradients between the cen ter as the side of the culture dish the gas can be used to maintain the partial pressure of gases in the culture medium e g 5 CO and 95 at a flow rate of 0 5 to 2 L min the pH of a bicarbonate buffered medium and its PO can be maintained Grounding Two independent ground pathways are also available when single channel or similar demanding electrical recordings are to be performed 1 Salt Bridge Ground The Support ring is fitted with a well that can be used as Bath Earth Pot for electrophysiology experiments A silver silver chloride Ag AgCD electrode can be introduce in the pot then is filled with KCI solu tion and an AGAR bridge used to complete the circuit with the medium in the culture dish during the experiment 2 Shield Ground A grounding terminal is connected at the end of the output cable connecting this to recording instruments or local grounds such as microscopes or Faraday cage Publication 5403 006 REV B LU CB1 Leiden Micro Incubator User s Manual Advantages and Accessories The two main advantages of the oil covered micro incubator are that it is freely acces sible for experimental manipulations and that disposable or reusable culture dishes can be used For biochemical analysis aliquots can be taken from the culture under the mineral oil surface and medium can be easily refreshed or exchanged in the same way In light microscopical studies f
4. LU CB1 Leiden Micro Incubator User s Manual LU CB1 Leiden Micro Incubator Users Manual MAI 65 0040 HARVARD APPARATUS Publication 5403 006 REV B WEEE RoHS Compliance Statement EU Directives WEEE and RoHS To Our Valued Customers We are committed to being a good corporate citizen As part of that commitment we strive to maintain an environmentally conscious manufacturing operation The European Union EU has enacted two Directives the first on product recycling Waste Electrical and Electronic Equipment WEEE and the second limiting the use of certain substances Restriction on the use of Hazardous Substances RoHS Over time these Directives will be implemented in the national laws of each EU Member State Once the final national regulations have been put into place recycling will be offered for our products which are within the scope of the WEEE Directive Products falling under the scope of the WEEE Directive available for sale after August 13 2005 will be identified with a wheelie bin symbol Two Categories of products covered by the WEEE Directive are currently exempt from the RoHS Directive Category 8 medical devices with the exception of implanted or infected products and Category 9 monitoring and control instruments Most of our products fall into either Category 8 or 9 and are currently exempt from the RoHS Directive We will continue to monitor the application of the RoHS Directive to its products
5. TC 202A Set the Control Point of the TC 202A to BATH 10 Turn the TC 202A ON and adjust the set point to the desired temperature from ambient up to 5000 Depending on the ambient the temperature the bath temperature will stabilize within 15 to 20 min after initial set up Publication 5403 006 REV B
6. and connected to external 1 mm jacks Gas perfusion inlet e Polyvinylchloride PVC support ring with magnetic base metal top to allow mounting of thermistor probes or other devices with magnetic holders around the incubator Support ring is fitted with wells that can be used as reference electrode wells for electrophysiological experiments and two screws to hold the micro incubator in place 3 PVC insulating rings to avoid contact with the insert plate of the micro scope stage and to prevent heat loss Publication 5403 006 REV B LU CB1 Leiden Micro Incubator User s Manual Operation The incubator is heated by passing electric current through the heating coil inside the anodized alu AC minium ring The duct inlet Ground in the ring directs and heats the incoming gas The cell culture is thus heated by direct heat conduction from the incubator as well as by convection as result of the heated gas blown over the surface of the bath or chamber The heat trans fer by convection reduces considerably the tempera ture gradient between the center and side of the culture dish A 35 mm OD x 10 mm high disposable culture dish or a Glass cover slip dish fits snugly into the incubator A layer of non toxic mineral oil 2 to 3 ml added to the dish will prevent the evaporation of the medium in the dish 2 to 4 ml For temperature control the TC 202A and its appropriate sensor BSC T3 are used The temperature contr
7. and will comply with any changes as they apply Do Not Dispose Product with Municipal Waste Special Collection Disposal Required 2 LU CB1 Leiden Micro Incubator User s Manual Table of Contents SUBJECT PAGE NO Warranty and Repair Information Serlal N mibersss te nase 2 BallDrallons ta aan 2 WENN ee aio c ate aeu d etta bids 2 Repair Facilities and Parts 2 Introduction Micro Incubator Construction 3 OpBeraON NR 4 Perfusion Fiud Perfisiort aste eio aite SD e Rec 5 Gas Perfusion and Grounding Gas Perfusion asc eacus 6 GOUGING exactes antenna kate 6 Advantages and Accessories Included Accessories suus 7 Optiorial ACCOSSOTIOS Rr neden 7 TT nasse T 8 Publication 5403 006 REV B LU CB1 Leiden Micro Incubator User s Manual Warranty and Repair Information Serial Numbers All inquires concerning our product should refer to the serial number of the unit Serial numbers are located on the rear of the chassis Calibrations All electrical apparatus is calibrated at rated voltage and frequency Warranty Harvard Apparatus warranties this instrument for a period of one year from date of purchase At its option Harvard Apparatus will repair or replace the unit if it is found to be defective as to workmanship or material This warranty does not extend to damage resulting from misuse neglect or abuse nor
8. ible preferably using our part numbers If practi cal enclose a sample or drawing We offer a complete reconditioning service CAUTION This apparatus is not registered with the FDA and is not for clinical use on human patients A CAUTION Not for clinical use on human patients Publication 5403 006 REV B E LU CB1 Leiden Micro Incubator User s Manual Introduction for Nikon x Insulating Olympus or Euri Rings Zeiss Micro p Scope Stage Screws 3 adjust height Q Output Cable Sensor Temper ature Probe Connector to Holder Temperature j Controller z y 1 mm pin Connectors to Heating Assembly The LU CB1 is a simple micro CO7 incubator designed for use on the microscope stage of a inverted microscope The micro incubator is easy to use and offers a handy tool for the culture of cells under the microscope The performance of the LU CB1 com pares well with that of a conventional COz microincubator A standard 35 mm plastic or glass culture dish can be placed in the micro incubator The culture medium is then covered by a gas permeable layer of mineral oil which protects the culture from the environment without affecting culture conditions allow ing easy cell manipulation under microscopical control Micro Incubator Construction The LU CB1 Leiden Micro Incubator consists of 3 torroidal rings anodized with black aluminum oxide film Two turns of cantal or nichrome wire wound around the middle ring
9. ol unit regulates the heating current by sensing the temperature of the incubator or of the medium via its sensor The TC 202A has been designed as bi polar and mono polar unit To achieve the feed back control and obtain maximum benefits from your LU CB1 Micro Incubator the TC 202A must be used on its mono polar mode See TC 202A setup for more information Gas for Duct Culture Bath Mineral Oil Support Magnet 2X Se Support Ring Heating Current Heating Coil Petri Dish 3X Insulating Ring Microscope Thermistor Figure 1 Cross Sectional Drawing of Micro Incubator Publication 5403 006 REV B E LU CB1 Leiden Micro Incubator User s Manual Perfusion ts ts ir Traps SWITCH EE ha Reservoirs Heat Exchange _ A OUT j f Figure 2 Perfusion System Fluid Perfusion The perfusion system above uses a peristaltic pump for both inflow and outflow of the bath On entry into the Faraday cage a bubble trap ensures electrical isolation of each channel The volume of fluid in the chamber is regulated by the changing height of the suction tube LU ASP Although the micro incubator was not designed with internal perfusion ports perfusion can be achieved by using a system that allows the fluid to be pre heated before entering in the bath However oil cannot be used in this configuration The temperature of the medium and the pH should be controlled just before it enters the culture dish The fluid tem
10. or example a transparent lid can be placed on the micro incubator to reduced the gas flow and drugs can be easily applied to the culture by opening and closing this lid During electro physiological experiments the lid can not be used however because free access to microelectrodes is necessary Gas flow then should be at least 1 liter min to minimize temperature gradient and obtain ade quate equilibration of the gas and the medium For all cases the use of more than 2 ml medium is recommended Included Accessories 1 Transparent Cover Lid Pin Magnet Set of 3 Insulating Rings with 40 mm 48 mm and 50 mm Diameters 6 ft Output Cable Sensor or Thermistor Holder NWR WON 1 mm Pins Spare 2 pcs Optional Accessories 1 Peristaltic Pump MP ID 2 Aspirator LU ASP for Fluid Perfusion 3 Alignment Rings to replace Insert Microscope Stage for Nikon Olympus Zeiss or Leica Microscopes 4 Fixing Platform LU FPZ for Zeiss Axiovert with Attachable Mechanical Stage M Type 451335 6 5 Fixing Platform LU_FPL for Leica DMIRB Mechanical Attachable Stage 520804 6 Cover Slip Glass Teflon pish Publication 5403 006 REV B LU CB1 Leiden Micro Incubator User s Manual 1 Set the temperature controller to Mono polar This is done by set ting the Incubator selector switch in the back of the TC 202A to BSC BU LU CB1 amp LU CPC 2 Assemble the micro incubator into the appropriate support ring as follows a Align
11. perature must also be as close as possible to the tem perature in the culture dish and have a constant flow rate minimizing the temperature oscillation of the medium The heating system of the culture bath then can be used to support the pH and the temperature control of the medium as it passes through the culture dish For reproducability at low flow rates a peristaltic pump is preferable over gravity feed This configuration also makes it easier to maintain the fluid level while changing solu tions An aspirator LU ASP can be used to remove the perfusate in the chamber via direct suc tion The aspirator has a magnet ic base to grip the magnetic metal base on top of the support ring The level of fluid in the chamber is determined by the height of this aspirator and can be changed by adjusting the nut on top of the threaded post The EN ar oscillations in inherent in any ERS peristaltic perfusion are easily QU AW damped with bubble traps N ee DE RS Publication 5403 006 REV B gt LU CB1 Leiden Micro Incubator User s Manual Gas Perfusion and Grounding Gas Perfusion This technique is intended primarily to reduce the radiant heat loss to or gain from the local atmosphere thereby reducing temperature gradients across the dish by con trolling the gas mixture above the chamber The gas passes over the heat exchange Any desirable gas mixture may be introduced into the micro incubator Besides its use to achieve
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