NucleoCounter SCC-100 User's Guide Revision 1.
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NucleoCounter SCC-100 Somatic Cell Counter P/N 991-0200 (English) Revision 1.4 December, 2006 ChemoMetec A/S Gydevang 43, DK-3450 Allerød, Denmark Phone: (+45) 48 13 10 20, Fax: (+45) 48 13 10 21 Internet: www.chemometec.com E-mail: info@chemometec.
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Caution! This equipment must be operated as described in this User’s Guide. Please read the entire guide before attempting to use this unit. Please pay attention to that gloves or protective clothing are not worn on the illustrations/pictures shown in this User’s Guide. However, ChemoMetec A/S does recommend that the user wear suitable protective clothing etc.
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Declaration of Conformity Name of product: Type: NucleoCounter SCC-100 Other identifying data: Part no. 900-0200 The product complies with requirements of the following directives: 89/336/EEC - Electromagnetic Compatibility (EMC) 89/392/EEC - Machinery Directive Harmonized standards, which have been used: EN61326: 1997+A1: 1998 (Class B) +A2: 2000, Electrical equipment for measurement, control and laboratory use – EMC requirements (emission and immunity). Annex B and C from A1: 1998 is used.
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WEEE directive information – Europe only Correct Disposal of This Product (Waste Electrical & Electronic Equipment) - Europe only This marking shown on the product or its literature, indicates that it should not be disposed together with other household wastes at the end of its working life.
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Introduction and intended use Introduction and intended use The NucleoCounter SCC-100 is intended for use for the determination of somatic cell concentration in milk samples. The instrument is a part of the NucleoCounter SCC-100 System, which also comprises the disposable SCC-Cassette and the lysis buffer (Reagent C). The NucleoCounter SCC-100 System allows reliable, fast and objective somatic cell counting to be carried out based on an automated method of fluorescence microscopy.
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Warnings and precautions Wherever the symbol appears on the NucleoCounter instrument, it indicates that the manual must be consulted for precautions and warnings. Power and cables Use the shielded USB and printer cables supplied by ChemoMetec A/S to ensure that you maintain the appropriate EMI classification for the intended environment. The USB interface connector of the NucleoCounter must only be connected to SELV circuits.
Table 2 Description of the NucleoCounter Printer Output connector Name Maximum Voltage level 2 2 Rx ±10 VDC 3 Tx ±10 VDC Metal Enclosure DGND 0 VDC Pin no 13 1 3 2 Metal Enclosure Figure 1 Printer Output connector seen from the cable entry side. NucleoCounter is powered by an external 12VDC power supply. For safe use, please follow the instructions for connecting the power supply.
The user shall be made aware of that, if the NucleoCounter and the external power supply is used in a manner not specified by the manufacturer, the protection provided by the NucleoCounter and the external power supply may be impaired. Electromagnetic interference NOTE: This equipment has been tested and found to comply with the limits for a Class B digital device, pursuant to Part 15 of the FCC Rules.
Never pipette by mouth. Avoid cross contamination of the samples when preparing of the samples. This can compromise the quality of the results.
Table of contents DECLARATION OF CONFORMITY ...............................................................................VI WEEE DIRECTIVE INFORMATION – EUROPE ONLY ......................................................VIII INTRODUCTION AND INTENDED USE .......................................................................... X WARNINGS AND PRECAUTIONS ................................................................................1 1 INSPECTION AND UNPACKING OF EQUIPMENT ...............................
5.5 I NSTRUMENT 5.5.1 5.5.2 5.5.3 5.5.4 5.5.5 5.6 6 SETTINGS ................................................................................... 22 Setting Date (F200) and Time (F201) ......................................................... 23 Reset Counter (F30) .................................................................................... 23 Display contrast (F220) ............................................................................... 23 Instrument Info (F100)..........................
1 Inspection and Unpacking of Equipment 1 Inspection and Unpacking of Equipment Upon receiving the order from ChemoMetec A/S, the box or boxes should be carefully inspected for any damage that may have occurred during shipping. Any damage must be reported to the carrier and to ChemoMetec A/S immediately. Unpack the order, saving the packing materials for possible later use. Also be sure to save the User’s Guide, for instruction and reference.
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2 Introducing the NucleoCounter 2 Introducing the NucleoCounter SCC-100 The NucleoCounter method provides a fast alternative to the manual techniques, yielding objective and reliable results. Furthermore the NucleoCounter technology provides improved safety to the user compared to other comparable methods.
2 Introducing the NucleoCounter 2.1.1 Fluorescence microscope The compact fluorescence microscope integrated in the NucleoCounter comprises LED´s (light emitting diodes) as excitation light source, excitation and emission filters, optics (lenses) and a CCD (charged coupled device) camera. Furthermore, the NucleoCounter contains advanced software for image processing. The fluorescence microscope, with a cassette inserted, is shown in Figure 3. Figure 3.
2 Introducing the NucleoCounter A fluorescent image of the content of the measurement chamber is thus recorded. Example of such an image is shown in Figure 4. Each spot in the image represents a cell containing DNA stained with PI. Figure 4. Fluorescent image (from SomaticView PC software) of cell DNA contained in the measurement chamber of an SCC-Cassette 2.1.2 Built-in image analysis The NucleoCounter is equipped with advanced software for image analysis.
2 Introducing the NucleoCounter Figure 5 The SCC-Cassette. The fluorescent dye, propidium iodide, is immobilized in the first part of the flow system. 2.2.1 Loading the cassette The SCC-Cassette is loaded by gently pressing the white piston, which creates a partial vacuum in the flow system. The tip of the SCC-Cassette must be immersed into a lyzate mixture when pressing the piston, resulting in lyzate being loaded into the flow system. Approximately 50µl is loaded into the flow system. 2.2.
2 Introducing the NucleoCounter 2.2.3 Sample volume During production each cassette is marked with a dot code, which specifies the precise depth or thickness of the measurement chamber of the cassette. The dot code is read and decoded in the instrument during analysis and the volume analyzed is determined by multiplying the imaged area with the sample depth of the cassette. The imaged area is only dependent on the optics of the instrument. Therefore, the area is constant and specific for each instrument.
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3 Control Buttons, Keypad and Display 3 Control Buttons, Keypad and Display 3.1 3.1.1 Interactive Controls Control Buttons There are four Control Buttons on the front of the instrument (see Figure 6 ): Table 3.
3 Control Buttons, Keypad and Display 3.2 3.2.1 Interface Display Status indicator A small Status Indicator light above the “On/Off” button (see Figure 6) illuminates when the unit is turned on. It is green when the instrument is ready to measure and it turns red while the instrument is measuring or displaying an error message. The Status Indicator turns green after successful measurement when the results are displayed. 3.2.2 Display screen The display screen communicates information to the user.
4 Installation and Start-up 4 Installation and Start-up 4.1 Power on, power off The NucleoCounter is powered by a 12VDC external power supply. To connect the power supply to the NucleoCounter, connect it to the DC socket (see Figure 7), and plug the mains power cord of the power supply into a wall outlet. Figure 7. Connecting the 12VDC power cord plug to the NucleoCounter The NucleoCounter can now be turned on and off by pressing the “On/Off” button on the front panel. 4.1.
4 Installation and Start-up The start-up display shows the version number and date (year-month-date) of the internal software of the NucleoCounter in line 3 (“v4.XX” in Figure 8). The serial number of the instrument is shown in line 4 in the display (“002-01” in Figure 8). After a successful start-up the display changes (Figure 9) and the status indicator on the keyboard panel turns green. NucleoCounter Ready Figure 9.
5 Operation of the NucleoCounter 5 Operation of the NucleoCounter SCC-100 5.1 Ready mode After the NucleoCounter has started successfully the display should read: NucleoCounter Ready Figure 11. The NucleoCounter is ready for cell counting. This display is also shown if the “Esc” button is activated in the result mode or any other situation, except during measuring or shutting down. 5.
5 Operation of the NucleoCounter After analysis the lid is lifted and the cassette is removed from the insertion slit and disposed of according to national or regional laws or regulations regarding the nature of the mixture it contains. Figure 12. Illustration of how to grip and insert the cassette into the insertion slit 5.3 Measuring - The Run button When a cassette has been loaded and inserted into the instrument, the measurement is activated, by pressing the Run button. 5.3.
5 Operation of the NucleoCounter 5.3.4 External computer If an external computer with the SomaticView software installed on it is connected, the results and the recorded fluorescent image can be viewed using the SomaticView software. After the measurement is completed the cassette can be safely removed (see section 5.2). Sometimes, however, the complete process described above may be interrupted. Please familiarize yourself with possible error handling of the NucleoCounter presented in chapter 7 . 5.
5 Operation of the NucleoCounter Table 4 List of SCC and the corresponding s Poisson SCC S Poisson (SCC) '000 S Poisson (%) 50,000 7 14. 100,000 10 10. 200,000 14 7.1 400,000 20 5.0 600,000 25 4.1 800,000 28 3.5 1,000,000 32 3.2 1,500,000 39 2.6 2,000,000 45 2.2 The table above gives an indication of the number of significant digits in the reported results, when expressed as one standard deviation of repeatability.
5 Operation of the NucleoCounter 5.5.1 Setting Date (F200) and Time (F201) The functions, F200 (“F + 2 + 0 + 0 + ↵ ”) and F201 (“F + 2 + 0 + 1 + ↵ ”), are used to set the time and the date, respectively. The date and time are used when a printer or a PC is connected to the printer port. Please refer to Technical Note 001 (Part No. 9940014) with respect to setting and using the Time and Date features. 5.5.2 Reset Counter (F30) The function, F30 (“F + 3 + 0 + ↵ ”), is used to reset the Counter.
5 Operation of the NucleoCounter 5.5.4 Instrument Info (F100) When typing “F + 1 + 0 + 0 + ↵ ” the Instrument Info will appear in the display as in the Display Text Type shown below. ChemoMetec A/S NC SCC-100 v4.xx 2005-MM-DD S/N: 002-01 Figure 16. Instrument Info The display shows the instrument type in line 2 of the display. The version number and date (year-month-date) of the internal software of the instrument is shown in line 3. The serial number of the instrument is shown in line 4 of the display.
5 Operation of the NucleoCounter and with the lid closed. The check determines the number of CCD-pixels that show elevated signal levels. If the number of pixels is above a predefined limit then the cassette insertion slit can be contaminated with an interfering particle, like a dust particle. An error message is shown in the display and a cleaning of the insertion slit is recommended (refer to chapter 6 Maintenance of NucleoCounter).
5 Operation of the NucleoCounter After a few seconds the instrument has performed the Zero Count Check. If no contamination of the insertion slit can be detected an “OK” message appears in the display (see the figure below). Press any key to return to “Ready mode”.
6 Maintenance of NucleoCounter 6 Maintenance of NucleoCounter SCC-100 A regular cleaning of the NucleoCounter is recommended in order to protect its surface and assure the quality of the collected images. 6.1 Cleaning Depending on the environment in which the NucleoCounter is operated it is recommended that regular cleaning of the cabinet be carried out. When cleaning the cabinet it is recommended to use a soft moist cloth and gently wipe the surface.
6 Maintenance of NucleoCounter Figure 24. 6.1.3 An image showing a contamination of the optical system, visible as a white cloudy phenomena in the upper right hand corner. If the contamination is stationary it should be removed using compressed air. Removal of dust particles The presence of a contaminant (e.g. dust particles) will normally not influence the counting of cells.
6 Maintenance of NucleoCounter 6.1.4 Spill of liquid If liquid has been spilled on the instrument it can contaminate elements of the optical system. This contamination can show up in the image in several ways and have many causes but a common feature is that it cannot be removed by the use of compressed air. Even though such phenomena are clearly visible on the image it only rarely affects the results of the NucleoCounter.
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7 Troubleshooting - Error messages 7 Troubleshooting - Error messages Under certain conditions the NucleoCounter will display error messages during operation. Corrective actions are suggested below but if they do not correct the errors, contact ChemoMetec A/S or the local distributor. 7.1 No valid cassette When trying to analyze a cassette, which is not valid, the NucleoCounter displays the error message shown in Figure 26. No valid cassette Press any key Figure 26.
7 Troubleshooting - Error messages The error will also occur if the cassette has not been inserted properly into the NucleoCounter. Then the instrument cannot press the piston down and the error occurs. Take out the cassette, insert it properly and run the analysis. 7.3 Actuator error messages The actuator moves the piston down into the barrel of the cassette. The error message is shown in Figure 28. Actuator error Refer to manual Press any key Figure 28.
7 Troubleshooting - Error messages mixture in the cassette chamber. If one or more of the sensors has a faulty operation an error message will be displayed, see Figure 30. Sensor error Refer to manual Press any key Figure 30. This error message is displayed if one of the sensors, which read the black dots on the cassettes, causes error.
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8 Technical specifications 8 Technical specifications 8.1 Technical specifications for NucleoCounter SCC-100 Specificity Somatic cells stained with the DNA specific fluorescent dye, PI. Sample types Milk samples containing dispersed somatic cells. Sample consumption The sample is mixed with a lysis buffer prior to analysis. It is recommended that at least 500 µl of sample are used for this dilution in order to reduce error caused by sample handling.
8 Technical specifications Other environmental operating conditions for the NucleoCounter Indoor use only. Altitude Up to 2000 m. Mains supply Refer to the specifications for the Power supply in section 9.2 Power supply. INSTALLATION CATEGORY II (Refer to UL 61010A-1) OVERVOLTAGE CATEGORY II (Refer to UL 61010A-1) POLLUTION DEGREE II (Refer to UL 61010A-1) 8.2 The SCC-Cassette™ Reagent Each SCC-Cassette contains approximately 2,8 µg propidium iodide.
9 Equipment and Accessories 9 9.1 Equipment and Accessories Equipment and Accessories List The accessories for the NucleoCounter are listed in the table below. Ensure that items marked with this symbol are the only equipments and accessories used together with the NucleoCounter. Table 5. Equipment and Accessories for the NucleoCounter SCC-100 System Item NucleoCounter SCC-100 Part no. 900-0200 Description Instrument 4 SCC-Cassette, 100 pcs.
9 Equipment and Accessories Item Part no. Description Dry, compressed air See section below Used for removal of dust from the optical system of the NucleoCounter Finnpipette 100-1000 µL 911-0009 Pipette for milk sample and Reagent C Finntip 1000, 96 pcs. 911-0012 Disposable tips for pipette 9110009 Eppendorf 1.
9 Equipment and Accessories UL listed, type SVT, rated min. 60C, 18 AWG, 3 conductors. Provided with molded-on grounding-type (NEMA 5-15P) attachment plug, rated min. 125 Vac, min. 2.5A. Opposite end terminates in molded on, IEC320 style connector, rated min. 125 Vac, min. 2.5 A. For Europe 250Vac Cord type min. H05RR-F or min. H03VV-F or min. H03VVH2-F, rated min. 60C, 3x0.75 mm2. Provided with molded-on grounding-type attachment Plug, rated min. 250 Vac, min. 2.5 A.
9 Equipment and Accessories For connection of a printer (Impact type CM no. 939-0003; thermo type CM type no. 939-0006) please refer to Technical Note 001 (Impact type CM no. 994-0013; thermo type CM no. 994-0014). The NucleoCounter is equipped with a printer output port.
10 Appendix 1: WEEE directive information in more EU languages 10 Appendix 1: WEEE directive information in more EU languages United Kingdom Correct Disposal of This Product (Waste Electrical & Electronic Equipment) - Europe only This marking shown on the product or its literature, indicates that it should not be disposed together with other household wastes at the end of its working life.
10 Appendix 1: WEEE directive information in more EU languages Norw ay Korrekt avhending av dette produkt (Avfall elektrisk og elektronisk utstyr) - Kun Europa Denne m erkingen som vises på produktet eller dens dokumentasjon, indikerer at den ikke skal kastes sammen med annet husholdningsavfall ved slutten av sin levetid.
10 Appendix 1: WEEE directive information in more EU languages Finland Tämän tuotteen turvallinen hävittäminen (elektroniikka ja sähkölaitteet) - Vain Eurooppa Oheinen merkintä tuotteessa tai tuotteen oheismateriaalissa merkitsee, että tätä tuotetta ei tule hävittää kotitalousjätteen m ukana sen elinkaaren päätyttyä. Hallitsemattom asta jätteenkäsittelystä ympäristölle ja kanssaihm isten terveydelle aiheutuvien vahinkojen välttämiseksi tuote tulee käsitellä muista jätteistä erillään.
The information contained herein is to the best of our knowledge accurate and complete. However cell species and cell environments may vary in property. Therefore systematic and/or random deviation between estimates obtained by the NucleoCounter and other cell counting methods may occur. As such, nothing contained or stated herein including results obtained from use of the NucleoCounter or SCC-Cassette™ shall be construed to imply any warranty or guarantee.
