ChemoMetec NC-100 User Manual

NucleoCounter®

User's Guide

P/N 991-0009

Revision 1.2

Technology that counts

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N u c l e o C o u n t e r

Mammalian Cell Counter

P / N 9 9 1 - 0 0 0 9 ( E n g l i s h )

R e v i s i o n 1 . 2

N o v e m b e r , 2 0 1 3

ChemoMetec A/S

Gydevang 43, DK-3450 Allerød, Denmark

Telephone: [phone], Fax: [phone] Internet: www.chemometec.com,

E-mail:[e-mail]

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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.

Contacting support

Technical information including product literature, answers to questions regarding the operation of the NucleoCounter not covered in this document and information on software upgrade is available through the following:

•For e-mail support, send questions to NucleoCounter Technical Support on the address [e-mail]

•To speak with a Technical Support Specialist, call ([phone].

Please note the NucleoCounter serial number and have it available when contacting ChemoMetec A/S for support. The NucleoCounter serial number is found on the label affixed to the bottom of the instrument and in the display upon start-up.

Sales and ordering information

For sales assistance with NucleoCounter or the NucleoView software, to place an order for a NucleoCounter or consumables, call ([phone], fax [phone], or send e-mail to [e-mail]

Disclaimer Notices

The material in this document is for information only and is subject to change without notice. While reasonable efforts have been made in the preparation of this document to assure its accuracy, ChemoMetec A/S assumes no liability resulting from errors or omissions in this document, or from the use of the information contained herein.

ChemoMetec A/S reserves the right to make changes in the product design without reservation and without notification to its users.

Copyright Notices

Copyright © ChemoMetec A/S 2005. All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, recording or otherwise, without the prior written consent of ChemoMetec A/S, Gydevang 43, DK-3450 Allerød, Denmark.

ChemoMetec and NucleoCounter are registered trademarks owned by ChemoMetec A/S. NucleoCounter , NucleoCassettes, Reagent A100, Reagent B and NucleoView are trademarks of ChemoMetec A/S.

All other trademarks are the property of their respective owners.

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Declaration of Conformity

Name of product:	NucleoCounter
Type:	Mammalian Cells
Other identifying data:	Part no. 900-0004

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.

This device complies with Part 15 of the FCC Rules. Operation is subject to the following two conditions: (1) this device may not cause harmful interference, and (2) this device must accept any interference received, including interference that may cause undesired operation.

Date: 2013-11-07

Signed:

Name: Sarah R. Rasmussen

Position: QA/QC Managers

Name and address of manufacturer:

ChemoMetec A/S

Gydevang 43

DK-3450 Allerod

DENMARK

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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. To prevent possible harm to the environment or human health from uncontrolled waste disposal, please separate this from other types of wastes and recycle it responsibly to promote the sustainable reuse of material resources.

Business users should contact their supplier and check the terms and conditions of the purchase contract. This product should not be mixed with other commercial wastes for disposal.

This information is listed in “Appendix 3: WEEE directive information in more EU languages”.

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Introduction and intended use

Introduction and intended use

The NucleoCounter® is a novel instrument designed specially to count mammalian cells. The instrument is a part of the NucleoCounter system, which also comprises the NucleoCassette™, Lysis buffer (Reagent A100) and Stabilizing buffer (Reagent B). The NucleoCounter system allows reliable, fast and objective cell counting to be carried out based on the method of fluorescence microscopy. It also makes it possible for the user to stain and count mammalian cells without handling hazardous dyes.

The NucleoCounter system has been developed to count mammalian cells in suspension. The mammalian cells could be cultured in T-flasks, bioreactors or on the surface of micro carriers.

The NucleoCounter is developed as a stand-alone instrument but optionally it can be connected to a computer via an USB-interface. When connected to a computer the NucleoViewTM software offers various features such as documentation of the obtained results. Read more about the software in the NucleoViewTM User´s Guide.

The NucleoCounter type at ChemoMetec A/S is 900-0004.

This user´s guide is valid for NucleoCounter software version 4.XX

The NucleoCounter system enables the user to obtain cell counts and viability determinations on cell suspensions from a wide range of cultured mammalian cell lines.

The cell counts and viability determination rest on the ability of the fluorescent dye to differentiate between viable and non-viable cells by means of different permeability of the plasma membranes. Thereby the concentration of viable and non-viable cells in suspension may be determined using the NucleoCounter system.

The NucleoCounter system is intended for fast, reliable and efficient cell counting and viability determinations of samples containing mammalian cells.

The NucleoCounter, NucleoCassette™, the Lysis buffer and the Stabilizing buffer are not intended for human or veterinary consumption.

The NucleoCounter system is for research use only, not for diagnostic use.

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Introduction and intended use

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Warnings and precautions

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 cable supplied with the NucleoCounter 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. External computing devices connected to the USB interface connector of the NucleoCounter has to comply with the standards, UL 1950 and IEC/EN 60950.

Table 1. Description of the NucleoCounter USB interface connector

	Pin no.	Name	Maximum
			Voltage level1
	1	+5V	+5 VDC
	2	D-	+3.5 VDC
	3	D+	+3.5 VDC
	4	DGND	0 VDC
	Metal	Shield
		(connected	0 VDC
	enclosure
		to DGND)

Use the shielded printer cable supplied with the NucleoCounter to ensure that you maintain the appropriate EMI classification for the intended environment.

The printer interface connector of the NucleoCounter must only be connected to the printer supplied by ChemoMetec A/S. External computing devices connected to the printer interface connector of the NucleoCounter has to comply with the standards, UL 1950 and IEC/EN 60950.

1 In normal operation mode (refers to Pin no 4)

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Metal Enclosure

Warnings and precautions

Table 2 Description of the NucleoCounter Printer Output connector

Pin no	Name	Maximum Voltage level2
13
2	Rx	±10 VDC
3	Tx	±10 VDC
Metal Enclosure	DGND	0 VDC

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Figure 1 Printer Output connector2 1 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 NucleoCounter shall only be used with one of the following external power supplies:

•Power supply Class I, Proton Electronic Industrial Co. Ltd., model SPN-260-12, input rated 100-240 Vac, 50-60Hz, 1.6 A. Output rated 12 Vdc, 4.3 A.

•Power Supply Class I, Powerbox Europe AB, model EBH 03 131, input rated 100240 Vac, 47-63 Hz, 0.8A. Output rated 11-13 Vdc, 2.7 A.

•Power supply Class II, Celetron USA Inc., model ZVC40LT12E, input rated 100-240 Vac, 50-60Hz, 1.2 A. Output rated 12 Vdc, 3.3 A.

The detachable power supply cord set and appliance inlet of the external power supply are considered as the disconnecting device.

The mains supply cord and plug of the external power supply shall comply with any national regulations.

2 In normal operation. Refers to DGND Metal Enclosure. Taken from the datasheet for MAX202ECSE.

3 There is no signal connected to pin 1.

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Warnings and precautions

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. These limits are designed to provide reasonable protection against harmful interference in a residential installation. This equipment generates, uses and can radiate radio frequency energy and, if not installed and used in accordance with the instructions, may cause harmful interference to radio communications. However, there is no guarantee that interference will not occur in a particular installation. If this equipment does cause harmful interference to radio or television reception, which can be determined by turning the equipment off and on, the user is encouraged to try to correct the interference by one or more of the following measures:

•Reorient or relocate the receiving antenna.

•Increase the separation between the equipment and receiver.

•Connect the equipment to an outlet on a circuit different from that to which the receiver is connected.

•Consult the dealer or an experienced radio/TV technician for help.

Caution! Changes or modifications not expressly approved by the party responsible for compliance could void the user's authority to operate the equipment.

Cassettes, Reagents and Dispensers

With respect to usage and handling of cassettes, lysis buffers and dispensers, please refer to appropriate package inserts for these items.

Caution! When using a bottle-top dispenser: To protect against accidental splashes protective clothing, eye protection and gloves must be worn when using potentially hazardous liquids.

General

Any biological specimen should be handled as if it is capable of transmitting infectious disease and disposed of with proper precautions according with federal, state and local regulations.

Avoid specimen contact with skin or mucous membranes.

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Warnings and precautions

Never pipette by mouth.

Avoid cross contamination of the samples when preparing of the samples. This can compromise the quality of the results.

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			1 Inspection and Unpacking of Equipment
Table of contents
DECLARATION OF CONFORMITY ................................................................................				VI
WEEE DIRECTIVE INFORMATION – EUROPE ONLY ......................................................				VIII
INTRODUCTION AND INTENDED USE ..........................................................................				X
WARNINGS AND PRECAUTIONS ...............................................................................				XII
1 INSPECTION AND UNPACKING OF EQUIPMENT ......................................................				3
2	INTRODUCING THE NUCLEOCOUNTER ...................................................................			5
	2.1	THE NUCLEOCOUNTER INSTRUMENT .......................................................................		5
	2.1.1		Fluorescence microscope .............................................................................	6
	2.1.2		Built-in image analysis ................................................................................	7
	2.2	THE NUCLEOCASSETTE ......................................................................................		7
	2.2.1		Loading the cassette....................................................................................	8
	2.2.2		Fluorescent dye ...........................................................................................	8
	2.2.3		Sample volume ............................................................................................	8
	2.2.4		Cassette handling ........................................................................................	9
	2.3	LYSIS BUFFER (REAGENT A100) AND STABILIZING BUFFER (REAGENT B) .........................		9
3 CONTROL BUTTONS, KEYPAD AND DISPLAY .........................................................				11
	3.1	INTERACTIVE CONTROLS ...................................................................................		11
	3.1.1		Control Buttons ..........................................................................................	11
	3.1.2		Keypad ......................................................................................................	11
	3.2	INTERFACE DISPLAY ........................................................................................		12
	3.2.1		Status indicator .........................................................................................	12
	3.2.2		Display screen ...........................................................................................	12
4	INSTALLATION AND START-UP ...........................................................................			13
	4.1	POWER ON, POWER OFF ...................................................................................		13
	4.1.1		Starting up.................................................................................................	13
	4.1.2		Shutting down ...........................................................................................	14
	4.2	INSTALLATION OF NUCLEOVIEW ..........................................................................		14
5 OPERATION OF THE NUCLEOCOUNTER .................................................................				15
	5.1	READY MODE ................................................................................................		15
	5.2	INSERTING AND REMOVING THE CASSETTE...............................................................		15
	5.2.1 Protection of the optical system ................................................................			15
	5.2.2		Inserting cassette ......................................................................................	15
	5.3	MEASURING - THE RUN BUTTON .........................................................................		16
	5.3.1		Actuator movements .................................................................................	16
	5.3.2		Result presentation ...................................................................................	16
	5.3.3		External computer .....................................................................................	17
	5.4	RESULT DISPLAY ............................................................................................		17
	5.4.1		Result mode ..............................................................................................	17
	5.4.2		Saving parameters .....................................................................................	18

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1 Inspection and Unpacking of Equipment

	5.4.3 Setting Date (F200) and Time (F201) .........................................................			18
	5.4.4		Reset Counter (F30) ...................................................................................	18
	5.4.5		LCD contrast (F220) ...................................................................................	19
	5.4.6		Instrument Info (F100) ..............................................................................	19
5.5		ZERO COUNT CHECK (F50) ...............................................................................		20
6 TOTAL AND VIABILITY COUNT ............................................................................				23
6.1		TOTAL CELL COUNT .........................................................................................		23
6.2		COUNTING NON-VIABLE CELLS ............................................................................		25
6.3		CALCULATE VIABILITY ......................................................................................		26
6.4		DILUTE THE SAMPLES.......................................................................................		26
7	MAINTENANCE OF NUCLEOCOUNTER ...................................................................			29
7.1		CLEANING....................................................................................................		29
	7.1.1		Cassette insertion area ..............................................................................	29
	7.1.2		Optical elements .......................................................................................	29
	7.1.3 Removal of dust particles ..........................................................................			30
	7.1.4		Spill of liquid .............................................................................................	31
8	TROUBLESHOOTING - ERROR MESSAGES .............................................................			33
8.1		NO VALID CASSETTE ........................................................................................		33
8.2		ANALYSIS ABORTED ........................................................................................		33
8.3		ACTUATOR ERROR MESSAGES .............................................................................		34
8.4		SAMPLE COULD NOT BE ANALYZED .......................................................................		34
8.5		SENSOR ERROR .............................................................................................		34
8.6		POWER-ON FAILURE........................................................................................		35
9	TECHNICAL SPECIFICATIONS ..............................................................................			37
9.1		THE NUCLEOCOUNTER ......................................................................................		37
9.2		THE NUCLEOCASSETTE™ ...................................................................................		38
9.3		EMC/EMI STANDARDS ...................................................................................		38
10	EQUIPMENT AND ACCESSORIES .......................................................................			39
10.1			EQUIPMENT AND ACCESSORIES LIST ..................................................................	39
10.2			POWER SUPPLY ..........................................................................................	40
10.3			MAINS POWER CORD ...................................................................................	41
10.4			DRY COMPRESSED AIR ..................................................................................	41
10.5			EXTERNAL PRINTER (OPTIONAL) ......................................................................	42
10.6			NUCLEOCOUNTER TEST KIT .............................................................................	43
10.7			NUCLEOVIEW .............................................................................................	44
11	APPENDIX 1: RESULT MODE 4 ........................................................................			45
12	APPENDIX 2: EXTENDED NUCLEOCOUNTER® NC-100T M SETTINGS .........................			47
13	APPENDIX 3: WEEE DIRECTIVE INFORMATION IN MORE EU LANGUAGES...............			49

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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.

Verify the ChemoMetec packing list received refers to the correct and ordered materials, and that nothing is missing.

If any part of the order was damaged during shipping or is missing, or fails to operate, please contact ChemoMetec A/S.

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1 Inspection and Unpacking of Equipment

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2 Introducing the NucleoCounter

2 Introducing the NucleoCounter

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.

The NucleoCounter system is comprised of the NucleoCassette for sample handling, the NucleoCounter instrument for particle analysis, a Lysis buffer for cell lysis (please refer to appropriate Application Notes provided by ChemoMetec concerning lysing of cells), and a Stabilizing buffer for optimizing the pH of the lysed cell suspension (please refer to appropriate Application Notes provided by ChemoMetec concerning stabilizing of the lysed cell suspension). Basics of the NucleoCounter system is explained on the next few pages.

The NucleoCounter system identifies and counts individual cell containing stained DNA. The sample pre-treatment is intended to allow the effective staining of cellular DNA but it will also aid the dissolving of cell aggregates. Depending on cell species and sample properties some additional effort might be needed to dissolve cell aggregates (please refer to appropriate Application Notes provided by ChemoMetec).

2.1The NucleoCounter instrument

The NucleoCounter is shown in the figure below.

Lid

			Weight	3 kg
	Control Buttons
			Height	26 cm
	Keypad		Width	38 cm
	Display		Depth	22 cm

Figure 2. NucleoCounter instrument

Refer to chapter 5 with respect to the operation of the instrument.

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