DNA-Technology DTprime User manual

Operation Manual
DTprime
Real-Time PCR instrument
PART I
Operating Procedure
TS 9443-004-96301278-2010
DNA-Technology, Research & Production LLC
Protvino

2

3
Safety Information
IMPORTANT: READ THIS MANUAL CAREFULLY BEFORE USING THE INSTRUMENT
General Safety
Whenever see the sign Caution! Danger! refer to the manual.
Before using the “DTprime” Real-Time PCR instrument (hereafter –the “instrument” or
“device”), please read this section carefully and pay particular attention to the safety
information. To avoid accidents, as well as breakdown of the instrument and equipment used
therewith, observe the safety rules given below.
If safety rules are violated during exploitation or in case of unintended use of the instrument,
the protection provided by the instrument may worsen and a health hazard may arise.
Do not operate the instrument if humidity exceeds 80% in the room. Condensate formation
may result of a short circuit in the instrument electronics.
The instrument should be saved from shocks and falling.
The instrument must be stored and transported only in a vertical position.
After transport or storage in humid and /or cold conditions, dry out the instrument not less
than 3 hours before connecting it to the supply voltage.
Avoid any liquids or objects getting inside the instrument's cabinet. This may result in
breakdown of the instrument.
“DNA-Technology, Research & Production”, LLC bears no responsibility for any injuries or
damage to health caused by unintended use or unauthorized modifications of the instrument.
Electrical Safety
Prior to connecting the instrument to an electric grid, check the presence of protective
grounding in the socket to which the instrument will be connected as well as integrity of the
power cable. Connection to the socket without grounding is prohibited. Please use the
power cable provided in the set of delivery. If liquid is spilt inside the instrument, disconnect
it from the power supply immediately and contact local service center or distributor.
Please connect the instrument to an electric grid with the corresponding voltage, which is
mentioned on the identification plate of it.
Caution! Danger of electrical shock! Replacement of fuses of the instrument has
to be carried out in compliance with the rules and guidelines of electrical safety
regulations. Fuses have to be replaced only when the instrument is switched off. The
instrument is considered to be unplugged if it is unplugged of the electricity socket and the
USB cable is disconnected from the USB 2.0 port of a PC.
During Operation
Do not expose the instrument to heat and sunlight or bright light from other intensive light
sources.

4
Maintenance
Do not open the cabinet of the instrument! It does not contain user serviceable
components.
Note! Settings of motor controllers, calibration parameters of optical and temperature
modules of the instrument cannot be changed by the end user. Calibration of the instrument
has to be performed by the manufacturer in accordance with the internal regulations of quality
control. Calibration data, if necessary, can be provided by the service department of DNA-
Technology Research& production
Operational Safety
The instrument is designed to be used under the following environmental conditions:
indoors;
altitude up to 2 000 m;
temperature from 5 °C up to 35 °C;
maximum relative humidity of 80% for the temperature up to 25 °C;
main voltage from 100 up to 220 V, frequency 50/60 Hz.
The istrument is IVD medical equipment and it is intended for studing of DNA samples using
the polymerase chain reaction (PCR) method and the detection of the accumulated product in
real time (real-time PCR). The instrument does not carry a direct biological hazard. Servicing
and maintainance of the instrument should be performed by specially trained, qualified staff.
While operation with the instrument:
wear suitable protective clothing, gloves and secure eyes and face;
do not pipette solutions by mouth;
do not eat, drink, smoke or use cosmetic products in the operational area;
after handling samples and chemical reagents wash your hands carefully;
dispose of leftover / used reagents and waste in compliance with the regulations in
force;
read the instruction for use provided with the PCR kit before running the assay;
while running the assay, follow the instructions for use;
do not use a PCR kit after its expiration date;
use only the chemical reagents provided along with the kit and those, which are
recommended by the manufacturer;
do not mix chemical reagents from different PCR kits or lots.
Environmental Effect
Deinstallation of the instrument for maintenance or recycling purposes: if properly operated,
the device doesn’t bear biologically hazardous.
Recycling of the instrument should be carried out in accordance with the regulations in force
at the area. The device does not contain any substances, which may represent a direct threat to
the environment.

5
Notice
This instruction for use contains information protected by copyright. No part of it may be
reproduced without preliminary written consent of “DNA-Technology, Research &
Production”, LLC. Any software products, equipment and designations mentioned in the text
may represent trademarks of their holders.

6
Table of Contents
1. General Information ............................................................................................................................ 8
1.1. Intended Use and modifications of the instrument....................................................................... 8
1.2. Delivery Set.................................................................................................................................. 9
1.3 Technical specification.................................................................................................................. 9
1.4. Operating Conditions.................................................................................................................. 11
1.5. PC Requirements........................................................................................................................ 11
2. Instrument description and principle of operation............................................................................. 12
2.1. Fast Thermal Control System.....................................................................................................14
2.2. Optical system ............................................................................................................................ 14
2.3. Control and Display System....................................................................................................... 15
2.4. Marking. Symbols used for marking.......................................................................................... 16
3. Setting up the instrument................................................................................................................... 18
3.1. Instrument Unpack ..................................................................................................................... 18
3.1.1. Instrument Carrying............................................................................................................. 18
3.2. Instrument installation................................................................................................................ 18
3.3. Software Installation................................................................................................................... 19
3.3.1. Instrument Driver Installation.............................................................................................. 19
3.3.2. Installation of RealTime_PCR software.............................................................................. 25
4. Operating Procedure.......................................................................................................................... 29
4.1. Instrument Switching-on and RealTime_PCR Launch ......................................................... 29
4.2. Preparing the Instrument for PCR.............................................................................................. 30
4.2.1. Checkup of Geometrical Settings........................................................................................ 30
4.2.2. Optical Measurement Exposure Setting .............................................................................. 31
4.2.3. Checking the Cleanness of Thermal Plate Wells................................................................. 32
4.2.4. Tubes Height Measurement................................................................................................. 32
4.3 Creation of Plate protocol and Amplification Program............................................................... 33
4.3.1. Filling in of Plate protocol................................................................................................... 33
4.3.2. Creation/editing of Amplification Program......................................................................... 34
4.4. Placement of Tubes with Samples............................................................................................. 36
4.5. Startup of the amplification program.......................................................................................... 37
4.6 Amplification program running................................................................................................... 38
5. Maintenance ...................................................................................................................................... 40
5.1 General Guidelines...................................................................................................................... 40
5.2 Maintenance Procedure ............................................................................................................... 41
5.3 Disinfection requirements of the instrument prior to maintenance and repair procedures.......... 41
5.4 Troubleshooting........................................................................................................................... 42
6. Storage and shipment ........................................................................................................................42
7. Recycling........................................................................................................................................... 43
8. EMS declaration................................................................................................................................ 43
9. Warranties.......................................................................................................................................... 44
10. Packing Certificate .......................................................................................................................... 45
11. Acceptance certificate...................................................................................................................... 45
12. Cards of warranty repair.................................................................................................................. 47
13. List of claims and instrument disinfection procedure...................................................................... 49

7
Introduction
The “DTprime” Detecting Thermocycler (hereafter –the “instrument” or “device”) is
intended to conduct qualitative and quantitative PCR analysis dropping out electrophoresis
stage of PCR products in agarose gel when using PCR kits based on the fluorescent detection
principle. Measurement of the quantity of accumulated product of PCR amplification takes
place right in the course of reaction temperature cycles (real-time PCR). The qualitative
analysis is based on evaluation of kinetics of PCR visible part, carried out using relevant
mathematical tool.
The use of fluorescent detection of PCR products has a list of substantial advantages:
high detection specificity (if oligonucleotide probes are applied, which allow detecting
only of certain amplicons);
high efficiency;
reduced assay time;
carrying out detection in a closed tube, which practically eliminates contamination of
further experiments;
availability of quantitative assessment of initial DNA-matrix;
registering and accounting of the data in electronic format.
Fig.1. Instrument General View

8
1. General Information
1.1. Intended Use and modifications of the instrument
The “DTprime” real time PCR instrument is intended for carrying out an assay using
polymerase chain reaction (PCR) method and accumulated product detection in a real time
(real-time PCR). Detection of the accumulated PCR amplification product occurs directly
during the execution of the amplification program, which increases the informative value of
the study.
The field of application - clinical laboratory diagnostics in vitro;
Purpose of use –diagnostics;
Sample type - samples of human DNA and microorganisms extracted from samples of
biomaterial;
The instrument is manufacturing in the following modifications: 2М1, 2М3, 2М6, 4M1, 4М6,
5M1, 5M3, 5M6, 6М6, 2Х1 4X1, 5X1, 6Х1.
Modification of the instrument is defined by the following structure –XXX, where:
1) 1st X is figure from 4 to 5 and it indicates the number of optical detection channels;
2) 2nd X is the symbol M or X and it determines the thermal block format: M - 96 wells of
200 l, X - 384 wells of 45 l.
3) 3rd X is figure 1, 3 or 6 and it means a thermal block version such as 1 –monoblock; 3 or
6 - the number of thermal block sections.
The instrument is designed for simultaneous analysis of up to 96 or up to 384 samples
depending on the modification (M or X).
Preparation and initialization of experiments on the instrument is realized with an IBM-
compatible personal computer, and further implementation of a preset program may be
executed autonomously. On completing the amplification program, the instrument goes to the
storage mode, and if the option “Turn off device after the run” is activated on the window
“Running”, the instrument goes to the power standby mode.
The optical measurements results may preliminarily be evaluated during the execution of the
amplification program, whereas the final evaluation –at any time after completing the
program. All data of the last amplification are saved in the internal memory of the instrument
and may be read out even in the case of power failure during amplification. As soon as the
mains supply is restored, the instrument continues to function with full recovery of the
amplification program status. Neither failure of the operating system, nor disconnection of the
external computer will halt the experiment. Thus, the instrument is highly protected against
major factors effect, which may result in a failure during PCR implementation.
One computer may control operation of several instruments, including ones of different types
(DTlite, DTprime) connected to it via USB ports.
To operate the instrument, the RealTime_PCR software is used, which allows performing the
following operations:
Test creation and editing;
Plate protocol creation;
Checking geometrical and optical settings of the device;

9
Creation, launching, and monitoring of the amplification program;
Monitoring the execution of the amplification program
Analysis of optical measurements data.
Data analysis report.
Settings and diagnostics of the instrument.
Loading and unloading of analyzed samples may be automatic in case if the instrument is
built in a robotic system.
Instrument is open system and it is not depended of concrete PCR kits.
1.2. Delivery Set
No.
Description
Reference
Producer
Quantity
pcs.
1
DTPrime real time PCR instrument
RT96-01IN
DNA-
Technology
R&P
1
2
Instruction for use (Part I and Part II)
RT96-04IFU
DNA-
Technology
R&P
1
3
Computer communication cable USB 2.0
High Speed А-В
—
China
1
4
Power cable (10А, 250V, 3х0,75mm2)
—
Taiwan
1
5
Fuses. (10А, 5х20, 250 V)
179200.10
Germany
1
6
Distribution package of software
RealTime_PCR
version 1.0.0.198
and higher
DNA-
Technology
R&P
2
7
Packing unit
RT96-03PG
DNA-
Technology
R&P
1
The instrument may be delivered along with portable PC with the pre installed software in
accordance with customer’s request. Note: PC should be purchased along with the order of
the instrument.
1.3 Technical specification
Specification
Value
Thermal block format (modification M)
96 test tubes of 0.2 ml (12 x 8)

10
Thermal block format (modification X)
384-well microplate
Tube type (modification M)
0.2-ml test tubes for PCR (individual, in strips, 8
pieces each, or in plates 12 x 8)
Tube type (modification X)
384-well microplate, 24x16 wells of 45 mcl
volume
Reaction volume (modification M)
10-100 mcl (10-50 mcl recommended)
Reaction volume (modification X)
5-30 mcl
Temperature range
0 C-100°C
Temperature step
0.1°С
Temperature accuracy
±0.2°С
Temperature uniformity
±0.15°С
Average heating ramp rate within temperature
range of 0…100°С (modification M)
Maximum heating ramp rate within temperature
range of 0…100°С (modification M)
3.3°C/s
3.5°C/s
Average cooling rate within temperature range of
100...55°С (modification M)
Maximum cooling rate within temperature range of
100...55°С (modification M)
2.1°C/s
2.5°C/s
Average heating ramp rate within temperature
range of 0…100°С (modification X)
Maximum heating ramp rate within temperature
range of 0…100°С (modification X)
2.1°C/s
2.5°C/s
Average cooling rate within temperature range of
100...55°С (modification X)
Maximum cooling rate within temperature range of
100...55°С (modification X)
1.0 °С /s
1.5 °С /s
“Hot lid” temperature
105°С ±1°С
Heating and cooling method
Peltier elements
Excitation source
LED
Detector
CCD camera

11
Number of optical channels
4 (versions 2M1, 2M3,2M6,2Х1) 4 (versions
4M1, 4M3,4M6) 5 (versions 5M1, 5M3,5M6,
4X1,5X1) 4 (versions 6M1, 6M3,6M6,6Х1) *
Excitation/Detection wavelengths for each channel,
nm
470/515, 530/560, 580/620, 630/660,687/731(*)
Threshold sensitivity of each channel for standard
fluorophors solutions
0,05хЕ-12 gram-mol
Data connection
USB 2.0 High-speed
Power consumption
550 W (max.)
Mains voltage
220 V
Line frequency
50/60 Hz
Dimensions, WxDxH
210х540х540 mm
Starting operation after switching on
no more than 5 minutes
Weight
27 kg
* spectral characteristics of optical channels can be changed within the specified limits 450 ÷
750 nm in accordance to customer request.
1.4. Operating Conditions
The instrument is intended for the use indoors under the following climatic conditions:
indoor temperature should be from 5°С to 35°С;
relative humidity for temperatures up to 25 °С should not exceed 80%;
at an altitude of not more than 2000 meters.
1.5. PC Requirements
The instrument operates under the control of IBM-compatible personal computer with the
following minimum requirements:
1 GHz Pentium IV;
512 Mb RAM;
Free hard drive space 100 Mb;
Screen resolution 1024x768 pixels;
USB 2.0 High-Speed port.
To control the instrument the following parts of software pack must be installed:
the instrument driver;
Realtime PCR software to control the instrument and perform data analysis.

12
The software may be downloaded for free from the company’s official website at
http://www.dna-technology.ru/eng/support/
The software is compatible with the following operating systems: Windows 98, Windows
ME, Windows 2k/XP, Windows Vista, Windows 7, Windows 8, Windows 10.
2. Instrument description and principle of operation
DTprime is a specialized instrument combining a precise programmable thermal cycler and an
optical system to collect fluorescent data at each stage of polymerase chain reaction.
Design of the instrument:
The design of the instrument is a supporting frame which bears the following installed
modules:
thermo block including positioning module of actuated parts of the instrument
designed for automatic moving of it;
the optical system module which consists of sources of excitation based on LEDs and
detection based on CCD camera;
optical path module for transporting light beams;
slide for the thermal block placement;
hot lid module, which presses the tubes from the top and heat them for preventing
condensation and also eliminates self-opening of the tubes in order to avoid a possible
contamination of the instrument by amplicones;
horizontal drive, which provides movement of the thermal block in horizontal plane
for loading and unloading tubes into it;
vertical drive, which provides movement of the thermal block in vertical plane to
ensure that the tubes are pressured to the thermal lid;
electronics modules including power supplies;
cabinet of the instrument, which has several ventilation holes.
The instrument face panel accommodates the following (fig 2):
LCD panel;
push-button control panel;
“hot lid” front panel;
thermal block front panel.
Side walls of the instrument have ventilation slots.

13
Fig.2. Front view of DTprime instrument
The instrument rear panel accommodates the following (fig 3):
power switch;
two fuses;
power input;
USB connection (type B).
Fig.3. Rear view of DTprime instrument

14
Functionally there are three main systems of the instrument:
high-rate temperature control system;
optical system;
control and display system.
2.1. Fast Thermal Control System
The system of high-rate temperature control includes a thermal unit, its movement and
positioning module, and the hot lid module.
The thermal block ensures thermal cycling of the tubes according to the amplification
program. Six Peltier elements are used for thermal plate cooling and heating providing high
accuracy of temperature regulation, noiselessness, good characteristics of mass-and-
dimension ratio and high reliability. Each of these Peltier elements has its own temperature
sensor and regulator that allows the preset of necessary temperature gradients across the plate.
The instrument thermal plate is made of aluminum alloy, which possesses an optimal ratio
between good thermal conductivity, weight, and processing characteristics. The plate design
is a cellular structure with reinforcing ribs and corresponding number of wells:
in M modification –96 wells for standard 200 mcl PCR tubes with distance between
the rows as in standard 96-well plates. The well shape provides optimal heat transfer to the
samples with volume up to 50 mcl.
in X modification –384 wells for standard 384-well PCR microplates.
Note! For stable results, tubes with the angle of tapered part of 17º20' must be used.
Movement and positioning module is intended for precise positioning of the thermal unit
within 180 mm and 50 mm in horizontal and vertical planes correspondently and provides
easy access to the tubes.
The hot lid module provides a reliable heat contact between tubes and the thermal plate and
allows maintaining temperature of the tube caps at the level of 105±1°С in order to prevent
condensate formation on the tube caps, since it may cause substantial distortions in the
luminous flux measurements. The hot lid design provides an adjustable force of tube-to-plate
hold-down that depends on the number of the tubes installed. During running PCR the
thermal block moves vertically from bottom to top. The moment of contact of the tubes with
the hot lid is monitored by a special sensor, which passes information to electronic module of
the instrument to calculate the necessary pressing force. The positioning module moves the
thermal block vertically by an amount that provides the necessary pressure to hold the tubes
against the surface of the hot lid.
2.2. Optical system
The optical system consists of an optical module and a light beam transmission module.
The optical module is a system of lenses, mirrors and light filters that provides folding of the
luminous flux from several light sources on one optical axis and its separation from the
luminous flux entering the CCD camera. The light sources are powerful LEDs. The light
source design provides efficient heat removing from emitting crystals of the light-emitting
diodes for stable luminous flux.
Optical path (light beams transmission module) ensures the transmission of the luminous
flux from light sources of the optical module to the tubes, and back transmission of the
fluorescent luminous flux from the tubes to CCD camera.

15
The module for light beams transmission is a rectangular duct. Rectangular mirrors located in
the duct corners provide the optical pathway in a required direction.
The system of light locks provides an absolute isolation of the optical channel from external
light.
2.3. Control and Display System
The control and display system comprises a push-button panel for manual control of the
thermal block movement and the LCD screen indicating the process of amplification program
running (Fig. 4).
Push-button of on-screen menu displays functions of other panel push-buttons within the
given operation mode. The push-button of thermal block control (the second one from the
left) is intended for launching the following operations:
opening and closing of the thermal block for placing the tubes into the thermal plate
before PCR running;
precise positioning of the thermal block under optical elements in their operating
position.
When the instrument operates under computer control, one can open or close the thermal
block using and buttons located under the button
in the Running window of RealTime_PCR
software.
LCD screen displays information about an amplification program and the running process.
It shows the following information:
operator’s name;
date of the amplification program start;
Plate protocol number;
name of the amplification program;
instrument serial number;
quantity of conducted measurements;
amplification program timer;
instrument status on completion of an amplification program;
current temperature of the reaction mix;
description of the amplification program;
fluorescence measurement channels enabled;
optical measurement indicator;
indicator of the amplification program run status.

16
Fig.4. LCD panel
2.4. Marking. Symbols used for marking.
labeling of the instrument meets the requirements of the following standards: EN ISO 18113-
1:2011, EN ISO 18113-3:2011, EN 61010-1:2010, EN 61010-2-101:2018.
Below provided example of the label on the rear panel of the instrument:
The following warning signs “Fragile”, “Top”, “Keep dry”, “Stacking is forbidden” are
placed on the shipping box as well as:
trade mark or information about manufacturer;
brand name;
date of production;
net weight;
gross weight;
requirements for transportation and storage.

17
Symbols used for marking:
For in vitro diagnosis only
Storage requirements (temterature)
« Fragile »
«Top»
«Keep dry »
«Stacking is forbidden »
Date of production
Manufacturer details
High voltage

18
3. Setting up the instrument
3.1. Instrument Unpack
DTprime real time PCR instrument is delivered packaged in a cardboard or wooden box with
inserts of polyethylene foam; the package is intended to protect the instrument from
mechanical damage when shipped or stored.
Before being packaged in the box, the instrument is placed to a special bag of durable and
waterproof material with zip fastener and transportation straps.
Caution! Heavy! The instrument has to be carried to the installation’s place in the
manufacturer’s package box by two persons.
Before taking the instrument out of the package box, perform the following procedures:
provide a vacant space on a work table for the instrument;
place the box with the instrument beside the table and open it;
take the components out of the box, and remove the polyethylene foam inserts;
compare the components with the delivery set (refer to item 1.2);
observing all the precautions, lift the instrument by two persons using transportation
straps and place it on a table;
remove the bag and inspect the instrument visually for external damages.
If any items are missing or damaged, contact your local “DNA-Technology” distributor or
“DNA-Technology” directly.
3.1.1. Instrument Carrying
During operation, the instrument may be carried over a short distance within the facility in
unpacked condition by two persons or using movable table (trolley) to other preliminary
prepared working place in accordance with corresponding precautions.
Note! In case the instrument is transported over long distances or shipped by a carrier, it has
to be placed into the original manufacturer’s package.
Caution! When instrument is moved the thermal block has to be in closed position.
Otherwise, in case of some problem with instrument please contact service department of
local distributor or “DNA-Technology” directly.
3.2. Instrument installation
Caution! The instrument has precision mechanical elements. To avoid displacement of them
any shocks during operation and movement of the instrument should be eliminated.
When selecting a place for the instrument’s installation, leave about 18 cm of space in the
front and 12 cm in the all others sides of the instrument for sufficient ventilation and free
access to the thermal block and power switch.
Plug the power cord into the back of the instrument and into an appropriate electrical outlet.
Plug the USB 2.0 cable into the back of the instrument and into an appropriate PC USB port.
Press the power switch on the back panel of the thermal cycler to start the system.
For a complete power off the instrument must be disconnected from the power outlet and PC.
The instrument does not require any additional devices to stabilize circuit voltage. When
plugging up the instrument to the devices of uninterruptable power supply, please note that
provided power must be not lower than 550W.

19
Prior to connecting the instrument to the supply voltage, check the presence of protective
grounding in the socket to which the instrument will be hooked up as well as integrity of the
connecting cable.
Caution! Hooking up to the socket without grounding is prohibited.
It is important that the instrument be correctly grounded. The instrument is supplied with
power cord that provides the instrument grounding, when connected to an appropriate power
outlet.
Also it is important that the instrument and PC must be connected to the same grounded
power outlet. Otherwise, there is a risk of the instrument damage.
Caution! After transport or storage under humid and cold conditions, dry out the instrument
(2-3 hrs) at indoor temperature (+18-25°С) before connecting it to the supply voltage. During
drying out the intrinsic protection may be impaired.
3.3. Software Installation
To control the instrument, software from the instrument’s delivery set should be installed on
PC. The software operates in Windows 98, Windows ME, Windows 2k/XP, Windows Vista,
Windows 7, Windows 8, Windows 10.
Software installation is required only if the instrument and PC are purchased separately. If the
computer is supplied as a part of the instrument set, the software has been already installed.
Caution! In case of purchasing the instrument without PC, any Windows operating system is
not provided.
Software installation procedure consists of two stages:
1. Instrument driver installation.
2. Installation of the software.
3.3.1. Instrument Driver Installation
Driver installation for the instrument is identical with one for any standard device and should
be performed as follows:
Switch on PC and wait until the operating system is launched.
Insert the USB flash drive with the drivers into the USB port of PC or download
drivers from DNA-technology official website at http://www.dna-technology.ru/eng/support/
Turn on the instrument using the power switch on the back panel.
The operating system indentifies a new device in 1-2 minutes (wait for message
on the computer screen) and start driver installation process using
the Found New Hardware Wizard (Fig. 5)

20
Caution! When installing the driver on the PC with Windows 7, Windows 8 or
Windows 10 operating system, the Found New Hardware Wizard window does
not appear automatically. Do the following to install the driver:
After connecting the instrument to the computer, open Device Manager window, then
right-click DT-964 and select Update driver in the list
In the next window Driver Updating select Manual driver search and installation,
click Browse, select Drivers_DT96x_W32_2014 folder (for 32-bit version) or
Drivers_DT96x_W64_2014 folder (for 64-bit version).
Click Next and complete the driver installation as described below.
Fig.5. Launching the Wizard for driver installation
Select point No, not this time and click