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View additional product information for 7500 Real-Time PCR System, desktop - FAQs (4351105)
48 product FAQs found
The 7500 software requires that the endogenous control be run on every plate within a study. If not, you will only see RQ values for the plate that contains the endogenous control on it. The other plates will have CT values for all the wells, but the software will not calculate RQ values.
If you designed your experiments in this type of layout, you will want to do your analysis in the free DataAssist software instead. This program does not have the endogenous control requirement for every plate. Simply export the study results file from the 7500 software as a *.txt or *.csv file, and use that to create a new study in the DataAssist software. You should now be able to see RQ values for all samples, even those without an endogenous control on the same plate.
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If you have the SDS v1.4 or earlier, you will need to open the dd CT plate in a Study first in order to see the CT or RQ values. (If you have v2.0.x this error does not apply.)
1.After the initial run is complete, open, review the plots and save it. Then:
-Choose File then New
-Choose: Assay then ddCT (Relative Quantitation) Study. Set the other parameters as appropriate and click Next.
2.Click on ‘Add Plates...', then browse to where your data file is located and select the plate (or multiple plates).
Click ‘Finish', and you should see your study.
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You may encounter this error message, which will abort the run before it even starts. If so, go into the software under Setup then Run Method and check how much time has been set for the data collection step. The instrument has a minimum collection time, which is dependent on the number of filters being used. (This is true in both the 1.x and 2.0.x versions of the 7500 software.) Increase the extension time to 30 sec and restart the run. It should now proceed as normal. If the error persists, please contact us at techsupport@thermofisher.com for further assistance.
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This error can be caused by the wrong plastics being used in the block, such as the wrong Precision Plate Holder being used in the tray. This can give an error message such as “Block up switch not activating” and will not allow the run to start. Open the drawer, make sure the correct plate/tubes and plate/tube holder are being used and in the correct orientation. Turn off the instrument and open the front cover to make sure the heated cover door is closed. Restart the instrument and retry the run. If the error still persists, please contact us at instrumentservices@thermofisher.com for further assistance.
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If you see this error message, please check the following:
1. Verify that the USB connection is secure between the instrument and computer.
2. Verify that the computer user power save settings are set to Never for all users (in Windows under Control Panel, Power Options).
3. Is there a screen saver in use? If so, set the screen saver so that it will not come on during a run.
4. Is there antivirus or other network software in use? If so, disconnect from the network while running the instrument.
If the error persists, please contact us at instrumentservices@thermofisher.com for further assistance.
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Sometime there is an error such that the ROI will not calibrate all 96 wells. If you run into this problem check the following:
- Is the ROI calibration plate old or expired?
- Spin down the plate and visually inspect the wells. Is the volume consistent and correct across all wells? If not, try a new ROI calibration plate.
- Try the manual calibration and increase the exposure time.
- Check the status of the halogen lamp. If it is over 2,000 hours, replace the bulb and try again.
If you have tried all the steps above but the ROI still will not calibrate, please contact us at instrumentservices@thermofisher.com for further assistance.
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You may get a message that the heated cover is not reaching the set temperature. If you get this error, or notice this issue on your own, please contact us at instrumentservices@thermofisher.com.
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“Fatal Error 1420” is related to the door or heated cover not being fully closed on the instrument itself. We recommend opening the front access door using a small screwdriver, ensuring that all inner components are correctly seated, then closing the door completely. Please then reboot the instrument and attempt to restart your run. Please see page 60 of the Installation and Maintenance Manual for more information: https://tools.thermofisher.com/content/sfs/manuals/cms_041437.pdf
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Click on the ‘Group By' button and choose ‘Well Position' (Column). Then highlight the plate and print the report.
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Go to ‘View Well Table' from the Amp B117Plot screen.
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Yes. If you have the newer version of the software (v2.0.1 or later), which creates *.eds files, your data will be directly compatible with our Protein Thermal Shift Software (https://www.thermofisher.com/order/catalog/product/4466038?ICID=search-product). If you have the older software (v 1.x), you will have to program the software differently (see below) and analyze the results independently. For more details on the analysis, you can refer to this paper: “The use of differential scanning fluorimetry to detect ligand interactions that promote protein stability.” Nat Protoc 2007;2(9):2212-21.
For an experiment using SDS v1.x, follow the directions below:
1.Create a new experiment, choosing Absolute Quantification assay type. Click ‘Next'.
2.Go to ‘Select Detectors'. If there is no predefined detector for this application, create a new one. Make sure the Reporter Dye is ROX, and Quencher Dye is none.
3.Assign the detector to the plate. Select none for the Passive Reference
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In a touchdown PCR experiment, you will either change the temperature or the time of a particular PCR step with every cycle. Most commonly, the annealing temperature is adjusted throughout the experiment, such that the specificity is increased in the early cycles and the efficiency in the later cycles.
In this example, we will set the method to do the following:
1. Go to File then New Experiment then Advanced Setup. Fill out the relevant options as you normally would.
2. Go to the Run Method under the ‘Setup' section and you should see the Graphical View of your thermal profile. Check the box next to ‘Enable AutoDelta'. You should see some grey triangles appear next to the Temperature and Time at every step in the Cycling Stage. (Note: If you want to start the changes at a later cycle, set this here under ‘Starting Cycles'.)
3. A new window called ‘AutoDelta Settings' will open up. Select the appropriate options. In this example we are decreasing the temperature by 0.4 degrees C per cycle, so choose (“-“) and (0.40). Click ‘Save Setting'. You will then see a green triangle show up next to the parameter you changed, in this case next to the 72 degrees C step. Your new method has now been applied
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Run files will be saved to a default folder on the connected computer, unless you change it. To find or change the default folder (in SDS v2.0.1 or later), go to Tools then Preferences then Defaults. Here you will see a Data Folder and an Import Folder. The default location is shown. If you want files to be saved to (or open from) a different location, click ‘Browse' and choose the new folder.
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No. The software will use the last data collection step in the cycling stage for all amplification plots and Ct analysis. So even if you were to set two separate steps with “Data Collection On”, you would only be able to view and analysis data from the latter step.
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The following volumes are supported for each instrument block:
-7500: 20-100 µL reactions
-7500 Fast: 10-30 µL reactions
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Using the instrument computer when a run is in progress is not recommended, as this poses a risk of corrupting the data.
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The 7500 and 7500 Fast Real-Time PCR Systems can be used to run assays designed with custom dyes (dyes not manufactured by Life Technologies). Custom dyes must fluoresce within the 520-650 nm spectral range measured by the 7500 or 7500 Fast instrument. To use a custom dye, you must first determine what the right concentration of the dye is. You will need to order an oligo with a 5' custom dye but no quencher. Make up a plate with different concentrations of this oligo (approximately 25-3,200 nM) and use the ROI Inspector to assess fluorescence. Choose the concentration that displays the brightest possible signal without saturation in all filters. Once you have found the correct concentration, create a full plate of custom dye at this concentration and perform the custom dye calibration. See Appendix B in the 7500 and 7500 Fast Real-Time PCR System Maintenance Guide (http://tools.thermofisher.com/content/sfs/manuals/cms_077749.pdf) for full details.
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The Applied Biosystems 7500 and 7500 Fast Real-Time PCR Systems use the following dye sets for calibration: Cy3, Cy5, FAM, JOE, NED, ROX, SYBR Green, TAMRA, Texas Red, and VIC dyes. Custom dyes that are read between 520 and 650 nm can also be used, although you will have to calibrate the system first for any new dye.
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Instrument Specifications
Block Options (Fixed): 7500: 96-well (standard); 7500F: 96-well (Fast)
Sensitivity: Down to 1 copy
Dynamic Range: 9 logs of linear dynamic range
Calibrated Dyes: FAM, SYBR, VIC, ROX, NED, TAMRA dyes (Cy3, Cy5, and Texas Red dyes - 7500F only)
Detection Method: SYBR dye, primer-probe detection
Resolution: Detect changes as little as 1.5-fold
Reaction Volume Range: 20-100 µL (7500); 10-30 µL (7500F)
Reaction Speed: Fast or standard (7500 has standard mode only)
Optics: Tungsten-halogen lamp, 5 excitation filters, 5 emission filters, CCD Camera
Temperature Range: 4-99.9 degrees C
Run Time: <2 hr (standard mode); ~35 min (Fast mode - 7500F only)
Temperature Accuracy: ±0.25 degrees C (between 35 degrees C and 95 degrees C, after 3 min)
Temperature Uniformity: ±0.5 degrees C (after 30 sec)
Thermal Cycling System: Peltier-based system
Available Applications: Gene expression, genotyping, copy number variation, HRM, protein thermal shift, protein detection, mutation detection, miRNA, presence/absence
Dimensions: 34 cm (W) x 45 cm (D) x 49 cm (H)
Weight: 34 kg (75 lb)
Remote Monitoring: No
On-Board Memory: No
Setup Configurations: PC-controlled only
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Follow the directions in the 7500 and 7500 Fast Real-Time PCR System Maintenance Guide, Chapter 6 (http://tools.thermofisher.com/content/sfs/manuals/cms_077749.pdf), to decontaminate the sample block. You can also watch this helpful video demonstration (https://www.youtube.com/watch?feature=player_embedded&v=dJyolVGhpjk).
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The RNaseP verification plate contains template, master mix, and a TaqMan assay for RNaseP. It is used to verify that the instrument is performing to specifications. If you have reason to suspect there is something wrong with the instrument, if the instrument has been moved, or if you want to rule out a chemistry issue, the RNaseP plate is a good way to test the system. The RNaseP verification plate is a single-use plate.
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The calibration plates can be stored and reused three times for up to 12 months after you first open them, so make sure to return them to their original packaging and return them to -20 degrees C storage until the next use. If needed, you can make your own background plate using deionized water. Please follow the directions in the 7500 and 7500 Fast Real-Time PCR System Maintenance Guide (Appendix C) (http://tools.thermofisher.com/content/sfs/manuals/cms_077749.pdf) for more details.
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Recommended Maintenance Schedule
Power on/off the computer controlling the instrument: Weekly
Check computer disk space. If necessary, archive or back up your experiment files and instrument settings: Weekly
Background calibration: Every month
Run disk cleanup and disk defragmentation: Every month
Perform an instrument self test: Every month
Pure dye calibrations: Every 6 months
ROI calibration: Every 6 months
Optical calibration: Every 6 months
RNaseP instrument verification: After the instrument has been moved, or as needed to verify instrument performance
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The minimum reaction volume supported by Applied Biosystems is 20 ul for Real-Time PCR runs and Allelic Discrimination plate reads on the ABI Prism 7300 and 7500 (Standard) Sequence Detection System. We have not validated the instruments with reactions volumes below 20 ul. We recommend you perform your own validation for comparable efficiency at lower reaction volume. On the Applied Biosystems 7500 Fast Real-Time PCR System instrument, you can run a minimum reaction volume of 10 ul reactions.
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The older versions of the software did not include this feature for all assay types. For the 7500 and 7500fast instruments, the latest software 2.0.5 allows a melt curve to be added to the PCR experiment. The other instruments/software require a separate program for melt curves. To run a Dissociation Curve after running a Relative Quantification Plate (ddCt) Assay, you will need to open a new assay from File -> New, select DISSOCIATION from the assay pull-down menu and run the Dissociation Curve as a separate assay. This will generate a separate SDS file for the Dissociation Run.
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The maximum thermal output for the Applied Biosystems 7500/7500 Fast Sequence Detection System is 3241.5 BTU/h (950W).
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The sample ramp rates for the 7500 instrument are as follows:
- Standard Mode: 1.6 deg C/sec up and 1.6 deg C/sec down
- 9600 emulation Mode: 0.8 deg C/sec up and 1.6 deg C/sec down. This matches the sample ramp rate achieved for the ABI PRISM 7700 Sequence Detection System.
- Fast Mode: 3.5 deg C/sec up and 3.5 deg C/sec down.
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With SDS 1.4 software, right-click on the graph, and choose "Export as JPEG" or "Export to Powerpoint". With SDS 2.0.x (for 7500/7500fast only), you can either click on the save icon above the graph "Save as JPEG Image" or click on the Export drop-down arrow (located on the upper toolbar) and choose "Send to Powerpoint"
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Yes, you can move the instrument carefully on your own. It is recommended that you recalibrate the instrument with the pure dye calibration kit(P/N 4349182, 4349180, 4351151, 4360788, or 4362201) and run an RNase P instrument verification plate (P/N 4351979 or 4350584), after the move.
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You can run an Applied Biosystems 7500 Fast Real-Time PCR System instrument in Standard mode as long the following set-up points are followed:
1) The reaction volume is between 10 ul-30 ul
2) The 96-well plate is a MicroAmp FAST 96 well reaction plate (P/N 4346907, 4346906 or 4366932)
3) Select the Run Mode to be “Standard 7500 “or “9600 Emulation” (2.0.x software versions only offer Standard or Fast - no 9600 emulation choice).
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All run files have an .SDS or .EDS extension. A Relative Quantification study will have an .SDM or .EDM extension and a template will have an .SDT or .EDT extension. The software for StepOnePlus, ViiA 7, and 7500 2.0.x software have .EDS, .EDT, or .EDM extensions; all other software have .SDS, .SDT, or .SDM extensions.
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The following dyes can be used as the reporter dye with MGB-NFQ as the quencher on the Applied Biosystems 7500 Real-time PCR System or Applied Biosystems 7500 Fast Real-Time PCR System when using TaqMan Gene Expression Master Mix (recommended for multiplexing) or any other Applied Biosystems TaqMan master mix: FAM, VIC, JOE, NED, TAMRA,CY3, ROX, TEXAS RED, CY5 dyes.
Of these dyes, we recommend a duplex reaction with FAM and VIC or FAM and NED dyes; if you are triplexing, we recommend FAM, VIC, and Cy5 or FAM, VIC, and TAMRA dyes. You must choose dyes that are well separated in emission wavelength. If you are using any probes with a TAMRA dye quencher, you cannot use NED, TAMRA, or CY3 dye as one of the reporter dyes in that multiplex reaction. ROX or TEXAS RED dyes can be used as reporter dyes only when not using an Applied Biosystems Master Mix.
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Please contact Technical Support for Real-Time PCR, and provide the serial number of your instrument. We will email you a copy of the user manual upon request.
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The error message is caused by a shifting of the heated cover. You can correct the problem by following this procedure:
(1) Shut down both the software and the instrument.
(2) Open the front cover by inserting and applying force with a pipette tip or a small Allen wrench on the right side of the front cover.
(3) Inside of the machine, you will see a metal plate with a plastic handle. This metal plate attaches to the heated cover. Pull the handle toward the front of the instrument and then close the front cover.
To prevent this from happening again, make sure you push the PCR plate completely down into the precision plate holder. Also, do not use compression pads on 7300, 7500, and 7500 Fast Real-Time PCR Systems.
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It is not possible to program a longer-term “infinity” hold on the Applied Biosystems 7500 Real Time PCR System. However, you can program a hold for up to 999 minutes and 59 seconds, which should be sufficient in most experiments.
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Yes, you can set this as an option. On the Detector Manager of the Applied Biosystems 7500 or 7500 Fast Real-Time PCR System, simply set the Quencher to "Non Fluorescent".
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Yes. you can choose either standard or fast ramp on the 7500 Fast System for software versions 2x or greater, and a 9600 emulation thermal cycling mode is available for customers who wish to run assays based on adjusted ramp rate. However, you should be aware that we cannot guarantee identical performance to the same modes run on a standard 7500 system block.
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Runs can be compared between standard and fast mode, as the data collected are collected in all five filters in the instrument. However, keep in mind that these data may not always show identical results and may have added variability due to the data collection processes and differences in ramp rate between standard and fast mode.
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Yes, older 7500 or 7500 fast data files are compatible with the current version of the SDS software. However, there is one exception: please note that the SDS v1.3 software will not allow RQ Studies to be created from RQ Plates from different instrument types, different thermal modes (i.e., fast and standard modes) or different numbers of thermal cycles.
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Yes, you can purchase the optional Applied Biosystems 7500 Fast Upgrade kit. However, this upgrade kit must be installed by a qualified Applied Biosystems service engineer.
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The 7500 Fast Real-Time PCR System contains a new high-speed thermal cycling block designed specifically to work with Optical 96-well Fast thermal cycling plates (Cat. No. 4346906) and TaqMan Fast Universal PCR Master Mix (Cat. No. 4352042). It also has a new precision plate holder design, for use only with Optical 96-well Fast thermal cycling plates, and looks slightly different with a front bezel and a darker tray drawer color.
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No, MicroAmp Optical Cover Compression Pads should not be used on the Applied Biosystems 7500 Real-Time PCR System.
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Yes, the standard 7500 instrument has a 96 x 0.2 mL block, whereas the 7500 Fast instrument has a 96 x 0.1 mL block.
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All SDS files generated on an ABI PRISM 7000, Applied Biosystems 7300 or 7500 or 7900HT Fast Real Time PCR System have a .sds extension (for example, test101.sds). If your computer has more than one version of SDS software, and you double click on the file icon to open the file, it will be opened from the last SDS software version installed. To work around this, simply launch the appropriate version of SDS software, and open the file from within the software.
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We recommend using the same thermal profile as listed for the 7500 and 7500 Fast systems (Table 8) in the TaqMan® Advanced miRNA Assays User Guide: https://assets.thermofisher.com/TFS-Assets%2FLSG%2Fmanuals%2F100027897_TaqManAdv_miRNA_Assays_UG.pdf
It is possible to import a standard curve into the Design & Analysis Software (v2.x) if you are analyzing results from one of the following Applied Biosystems real-time PCR instruments:
• 7900HT Fast Real-Time PCR System
• 7500/7500 Fast Real-Time PCR System
• StepOne/StepOne Plus Real-Time PCR System
• ViiA 7 Real Time PCR System
• QuantStudio 1 Real-Time PCR System
• QuantStudio 3 & 5 Real-Time PCR System
• QuantStudio 6 & 7 Flex Real-Time PCR System
• QuantStudio 6 & 7 Pro Real-Time PCR Systems
• QuantStudio 12K Flex Real-Time PCR system
Follow the instructions below to import a standard curve for analysis in the Design & Analysis Software (v2.x):
1. Open the data file in the Design & Analysis Software (v2.x).
2. Click on Actions.
3. Click on "Standard Curve Analysis Setting".
4. Select "External Standard Curves" and click on "Import".
5. Browse to the proper .csv file and click "Open".
6. Click "Apply".
There are two main differences between a Fast 96-well and standard 96-well block for the Applied Biosystems real-time PCR systems:
• Reaction well volume: The Fast 96-well blocks have 0.1 mL reaction wells and are compatible with 0.1 mL plates and tubes. The standard 96-well blocks have 0.2 mL reaction wells and are compatible with 0.2 mL plates and tubes.
• Ramp rate: The fast 96-well blocks have a higher maximum ramp rate than standard blocks for the same real-time PCR system. However, both the fast 96-well and standard 96-well blocks are capable of running standard and fast chemistry.
Note: For newer instruments (QuantStudio 3 and 5 Real-Time PCR Systems and later) "0.1 mL" and "0.2 mL" are being used in place of "fast" and "standard" as designations for the two 96-well block formats.
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All our real-time PCR instruments are compatible with Thermo Fisher Cloud Apps except the 7500 Fast instrument where the eds files alone are compatible with Thermo Fisher Cloud Apps.
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