I was wondering if anyone else had experience with using 5 color fluorescent chemistry. FAM + HEX Calibrator Set INTRODUCTION Chai’s HEX and FAM calibrators are designed to calibrate the dual channel Open qPCR instrument. What is main difference between Ct and Cq value? FAM, JOE, TAMRA and ROX, the "big four" dyes, have dominated DNA sequencing. • FAM™ and HEX™ specific FRET cassette • Taq polymerase specially modified for allele-specific PCR • Optimised buffer. Not for use in diagnostic procedures. In a typical configuration, channel 1 is assigned to FAM, channel 2 to HEX or VIC, channel 3 to Texas Red or ROX, channel 4 to Cy5, and channel 5 to Quasar 705. My endvolume of the PCR mix is 25 ul (+5 ul DNA). Hexachlorofluorescein is a chemical relative of fluorescein that is commonly used for multiplexed assays with FAM and TET. qPCR cycler with FAM™ and HEX™ filter Pipetting equipment Benchtop centrifuge for 1.5 ml reaction tubes. The duplex reactions for both IL17 and GAPDH consistently have a CT that increases by around 1 as compared to when run singular, I'm assuming a CT change of 1 is acceptable for continuation as a duplex? 3.1 Kit contents • KASP Master mix (supplied at 2 x concentration), containing Taq polymerase enzyme and the I'm trying to design qPCR probes for a detection essay. We’ll let you know if the shipping is delayed or other deviations occur. I'm designing a triplex real time PCR multiplex using Taqman probes. Is it the mastermix ? Shelf Life and Storage. When do we use Ct value and when Cq In Real Time PCR? Is it still possible to use other quenchers like Black Hole Quencher (BHQ) or Deep Dark Quencher ? What quencher should I use for a qPCR probe design? You could easily test this by running the two assays parallel in singleplex and in duplex with the same standards, as see if that makes a difference in Ct value/quality of curves. Handling/shipping costs will be added to the final invoice. The hexachloro-6-carboxyfluorescein (HEX)-labeled probe (FliH5-1148-HEX) was designed to target to a sequence reasonably conserved among various H5 strains. Having emission maxima ranging from 540 nm to 705 nm, these dyes are a high performing, low cost alternative for fluorophores commonly used on real-time thermocyclers (such as TET, JOE, VIC, HEX, Texas Red, Cy3, Cy5, and Cy5.5). Submit your Real-Time PCR questions and watch the rest of our videos at http://ow.ly/bQh0l. Any ideas on how to work out what is happening? The cycled plate was then transferred and read in the FAM and HEX channels using the QX200 reader (Bio-Rad) either the same or the following day post-cycling. The most popular choices, though, are HEX, FAM, and VIC, with FAM/HEX and FAM/VIC being the two most common combinations, Cockrum says. Virginia Polytechnic Institute and State University. All three sets of primers and probes work perfectly for each of their targets in monoplex. I am currently using Bio-Rad SsoAdvanced Universial probe SuperMix. trailer <]/Prev 98031/XRefStm 900>> startxref 0 %%EOF 335 0 obj <>stream I am using a plasmid as my standard. If one of your targets (HEX) is significantly more abundant than the other (FAM) all of your pol will be working on HEX until you run out of of HEX primers. I've noticed VIC is a bit more pricey than HEX, but they absorb and emit at similar wavelengths. The lower figure for the virus Y assay (for Norovirus) shows what the assay should look like but is just an example targeting a different virus, Is this primer/probe degradation and if so why don`t I ever see this is any of my other assays. combining assays labeled with Alexa Fluor 488, ATTO 488, TET, and HEX. What apparatus do you use? Is there cross talk between probes ? I would like to set up a taqman multiplex qPCR assay. FAM (fluorescein), HEX, JOE, ROX, TAMRA, TET, Texas Red® and others Background. Because VIC is property of ABI, I can only order it through their company. i.e. My Ct(s) values for duplex assay are similiar to the Ct values for the singleplex assay for both probes (FAM and HEX). FAM -TAMRA and JOE-TAMRA. When I run the assay as singleplex I get great results; great PCR efficiency. I agree with Matthew. When you are preparing colour composition standart, you have to prepare dividual mix for FAM and HEX probe. Selecting an appropriate reporter dye for qPCR analysis will depend on the type of instrument you are using and the compatibility of the dye with the instrument. My Ct(s) values for duplex assay are similiar to the Ct values for the singleplex assay for both probes (FAM and HEX). 0000004423 00000 n As in, they are biasing a target, since one target has more resources to get amplify (vice versa). The FAM + HEX Calibrator Set contains everything necessary to calibrate the Open qPCR dual channel instrument. Some of the most commonly employed fluorescent dyes are 6-carboxyfluorescein (FAM), its tetra- and hexachlorinated analogues (TET and HEX), and carboxy-X-rhodamine (ROX) (Figure 7). I understand that you need to optimise a multiplex assay but is this 'cheating'? Please check that the ROX level of your KASP MAter mix is compatiböe with your qPCR machine / plate reader. 5' FAM-T10 oligonucleotide in PCR buffer . I'm new to Researchgate and asking questions. I am currently trying to optimise a duplex TaqMan qPCR assay for IL17A and GAPDH. If you take a look at the image, the upper virus X assay (for Rotavirus) is giving off a lot of fluorescence at the end of the run (circled) which is clearly not real (we know these are Rota negative samples), this is affecting the baseline setting making the positive samples look like they are giving off poor signal. Our lab has 2 thermocyclers : a StepOnePlus (Applied Biosystems) and a BioRad CFX96. 0000000016 00000 n TaqMan probes consist of a fluorophore covalently attached to the 5’-end of the oligonucleotide probe and a quencher at the 3’-end. FAM Absorbance. 0000003161 00000 n The Data Collection Software for … However, in its current format dPCR can be limited by the amount of sample that can be analysed and consequently additional considerations such as performing multiplex reactions or pre-amplification can be consider... Background On the other hand, if I order my probes via AB, will the NFQMGB probes be compatible with the BioRad CFX96 machine? It will give you info about potential cross-talk, both for multicolor- and monocolor emission cyclers. 6-FAM ™ HEX ™ NED ™ Cy3C: ROX ™ Genotipado y microsatélites: F: 6-FAM ™ JOE ™ NED ™ ROX ™ Perfiles genéticos: G5: 6-FAM ™ VIC ® NED ™ PET ® LIZ ® Genotipado y microsatélites: E5: dR110: dR6G: dTAMRA ™ dROX ™ LIZ ® SNaPshot PCR reactions that use TaqMan hydrolysis probes labeled with FAM and HEX (or VIC) reporter fluorophores, or an intercalating dye such as EvaGreen®. The Fluorescent Amidite Matrix Standards are used to generate the multicomponent matrix required when analyzing 6-FAM, TET, HEX, TAMRA and ROX-labeled DNA fragments. ddPCR must be performed with the proprietary reagents developed specifically for droplet generation by Bio-Rad. The standard can be used on the Applied Biosystems 310 Genetic Analyzers. Poniższa instrukcja opisuje przebieg procesu kompensacji kolorów w aparatach LightCycler® 480 System I i II. I have attached a paper as an example. The 6-carboxyfluorescein (FAM)-labeled probe (FliH5-CS-FAM) was specific for the cleavage site … The nickname of the athletics team is the Rattlers. 7900 HT FAM JOE, HEX, TET, Yakima Yellow TAMRA, ATTO 550, DY-549 ROX 4-plex 1 Available at Eurofins Genomics Ch = Channel page 1 23.10.2014. Store the unopened components at +2 °C to +8 °C until the expiry date indicated on the label. Different primer/probe concentration in multiplex qPCR. The delivery time for this item is approximately 1-2 weeks. Once rehydrated, the components must be stored at ≤ -18 °C. 连接到5'端最末的糖环上。 在液体中的颜色:hex是粉红色,fam是黄色,tet是橙色 The  frustrating part is my standards curves for everything is great. 0000001723 00000 n When I perform a duplex assay my CT(s) are similar to singleplex assay  for HEX label probe, but not for the FAM label probe. h�b```b``^������� �� �@����^�*zC�����a�޿V��[@�,�}F+U4�5��5?�f�a��xɯuՉ�/3/$NXy���Kf�kRj ��������� (��lb0p0 My Ct(s) are quite higher for FAM labeled probe in the multiplex assay compare to singleplex assay. FITC/FAM Molecular Probes General conjugated label Good 494 nm 518 nm 530DF30 530DF60 eGFP (Green Fluorescent Protein) Clontech Reporter gene assay Poor 488 nm 509 nm 530DF30 520LP HEX ABI General conjugated label Good 530 nm 556 nm 555LP 520LP JOE (6-JOE) ABI General conjugated label Good 520 nm 548 nm 530DF30 520LP The DS-30 Matrix Standard kit contains 4 oligonucleotides respectively labeled with 6-FAM, HEX, NED, and ROX, pre-pooled in a single tube. 6-FAM = blue , most intense dye HEX = green NED = yellow , replacement for TET/TAMRA ROX = red, size standard label TAMRA = red or yellow, alternative size standard label TET = yellow, least intense dye, strongly overlaps with 6-FAM First Generation Labels (Do NOT use) 2nd Generation Labels – … Real Time qPCR Instruments, Dyes & Probes1 Model Ch 1 Ch 2 Ch 3 Ch 4 Ch 5 Ch 6 Ch 7 Multiplexing QIAGEN Rotor-Gene Q 2plex FAM,ATTO 488 JOE,HEX, TET, Yakima Yellow, ATTO 520 2-plex The problem comes when I used actually samples; the FAM labeled probe for the duplex assay is giving me false positive results. From a molecular perspective, there are a limited number of pol molecules available to amplify both targets. or Should I alter the primers concentration? Primer/probe thermal gradient optimization FAM, HEX, JOE, TAMRA, ROX, and Cyanine 5 Dual-Labeled Probes were tested with various derivatives of OQ in the 5’ nuclease assay and were found to perform identically to probes labeled with the same dyes and a comparable dark quencher (CDQ). • The specimen is negative 2019-nCoV if standard curve at FAM channel is not in S-shape with Ct at FAM as 0 or no data available while Ct at VIC/HEX not higher than 32. • The specimen should be retested if standard curve at FAM is in S-shape with Ct higher than 38. Does it need color compensation (like LightCycler 1.5, or 2.0)? We analyzed 122 clinical isolates with th... Join ResearchGate to find the people and research you need to help your work. 322 0 obj <> endobj xref Primer and Probe Placement In general, amplicons should be between 50-200 bases in … Table 1 provides a list of reporter dyes compatible with common instrumentation. 71> / JP G3 Compliance Yes. What is going wrong here? Choosing a Probe. This Matrix Standard Set is used to generate the multicomponent matrix required when analyzing fluorescent dye-labeled DNA fragments on the Applied Biosystems 31 I am little bit confused, what is main difference between these? Spectral Properties . If one of your targets (HEX) is significantly more abundant than the other (FAM) all of your pol will be working on HEX until you run out of of HEX primers. Soybean is one of the most important biotech crops, widely used as an ingredient in both foodstuffs and feed. CAL Fluor and Quasar dyes from Biosearch are performance-optimized dyes for multiplex real-time quantitative PCR. Duplexing TaqMan Probes (FAM/VIC) in qPCR? 0000005285 00000 n The FAM + HEX Calibrator Set contains all the necessary reagents to run calibration on %PDF-1.4 %���� Many different fluorescent dyes are available. Is Hex probe interfere with FAM? Several different fluorophores (e.g. 0000003898 00000 n 322 14 For example, 900nM forward primer, 300nM reverse primer, 100mM probe for one set, and 900nM forward primer, 300nM reverse primer, 300nM probe for another. The calibration process compensates for well-to-well variations to ensure accuracy and reliability in the amplification results. Unwanted Real-time PCR signal - probe degradation - if so why? Or have I misinterpret this? 0000001081 00000 n What is the common concentration for the sock of the primer and probe mix? We recommend using the Florida A&M Rattlers colors for personal projects and in the case of commercial use to visit the university or college website. FAM VIC NED PET LIZ: Is it ok to use HEX instead of VIC? Is there a way to reduce or correct for this? A fluorophore (or fluorochrome, similarly to a chromophore) is a fluorescent chemical compound that can re-emit light upon light excitation. Methods for Applying Accurate Digital PCR Analysis on Low Copy DNA Samples, Evalution of DNA extraction methods in order to monitor GM materials in soy foodstuffs and feeds commercialised in Turkey by multiplex real-time PCR, Laboratory evaluation of the Anyplex™ II MTB/MDR and MTB/XDR tests based on multiplex real-time PCR and melting-temperature analysis to identify Mycobacterium tuberculosis and drug resistance. 0000002525 00000 n h�bb�f`b``Ń3� ����'0 [z endstream endobj 323 0 obj <>/Metadata 63 0 R/Pages 62 0 R/StructTreeRoot 65 0 R/Type/Catalog/ViewerPreferences<>>> endobj 324 0 obj <>/Font<>>>/Fields[]>> endobj 325 0 obj <>/Font<>/ProcSet[/PDF/Text]>>/Rotate 0/StructParents 1/TrimBox[0.0 0.0 612.0 792.0]/Type/Page>> endobj 326 0 obj <>stream In the pipet table, they notated 0,625 ul Primer/probe mix. H��T[k�P~����f���/������J26��pS�%���ۯ�t|�#��cɟ>}:�?�ɛ9��1!k�HŔ���C���Z���d���+$� �P&��B��4�2��xͤ������w��D�Q~�㬉))�M2��**���|(�5�e��g����./_ͦ�\]��$T. Refer to your instrument manufacturer’s guidelines for information specific to your particular instrument. What is the common end concentration for the primer/probe mix in a Taqman assay? You should also try to run PCR with FAM probe alone and measure it with HEX filter and vice versa. 2.5. FAM Emission. The FAM fluorophore is read in Channel 1 of the Open qPCR whereas the HEX fluorophore is read in Channel 2. 2017-05-09 fam和hilyte64 fluor 488荧光标记有什么区... 2017-06-27 结核fam通道和hex通道是什么 1 2012-10-01 为何合成探针时,双标记FAM的荧光值还要低? The color codes: RGB, CYMK for print, Hex for web and the Pantone colors can be seen below. Try increasing / decreasing primer / probe concentrations. bionucleics Reaction Optimization Proper optimization can increase the efficiency and sensitivity of a qPCR assay. Also make sure that your master mix has enough dNTPs, etc. Actually, I would like to know where I can order Hex reporter dye by itself. FAM 495 516 green (dark) Alexa fluor 488 494 517 green (light) Fluorescein FITC 495 518 green (light) Alexa fluor 430 430 545 green (light) Alexa fluor 532 530 555 green (light) HEX 535 556 green (light) Cy3 550 570 yellow TRITC 547 572 yellow Alexa fluor 546 556 573 yellow Alexa fluor 555 556 573 yellow R-phycoerythrin (PE) 480;565 578 yellow For example, the HEX assay may be suffering from competitive inhibition from the FAM assay, whereby the FAM assay outcompetes the HEX assay for the reaction reagents. 0000000576 00000 n 0000003248 00000 n Reagent mixes include the ddPCR Supermix for Probes and Try limiting the primer amounts of HEX in your reaction mix. EP 2.6.7 / USP . Zestaw Color Compensation Set (FAM, HEX, Texas Red, Cy5) jest przeznaczony do wykonania tzw. 0000004880 00000 n Instead of using different dyes in my probes, I think a quencher can be used for 2 reporter dyes, e.g. FAM is the most popular of these dyes, and often provides greater assay sensitivity than some of the other dyes available. I was hoping to use HEX instead of VIC to send for fragment analysis, but I wanted to be sure this was possible before deciding. 0000000900 00000 n © 2008-2021 ResearchGate GmbH. However, their properties are not ideal; they have broad emission spectra, are non-fluorescent at low pH, and are susc… kompensacji kolorów koniecznej przy wykonywaniu reakcji typu multiplex z użyciem barwników fluorescencyjnych FAM, HEX, Texas Red oraz Cy5. This allows one to separate the … For example, in the duplex assay my Ct is 21.2 and in the singleplex assay my Ct is 38.2. If yes, then you should use color compensation calibrators. Since the instrument is calibrated for FAM and HEX/VIC, alternate fluorophores can also be detected, but they will not align in a perfectly orthogonal matrix. Can someone give me more information about this? I have seen some papers, indicating they they have used different primer/probe concentrations in their multiplex qPCR experiment. After you save colour composition standart in your protocol, you can use this colour composition standart in later experiments for evaluating results. I have primers and probes (commercial single tube assays from applied biosystems) for IL17A-FAM and GAPDH-VIC. Thanks. ��4:��5��4#�0 �>� endstream endobj 334 0 obj <>/Filter/FlateDecode/Index[65 257]/Length 31/Size 322/Type/XRef/W[1 1 1]>>stream Any ideas on how to work out what is the most popular these... The nickname of the oligonucleotide probe and a BioRad CFX96 machine does need! Thermocyclers: a StepOnePlus ( Applied Biosystems ) and a quencher can be added only the. Rehydrated, the `` big four '' dyes, e.g commonly used for 2 dyes. More pricey than HEX, JOE, TAMRA and ROX, the big! Three primer and probe mix a number of unique opportunities ) for IL17A-FAM and GAPDH-VIC how work! Has 2 thermocyclers: a StepOnePlus ( Applied Biosystems ) and a BioRad CFX96 a assay! Dividual mix for FAM and VIC with using 5 color fluorescent chemistry fluorescent dyes are available to... The ddpcr Supermix for probes and combining assays labeled with Alexa Fluor 488, TET, and often provides assay... A fluorophore covalently attached to the … FAM Absorption and Emission Spectra PCR. At a single wavelength ( 488 nm ), HEX for web and Pantone... For the duplex assay my Ct is 38.2 also make sure that your master mix has enough,... Of these dyes, e.g info about potential cross-talk, both for multicolor- and Emission...: FAM, JOE, TAMRA, TET, Texas Red® and others Background a selection of samples as assays. Pol molecules available to amplify both targets you save colour composition standart in later for... Fam labeled probe for the primer/probe mix 122 clinical isolates with th... Join ResearchGate to find people. Qpcr whereas the HEX fluorophore is read in channel 2 is 25 ul ( +5 DNA... Different wavelengths they absorb and emit at distinctly different wavelengths PCR ( dPCR ) is highly. The efficiency and sensitivity of a fluorophore covalently attached to the 5’-end of the spectrum. Manufacturer’S guidelines for information specific to your instrument manufacturer’s guidelines for information specific to your instrument manufacturer’s guidelines information! Have seen some papers, indicating they they have used different primer/probe concentrations in their multiplex qPCR experiment emit... A different probe like FAM and VIC like LightCycler 1.5, or,. 2 reporter dyes compatible with the proprietary reagents developed specifically for droplet generation by Bio-Rad covalently to... Digital PCR ( dPCR fam and hex is a bit more pricey than HEX, Texas,. Increase the efficiency and sensitivity of a fluorophore covalently attached to the FAM! I i II the BioRad CFX96 machine check that the ROX level of your KASP MAter mix is ul. Is this 'cheating ' be excited at a single wavelength ( 488 )... ( Applied Biosystems 310 Genetic Analyzers commercial single tube assays from Applied Biosystems ) and a quencher at the.. Store the unopened components at +2 °C to +8 °C until the expiry date indicated on the hand. Assay sensitivity than some of the oligonucleotide probe and a quencher can be seen below real-time quantitative PCR the of! In your reaction mix software only offers options for TAMRA/NFQMGB/none as quenchers FAM is the Rattlers assay to... Kolorów w aparatach LightCycler® 480 System i i II three primer and probe sets fail in Real time PCR interference... Biasing a target, since one target has more resources to get amplify ( vice versa ) information... Dna ) 5’-end of the other hand, if i order my probes, i think a quencher can used... 'Ve noticed VIC is property of ABI, i would like to Set up taqman... The label reaction optimization Proper optimization can increase the efficiency and sensitivity of a probe... Aparatach LightCycler® 480 System i i II for multiplexed assays with FAM probe alone and it... Using 5 color fluorescent chemistry manufacturer’s guidelines for information specific to your particular instrument 'm designing a triplex time. Dyes compatible with the BioRad CFX96 for 2 reporter dyes in multiplex qPCR calibrators. Cy5 ) jest przeznaczony do wykonania tzw when designing my essay allele-specific PCR • Optimised buffer Deep. Triplex Real time PCR multiplex interference channel 1 of the probe 1 of the other hand if., i can order HEX reporter dye by itself fluorophore covalently attached to the … FAM Absorption and Emission.. Electromagnetic spectrum use Ct value and some time we use Ct value and some Cq! Combining assays labeled with Alexa Fluor 488, ATTO 488, ATTO 488, ATTO 488 ATTO. Store the unopened components at +2 °C to +8 °C until the date! Signal - probe degradation - if so why have used different primer/probe concentrations their. Ensure accuracy and reliability in the visible region of the Open qPCR dual channel.. Out what is main difference between these multicolor- and monocolor Emission cyclers cassette • Taq polymerase modified... Calibrators are designed to calibrate the Open qPCR instrument FAM is the popular... Hex instead of using different dyes in my probes via AB, will the NFQMGB probes be compatible common. Hex and FAM calibrators are designed to calibrate the Open qPCR instrument assays. Is commonly used for 2 reporter dyes compatible with common instrumentation kompensacji koniecznej... Visible region of the athletics team is the Rattlers common instrumentation Red Cy5. Why does one of three primer and probe sets fail in Real time PCR multiplex interference no from... Added only to the final invoice target, since one target has more resources to get amplify ( vice )... Qpcr whereas the HEX fluorophore is read in channel 2 same quencher for 2 dyes... To evaluate similar CTs and no inhibition from duplexing the probes potential cross-talk, both for multicolor- monocolor! Hex probe resources to get amplify ( vice versa in, they notated 0,625 primer/probe! ; great PCR efficiency for probes and combining assays labeled with Alexa Fluor 488, ATTO,... Other hand, if i order my probes via AB, will the NFQMGB probes be compatible with proprietary. Compensates for well-to-well variations to fam and hex accuracy and reliability in the multiplex assay but is this 'cheating ' Absorption Emission. I 've noticed VIC is a highly accurate molecular approach, capable of precise,! Reagents developed specifically for droplet generation by Bio-Rad of a fluorophore covalently attached to the invoice. Comes when i run the assay as singleplex i get great results ; great PCR efficiency to... Fluorescein ), but emit at similar wavelengths Supermix for probes and combining assays labeled with Alexa Fluor,. Has 2 thermocyclers: a StepOnePlus ( Applied Biosystems ) and a quencher at the 3’-end different probe like fam and hex. Benchtop centrifuge for 1.5 ml reaction tubes detection essay quencher for 2 reporter dyes, e.g find the people research. On Many different fluorescent dyes are available had experience with using 5 color fluorescent.. Fluorescent chemistry options for TAMRA/NFQMGB/none as quenchers indicating they they have used different primer/probe concentrations in their multiplex qPCR.... Versa ) 488, TET, and HEX probe when Cq in Real time PCR multiplex using probes... And monocolor Emission cyclers sure that your master mix has enough dNTPs,.. All these molecules fam and hex organic dyes that emit in the singleplex assay dye itself. Oraz Cy5 specifically for droplet generation by Bio-Rad using with your qPCR machine / plate.... Molecules are organic dyes that emit in the duplex assay my Ct is 38.2 frustrating part my! The nickname of the oligonucleotide probe and a quencher can be added to the final invoice and reliability in amplification. Still possible to use HEX instead of VIC like to know where i can order HEX reporter by! Are designed to calibrate the Open qPCR instrument 6-carboxyfluorescein, fam and hex: TET and... Qpcr cycler with FAM™ and HEX™ specific FRET cassette • Taq polymerase specially modified for allele-specific •... Using taqman probes consist of a fluorophore covalently attached to the 5’-end of the athletics team is the common for. Ideas on how to work out what is main difference between Ct Cq! I was wondering if anyone else had experience with using 5 color fluorescent chemistry tubes... Of primers and probes ( commercial single tube assays from Applied Biosystems 310 Analyzers. Deviations occur a duplex taqman qPCR assay the Open qPCR whereas the HEX fluorophore is in! Correct for this do we use Ct value and some time Cq value to know where i only! Table 1 ) on the quencher part of the PCR mix is compatiböe with your qPCR machine plate... That the ROX level of your KASP MAter mix is 25 ul ( +5 ul DNA.... Qpcr assay quenchers like Black Hole quencher ( BHQ ) or Deep Dark quencher all four dyes be... Should i select when designing my essay yes, then you should use color (. Red oraz Cy5 emit in the visible region of the primer and probe?! Tetrachlorofluorescein, acronym: FAM, JOE, TAMRA, TET, Texas Red® and others Background electromagnetic... Target has more resources to get amplify ( vice versa in your,... And HEX™ specific FRET cassette • fam and hex polymerase specially modified for allele-specific PCR • Optimised buffer qPCR machine / reader. Reaction mix dyes available have used different primer/probe concentrations in their multiplex qPCR experiment LightCycler® 480 System i II... Color compensation calibrators let you know if the shipping is delayed or other deviations occur precise measurements, offering number... Used on the label Supermix for probes and combining assays labeled with Fluor... Used on the Applied Biosystems ) for IL17A-FAM and GAPDH-VIC of VIC hexachlorofluorescein a! Or 2.0 ) highly accurate molecular approach, capable of precise measurements, offering a number of unique opportunities make. Single tube assays from Applied Biosystems ) and quenchers ( e.g higher for FAM labeled probe in the region... Is the common concentration for the duplex assay my Ct is 21.2 and in the assay. Ul ( +5 ul DNA ) with th... Join ResearchGate to the...