Thermo Fisher Q Exactive Focus Mass Spectrometer – Slides full screen

so today we’re going to hear about routine applications what’s called routine everyone knows that this not so much routine about routine so we’ll be talking obviously about food analysis and the clinical applications of mass spectrometry and the thing we’re going to talk to you about today is a completely new product called the cue executive focus this an instrument has been developed with routine applications in mind in terms of the performance being exceptional for the price point so we’ve obviously thought about the pricing as well when we’ve developed this system for you so we’re going to have a little change to the session because of organizational errors from thermo not from anyone else and the first talk will be from Olaf sniper Scheibner and he’ll be talking to you about food and clinical applications and we’re gonna invite Bertrand Russia to stand up who’s had an early access to the cue executive focus you’ll stand up and he’ll talk to you he doesn’t have slides again that’s how fall he’s gonna come up and talk to you about his experience just for one or two minutes on the cue exactly focus the very last slide is it’s basically a 3d barcode if anyone has those applications on their phone you can scan that now it seems to work from even far back and then you can get access to information very early on by signing up to this sort of mailing list so our first speaker Olaf will come and hopefully engage you interest you excite you about the key exact to focus applied a high-resolution accurate math system in the routine markets thank you Thank You Martin so what I want to show to you is what the new newly introduced gear exacta focus is and which application is we can run on that system with which results some of you maybe have seen ma chase introduction yesterday already here we will go about a couple of things a little more in detail and show applications of that system the cue attracted focus is based on the known qyx active platform with the three instruments we have so far QX active to accept is plus and QX active H F which always are development in yeah new new options we have and technologies are always stepping up with new features but of course these systems have a certain price point so in this case we were more focused on the question to generate and to create an instrument which is capable of all the work small molecule routine analysis has to do but on the other hand address the price point which is clearly a different one than on a high level research labs with different fundings of course and that’s result which we came up with so we have a qyx active instrument so quadrupole Orbitrap hybrid with the maximum resolution of 70,000 M over Z full width half maximum we have a mass range up to 2,000 M over Z we have the mass accuracy known from cubics active instruments so that’s less than 1 ppm with internal calibration less than 3 ppm with external calibration we have fast related to switching so one full cycle one positive spectrum switching one negative negative switching a positive spectrum and switching back inside less than one second on 35,000 resolution we have four data dependent analysis a maximum of top two and we introduced with this system absolute collision energy because this fits better for small molecule applications if you maybe use the q attractive systems already you know that on these systems you have normalize collision energy that works very nicely for pulley for peptide and protein work where the rather easy equation smaller mass smaller collision energy works very well but we all know that for small molecules we in enough cases have the exact opposite so that doesn’t didn’t fit so we switched over to absolute collision energy on that system so in that orbit in that q to focus we have some known Orbitrap detection we have two known quadrupole with hyperbolic rods we have the RF lens ion optics for great robustness and we have to know an HDD fragmentation very robust giving very fragment rich spectra for an unambiguous identification of compounds this is the schematics of the system you have the ions ion source and

ion optics here with the RF lens we have the quadrupole for precursor ion selection the C trap the Orbitrap and in the back the HD so that’s the known layered of all QX active instruments so what can we do with this instrument one example for high resolution and the need of high resolution in this case of course it is made up just to keep things under control we have two components being very close together and you see with 17,000 resolution you are not there you get a completely wrong mass on 35000 solution we get a sort of separation but this is definitely not yet suitable for routine application just little slip in there and you again don’t see that flank here and with 70,000 solution we clearly can separate these two components and society that’s made up of course it’s an example but look into the data of high-res instruments you will see that often enough it’s not necessarily two components you want to look at but you have often enough matrix ions being just as close which would completely destroy your analysis when they’re inside here and especially if you’re looking into a detection at the limit of detection this matrix signals often are bigger than the component you want to look at and you get a completely wrong quantitation even if you can detect it we run the system for five days injecting wheat matrix with a couple of components I took one out of here this is aflatoxin g2 and we are looking into the stability of the peak area integrated with our processing software and on the other hand looking at the mass accuracy of the whole time of 900 injections during Mallis five days and you see that the variation of the peak intensity is rather small we get a flat line here so we don’t get any decrease due to due to contamination and on the other hand for the mass accuracy we still stay inside 1 ppm + – 1 ppm window with internal calibration so the RMS value is 0.45 ppm of the whole time and we have on the intensity and our SD of nearly 12% so you can get additional warranty on that system that’s just the side mark here that it can be easily purchased with a long time warranty to give you more security that the system will do the work it’s intended to in your lab we have new instrument control software on that system just aside markets called 2.5 that’s not that important it’s software which is valid for all q x-axis exact of instrumentation but it’s specifically for the QX active focus that’s added in here so the scan modes we have so we cleaned up the instrument software a little bit for Q extractor focus for more easy operation so basically what you will find in the end in the method setup that is it that you find full scan you will find sim you will find PRM which is a targeted MS 2 mode and you will find full scan AAF and full scan VGA a this already directs you into the direction that on such a system the major application is running everything in full scan and using the ms2 data mainly for confirmation but the main detection is done in full screen so that’s the sort of shift operating if you come from triple quad that you don’t think and fragmentation anymore you detect a full scan of course the resolution in full scan is the prerequisite for that that you can unambiguously detect your components and in false can we have two options of confirmation one would be fullscreen confirmation which is the classical data dependent name is – you have an inclusion list the system is looking for specific masses that can be 10 that can be 2,000 no matter what and whenever such a mass comes up it will trigger one ms to spectrum so you have to fragment ions for a confirmation all your own run full scan discovery which is the top end experiment so the most abundant ions will be taken fragmented going on to an exclusion list you will check the next one the difference is of course you don’t get that deep into your into your components so the low-level components you will not attract most likely with such an scan mode that’s more for metabolite ID than for quantitation we have the same mode with up to 10 times more sensitivity for specific detection of a lower number of components if sensitivity and full scan should not be

enough PR I’m going the same way with added selectivity fall scan AAF that’s known from from all ex active instruments so far you have the full scan and the fragmentation of all ions together and we have the VBA a mode and on that I want to leverage a little bit more so VD a is data dependent data independent acquisition for a full digital record of your acquired data video a stands for variable data independent acquisition dead append data independent acquisition is known from proteomics where you try to mimic sort of the data dependent M is 2 and then unbiased matter in this case here for the small molecules is more the other way around we come from the AF mode which lacks sort of selectivity and by this way sensitivity slice the AR f windows to get higher selectivity and sensitivity so you can do up to 8 windows with the many more my relation with of $50 and so that’s the difference to classical daa applications so that’s what Sachin Skan Skan setup looks like so you have for example a full scan which covers the mass mass range from 100 to 1000 which is fine for most of small molecule applications and subsequently we have in this example five windows for fragmentation four of them having an isolation width of 100 dalton and the last one at the upper mass range has a wider windows so we don’t slice everything the same size but to save some time we give the upper mass or energy bigger window so in total we are here at a full cycle time of roughly 600 60 milliseconds so that’s basically still fine even to be compatible with fast chromatography if you run really ultra fast chromatography with one second speaks you still should think about something else but even with five seconds Peaks you can work with this very nicely and I will show you what’s the beauty of that so basically VJ AAA fills the gap between data dependent MS through acquisition and a AF conformational ms2 has still the best selectivity and sensitivity that’s well known but if you want to do anything beyond your very limited target acquisition you cannot do very much in this mode because you don’t have fragments for components you are not thinking about beforehand so you could detect a non full scan but that’s it discovery ms 2 as I already said very nice selectivity as well but you don’t get to the low abandon ions in most cases a AF has all the fragments all the time that’s basically what we want but since we have all the fragments inside you’re lacking sort of cell activity so confirmation in many cases fails where you need it so in the end that’s not appropriate for the same to do quantitation and qualitative work and the same data and by segmenting the era of video enhances the selectivity and dynamic range on this data so that’s an example if you look into the passage of the antibiotic oxacillin and pick his name so we get a nice in AF we get a very nice extracted iron chromatogram of the parent iron that’s clear full sky and high resolution everything’s nice we got a good calibration curves as well as well but here you see if you look into the fragments it’s an iron overlay of all the fragments available for that there is nothing specific behind it so in terms of regulations this would be a no detect because you cannot confirm you can see the iron here but you cannot confirm in terms of regulation you cannot say you have detected it the same sample the same instrument the same chromatography just run directly behind it in VGA mode you see again a nice extracted and chromatic form of the peak of course the calibration curve stays the same but on the fragments you see the huge difference here this is something which you can take for confirmation yes confirmation there you see at least three fragment ions together with the with the parent ion you see a nice core illusion here so everything’s fine you can go report that as detected and by this selectivity is higher and by this the sensitivity is enhanced because you can confirm to lower limit and as it comes out in the end if we do a comparison on the same instrument it compares very well with full scan data dependent detection so the limits of detection are basically the same on both modes so why use vta now if it is the same as full scan data panama soon as i said you can go back and look for other components even confirm them so the whole unknown screening with confirmation is possible here you can go back at every time and look at any component you want to look at which it didn’t have in mind

beforehand to aid us in that we created a new library specifically for QX active instruments so we have library on environmental and food safety clinical and toxicology monitoring done on different collision energies I will show you a couple of numbers in the next slide so for identification with high confidence very quickly for HRM high resolution accurate mass screening so we have the database created at the 140,000 resolution we have pesticides mycotoxin veterinary drugs environmental contaminants we have tracks of use therapeutic drugs we have multiple collision energies multiple spectra per component and we populated that with retention times and relative retention times which are accompanied already-made methods you get with the library so you can see which condition have been used if you do it the same way in your lab you even can use the retention hands from the library for a dafuk Asian so currently we have roughly 1600 EFS components and they are roughly 1100 clinical toxicology components so the whole thing is represented by roughly 13,000 spectra at the moment so that’s the current number it’s still counting we are in the end heading for something like 6,000 components in there in the latest stage so we’re still working on that take some time to acquire the spectra and make sure that the quality is fine so with time over the next month we will even increase these numbers and on the other hand what we have as an addition is the online repository MZ cloud which is free to everybody to look at and to search data in which at the moment contains 19,000 spectra in EFS 12000 spectra for a clinical toxicology and this is still counting and for the future this is even a user library so even users can contribute spectra from their own components put them there and use them for subsequent search for their own purpose ok some applications as I told you I want to show you what this instrument can do if you treat it under real conditions so for example for food environmental what we see here is vetrix screening pesticide screening non-targeted screening food safety testing water analysis just to name a couple of elephant’s we have already run on that instrument and for one example we had veterinary drugs in food samples using four can VTA a I don’t name all the details here that would take a little too long just for reference here and these are a couple of components basically the antibiotics used in this in this study and you see here the limits of detection the limits of connotation are fairly low there in most cases below 1 ppb and so they are way lower than the administrative regulations at the moment give us the limits this is even future safe so even if the limits get tighter they will still far away from from these detection limits here and we didn’t have only antibiotics in there we had evam Acton’s and imidazoles in there as well just to have a couple of different components which behave quite differently in chromatography and mass spectrometry but we had all these components in the same samples random on the same chromatographic method with the same instrument method so everything’s generic you just go shoot your samples and get the results you don’t need to optimize anything the system is not dependent on any component optimization the chromatographic method has been chosen that way that it should do for the vast majority of all the components to find in environmental and food safety applications and even clinical applications most of the time and this is one of the Epley one of the methods you will get together with the library on the system when you buy it and so this is really just put the samples onto the autosampler go away let the system do the work and come back and find it ready to processed and trace finer software which can look like that so we see all the components here we see all the samples you measure it we get the extracted and chromatogram for example to see the opponent ativ peak here and then we get the confirmation steps of course one confirmation step is the retention time here clearly but we can use for we can look into the isotopic pattern of the parent peak that comes for free with full scan analysis and since we know their nominal composition it’s easy to do a comparison of the theoretical and the practical

spectrum we can look for fragments the library contains up to five fragments per component which comes with the software so you have the fragment information you just need to click it on and it will do it for you and of course we have the calibration curve so linearity is given so this is a check mark this component is detected and confirmed go for it report it just a couple of examples ampicillin cephalonian from the antibiotics range time’ try does all metronidazole from the nature emitters all range just to give two different types of components here which still perfectly knee arity here we cover a range of five orders of magnitude here we can do up to five up to six orders of magnitude on the instrument but what as I said what you can do as well as a subsequent screening it can be a targeted screening just with a big list but you can do unknown screening as well matter of time is that I stopped here at the targeted screening weather will show you what I did here so I just applied a big list it contains two thousand components and here I looked into the isotopic pattern I looked into the fragment ions and I looked into the library as I said we have this newly created library so this is the day this is the base which are worth working on and just running it through the samples here and it just this is just one one example nicotinamide i picked here from the components i detected additionally and as you can see again we have an isotopic pattern match of 1% for fragments were searched for four fragments were found and the library as well gahafer gave a hit basically I don’t have another number here but it must be greater than 80 percent that was limit here so just to show it a little bit in detail extracted and chromatogram we have the isotopic pattern match you can see the overlay easily quality control that it really matches well and the fragment search result you see the fragments how well the match in intensity and mass security for all your components so one generic methods search for all components it’s easy to set up and operate no optimization needed we have to integrate it software doing all the data processing for you so you just can set up the samples go away and find in the end the printed report if you want and with variable dia we are sensitive selectively and we can do retrospective analysis on it another example were pesticides in beat metrics in this case run again of course on the cue x-axis focus this time with data dependent ms2 and again the library was used for confirmation as well these are the components we looked at in this case so a couple of pesticides here and the food matrix and if you look into the LOD is here a lot use here it’s all around 1 ppm with only few exceptions but look at the regulations here where the limits are we have 50 we have 100 ppb we are fairly below so there’s really future safe even if the regulations get tighter linearity in all cases very good here so no question that this is a valid quantitative approach which you can use for routine analysis very nicely again of course automatic processing you get this this traffic light system here to see if all your criteria have been matched you see again the quantitative Peaks to see the fragments and here we have an example for the library screen where this component in this case has been found to the score of 91% trace final offers you a lot of quality criteria which will flag immediately if a criterion like linear it is not mad at something like that to get an easy overview over a daily you don’t need to look into everything which can be very tedious with hundreds of components and hundreds of samples another example for the library scheme that’s what it looks like so you have with one-click easy access to that if you want really to prove that the match is what you expect it to be and that this is not sort of one-hit wonder on some random spikes okay so with full scan data dependent ms2 you can meet all the detection limits of EU regulations we can confirm very nicely using the new spectral library and the key focus can do all that easily so we had an early user of that that’s hands mold from the wrinkled Institute in bargaining and he said I consider the QX active focus as a fit for purpose alternative for pesticide residue analysis currently done using chocolate quadrupole instruments besides actually equivalent quantitative data for frequently detected components the

instrument provides a complimentary screen of many other components from the same injection on one instrument and this comes from his comparison with as well competitors instruments being high rest being triple quad and our own cue extracted family instruments he has as well and the QX active focused that’s as well in terms of selectivity insensitive as the other key exact instruments do and it is equally or better than the other instruments he has in-house of course we don’t run only food safety or environmental applications we look into clinical applications as well for example so we have to target our screening and urine using full scan data independent names to in this case with permanent polarity switching so we have the full scan on high-resolution a data dependence to on low resolution switching the polarity doing the same on negative polarity and back so what are the results are looking like we have 300 components positively and negatively ionized we have component spikes in in some sort of real-life samples and analyzed that yeah here from the real life samples from a collaborator so that’s that’s not only made up this is a real sample what does it look like if we look into the results so for fentanyl for example we can easily match the mo VZ accuracy extract that as in the other examples as well with 5 ppm mass security we have retention time we have the isotopic pattern which matches well and we can apply the library search so an easy confirmation here I said already the toxic collect toxicology library I mentioned that already and starts to repeat as you can see we again use trace finder we again use the same criteria as a different field of application but the way you’re on it is the same and you have the easy steps of confirmation here so we have the libraries that match again we have the isotopic pattern match again but looking into the these donor samples which are real life easy to detect phenobarbital fentanyl nor fentanyl and you see here that’s the point I want to draw you to this is the number of scans across the peak with polarity switching on such an application it’s not 1 second peak but it’s not a really wide peak and still we get lots of confirming to lots of quantitative scans here and lots of confirmatory scans as well which you can rely on for fragment confirmation and the second sample even contained a couple more so don’t say show the scans across the peak but we get nice Peaks here they are not somehow edgy because they only contain two scans or something like that so we get really the nice peak shape because we get enough scans across the peak despite the polarity switching ok what we can do as well is quantitative analysis of opiates and urine in this case we are using the PRM mode so this is a target that I was to mold we have an inclusion list with the precursors all precursors will be fragmented all the quantitative qualitative processing in the end of course works on the fragment side so it should be the most specific method our collection of high resolution MSM spectra for each analyte and the internal standards this is what such a method looks like we have an inclusion list with certain parameters like the retention time like the collision energy like the polarity in this case only positive what we could do it negativity as well it’s an easy setup you don’t have to think about many parameters here and so we have a wide calibration range 2 to 5000 nanogram per liter a milliliter a couple of QC samples and this is what what it looks like in the end for example the oxymorphone you get a very nice peak on the quantifying iron and you get enough qualifier ions for confirmation so again whatever the regulatory needs are you can meet them because it’s all a high-resolution detected like if you’re if you’re in need of some edification points this always gives you enough enough edit identification points and since we are not selecting the fragments in a q3 like on a tropic what we see always all fragments you even could use more fragments if you want these are only the most abundant fragments for confirmation here again let’s look into the scan speed of the instrument on the in that case you see again we get more than enough scans across the peak for connotations no worries that you need to

quantitate on a 3p representation that’s really enough that you get good peak shape and a reliable peak area of course we want to see that the instrument behaves linear the same way even in that matrix but you see no problem here it’s perfectly linear and what we have is well of course valid for the other applications as well but not just not yet mentioned that is that we can use that you of course can apply iron ratios this is automatically limit raised finer as well trace finer even can derive the iron ratios from a high concentrated standard it’s a one-click application for you to say okay just look under this standard file and get the iron ratios from there and apply it for the rest of to the rest of the analysis so that’s easily done you don’t need to put it in if you’re dealing with 500 components it could be very tedious trace when it does that for you so looking into the sensitivity of this approach here so you see a lock use 2.5 – ppb and knowing especially in drag analysis what the active concentrations are you need to have an effect we are way below of the active of the active concentrations so no problem to detect everything you need so and one of their customers using the new cue exactly focus that’s that humble shell you can see his quote here but I would like to invite him to come up here and give you a couple of words on his first experience on the instruments he had during the last two months yeah hello everybody again so I have this instrument only for a couple of months and I didn’t have time to produce data yet to present here today but I still work with cue exactly and executive plus instruments and as I said this morning I invite you to consider this kind of technology high resolution for for your routine analysis pushing that for the last few years already I think with this instrument this is a kind of silent revolution on one point the price I think I don’t know exactly the price but I think what they told me this is the same price in a triple quad right so the next pressure is I think you should consider to do a comparison side-by-side with triple quads and this type of instruments and and then look the data as I did three years ago with the first executive instruments generation are just compare triple quad data and and executive plus executive instrument at that time and I realize that was really comparable so I just invite you to actually send your samples to these application specialists and then make your mind of course with your own analysis and own own demands I don’t remember what I said but basically I think this is a trend as I said this morning a trend where MS makers will push for i resolution especially if has customers we are asking for it so a point the vendors are looking for what we need but if we don’t move they will not move so I think they understood that more and more people are pushing for that and of course they have to follow us right so that’s only what I can say today if you have questions you can ask me or other specialists in the field performing routine analysis whatever clinical or food safety and I think you will see that they all think the same basically alright many thanks but while to come up rather spontaneously and give us a little introduction little overview with our impression of what you did so far with that instrument okay so in total in the end we have as you can see a nice Lagoon universe around you exactly instruments and of course the cue exactly focus is integrative part of that one of that universe the software we are mainly dealing on with this instrument is tradesmen or software we are lots of examples we saw a lots of examples from trace finder we have just released yesterday newly released together with this instrument our new tox finder software some of you may have heard about tox ID which was a long time or simple to use software just for

screening results quick overview and tox Finder is the successor of that in the background it’s based on trace foreigner but much more simplified it’s for screening and relative Quan it’s not for absolute Quan but very easy to use very quick to use with the same databases in the background for the same way of confirming your results so as the concluding remark so this is the portfolio we have on the Q exact a family and you see it’s growing bigger and bigger with time we have the exact of plus-x active + EMR which is sort of the still the entrance instrument you have the q exact if in q x active + as our so far working horses at the q exact of h f as the top-tier instrument with the best performance of all and as a federal rating so far the development has always been up and of course up in price as well the QX active focus is the first one to go down from a certain point and when we say always affordable durable proven about durable and program i think i’ve given to you some examples here the affordability we left out so far here and of course this is not the place to give you numbers but i can tell you the next time you’re thinking about buying a new LC ms/ms system in your lab or a high-resolution instrument are namely acute off instrument you should consider this one definitely will fit on the pot into the budget you will need for the other types of instruments as well so we’re a little bit advanced in time already so i don’t want to go over this really in detail just an overview what can the different instruments do so as a conclusion from my side why should you buy now eq exactly focus so you have high up of higher performance analysis and the most affordable Orbitrap ever refined for reviewing laboratory experiments so the performance you need the software you need for processing it you can use it for routine qualitative and quantitative work with optional retrospective analysis two techniques and one platform so you can do whatever you want to do in a triple part and you get something extra for the same price it’s easy to use most method set up on an instrument like this is without any evaluation optimization you just go and shoot you have versatile processing software trace finder and toxifying there are only two examples of the software portfolio we have for this instrument if you need more sophisticated software ask us we have more sophisticated software and we have user-friendly daily operation this instrument is really designed for 24/7 operation and high throughput ok and so far thank you for your attention and maybe you have some questions for us so if there are no questions I guess I’ll remind everybody that you have a short break now before the next session so there’s plenty of time for coffee Haroon you have a question which a says yeah I don’t know that we’re gonna commercialize assays for antibiotics right I don’t know that we will all want this is honest answer but this yes yes for Microbiology absolutely so the software you have right now the trace finder will actually operate the instrument and will actually acquire the data will process the data and generate a standard format of report so you kind of have them multiplexed I say already but if you mean are we going to sell kits and so on I don’t know the answer yes so we saw Bertrand show in one of his slides ago today the first publications came out I think 2008 with executive products so there’s quite a long history now showing that the the

the metrics of a quantitative assay on these types of instruments are more or less exactly the same as the the performance metrics on triple quadrupole in some instances they’re better is compound specific sometimes the triple is better on a compound sometimes high-rez approach is better on a compound but on average over a wide range of compounds they’re almost identical okay I’m gonna get a coffee