How to save your samples before it’s too late!

Picture this: You’ve planned your experiment, collected all of your samples and you’re ready to run them on your LC-MS. Time is tight, so you get your samples through as quickly as possible, making the best use of the time available to you. Finally, it’s complete and your data is ready for analysis – great, so far so good. You start analysing your data in your preferred software — or simply what’s available to you — but after several painstaking hours of frustrating analysis, you’ve realised that something’s not quite right. After a bit of investigation, you find that your data just isn’t usable, due to sample running problems at the time of acquisition. Time to beg for some more time on that LC-MS… assuming you have some sample left, of course!

Sound familiar? Wish there was a quick and easy way of checking your data for problems while you’re still running samples, so that any issues could be identified and rectified quickly to prevent wasting time collecting “garbage” data? Here’s how Progenesis can help:

Quality control comes first in Progenesis

The first stage of the workflow in Progenesis is Import Data. Here, the raw data is imported and immediately visualised in the form of a 2D ion intensity map.

The ion map for a single LC-MS run viewed at the Import Data screen

It’s this ion map that can be used to quickly spot a multitude of common sample running problems. We recommend installing Progenesis on the acquisition PC and checking the quality of your data every set number of runs using the ion intensity map to prevent any nasty surprises during the analysis.

If you’re worried about needing an additional Progenesis licence for this QC, have no fear: importing data into Progenesis doesn’t require a licence, so you can–

Wait a second! That’s a really important point that’s worth repeating:

Importing data into Progenesis doesn’t require a licence.

This means you can install it on as many PCs as you like without fear of delaying anyone else’s analysis (or even having to make an initial purchase of the software!). You’ll also be pleased to hear that as you’re not using any of the more powerful analytical processes and algorithms involved later in the Progenesis workflow, you can carry out this QC step on a reasonably basic PC.

Don’t just take our word for it

If all of this sounds too good to be true, here’s what Dr Paul Langlais from the Research Division at the Mayo Clinic (Arizona, US) had to say about the early QC in Progenesis:

“Progenesis is label-free quantitative proteomics at its easiest. What surprised me is how much I was able to improve the quality of my mass spec runs, using the 2D map feature combined with the charge state distribution analysis…

What nobody realizes is how powerful Progenesis is in analysing LC and instrument method design. I’ve completely redesigned everything. I look at other people’s .RAW files and I see how much they could use the 2D feature alone.”

And when you consider that Progenesis supports data from a wide range of instrument vendors as well as several generic file formats, this makes it a suitable and important resource for pretty much any LC-MS lab.

Spread the word

Did you find this article helpful enough to share with other researchers like yourself? If so, click here to share the tip on Twitter – if we can save even one experiment together, it’ll be well worth it!

Back to basics – No missing values

Missing values

When writing the blog, it’s sometimes easy to get distracted by what’s new and exciting with a product rather than to focus on its core features and functionality. With this in mind, I thought I’d take us back to basics and talk about the “no missing values” approach taken by Progenesis – one of its core, and arguably most important, features.

Missing values can occur for a number of reasons:

  • Not present (the analyte truly isn’t in the sample)
  • Present but below threshold detection limits on the instrument
  • Present but missing due to instrument error
  • Analysis error: present but misaligned, misdetected or misidentified

Missing values in your data can cause a number of problems:

  • Reduced effectiveness of statistical analysis through lost data points
  • Misleading statistics – missing values can result in the data being misleadingly reported as statistically insignificant.
  • Problems with useful multivariate visualisations caused by the missing data

The chance of missing values increases with the number of replicates you run and since replicates are important in increasing the statistical power of your experiment, this poses a significant problem. There are a few approaches used to attempt to address this:

  • Imputation (assigning values for missing data) which must be done with care and carries a risk of biasing the data depending on why the values are missing
  • Removal of observations with missing values (increasing fidelity for the remaining measurements but potentially sacrificing useful intact data or introducing bias)
  • Interpolation (e.g. modelling missing parts of a peak from the rest)

Or the Progenesis approach: co-detection, using accurate alignment and combined aggregate detection to eliminate missing values

How does Progenesis do this?

While no software can perfectly restore raw data that are absent due to instrument error / inaccuracy, Progenesis does remove the possibility of missing values caused by misalignment, misdetection and misidentification, giving you a true reading for everything reaching the detector. It does this by accurately aligning all the runs in a dataset to a reference run on the retention time axis, with the reference run being the run with the greatest similarity to all other runs in the experiment.

Once the runs are aligned, a single aggregate run is created containing all analytes from all the runs in the experiment, with the retention time alignment correcting any drifts due to inconsistencies in the chromatography. Co-detection is then performed on this aggregate run with the detected isotope profiles being applied to all the individual aligned runs:

Co-detection

This ensures that like-for-like measurements are made for the same point of the intensity profile across all the runs, with the same boundary. Abundance is consistently and accurately measured for every analyte in every run, with a value of 0 only being reported where there is truly no signal present above background. This approach also allows identifications to be confidently aggregated and passed across all runs for the same analytes. This aggregation of MS2 data allows more confident results and also eliminates missing identifications as information in one run can be applied to all runs.

If you’d like to try the Progenesis approach to no missing values with your own proteomics or small molecules LC-MS data, get in touch and we’ll arrange a demo.

“Making decisions is easy. Getting them right is the hard part.”

A phrase I’m sure we can all relate to, and one that’s got me thinking about the way we make decisions here at Nonlinear – decisions that affect our users and, ultimately, the way they analyse their data. So, how do we ensure we make the right decisions?

Listening to our customers

We regularly conduct surveys for both Progenesis QI and Progenesis QI for proteomics, asking about the top 3 challenges people face while using the software. So far, we’ve had great responses to these and they’ve given us valuable insight into anything we’re missing or just not doing quite right. While we can’t guarantee to implement all the suggestions we receive, every single one of them is reviewed by the development team, and we do our best to ensure we use these suggestions to improve the “Progenesis experience” for all our users. For example, lots of our users asked for more automation which has led us to develop the automation wizard for Progenesis QI for proteomics v2.0.

As well as specifically targeted questionnaires, we’ll happily receive comments and suggestions via emails to our support team. It might be that a request comes in for something Progenesis can already do, in which case we’ll guide you through the process; it might even prompt the writing of a new FAQ or a redesign of the functionality in the software.

The Progenesis Improvement Program

When you opt-in to the Progenesis Improvement Program, the software will collect information about how it’s being used and about any problems you encounter. Back at Nonlinear HQ, we can then use this information to identify which features are used most heavily, which are rarely used, and which parts of the analysis are fast or slow. This helps us to see where we need to improve performance, where we could add a small but helpful feature, where we need better instructions, and much more. If you haven’t opted in already, give it a go – you can always opt out again at any time.

Our Scientific Advisory Board

Our long term decision making is helped by our Scientific Advisory Board.  This is a group of prominent scientists who meet once a year and discuss with us what we should be considering long term.  These are lively and interesting sessions;  the board is candid and tells us if it thinks we are going in the wrong direction or wasting time and resource.  An example of this is that we were considering developing a pathway analysis tool but the board said, “Don’t.  Better to integrate with what’s already available”. Hence Progenesis QI for proteomics v2.0 now includes direct support for export to third party pathway analysis programs such as IMPaLA and PANTHER.

So if you’ve got an idea on how to make Progenesis even better, or simply want to try the software for yourself, get in touch.

Three years and a million thanks!

andyHi, I’m Andy, a software engineering intern at Nonlinear Dynamics. I recently graduated from Newcastle University with a degree in Computing Science, having also spent the past three summers at Nonlinear as a way of gaining hands-on experience to complement my academic studies.

What I’ve worked on

In my time here, I’ve worked on tasks of increasing depth and difficulty:

  • My first internship saw me working with another student to develop an in-house tool to analyse and react to trends in the issues being reported from our software.
  • During my second internship, I was responsible for my own project, indexing all of our test datasets, allowing the development team to efficiently find the data they need to test new features. Managing the project myself meant that I was in charge of organising and prioritising tasks, and responsible for the project in each stage of the software development life cycle.
  • This year, I’ve been working on something slightly different: an SDF Viewer. The tool will be made available later this year and will make processing SDF files much easier by helping to combat the issue of badly formatted compound databases for use with Progenesis QI.

All of my internships have been great experiences, giving me first-hand insight into working as a professional software engineer in a world-leading company. This year’s project, however, has been especially rewarding; knowing that I’ve contributed to software that will make it into the hands of research scientists all over the world is a real buzz. :) I’m delighted to have been able to develop it for you and, while it’s not quite complete yet, I’m really looking forward to its launch!

The Nonlinear Dynamics Experience

One of the best things about Nonlinear Dynamics is the relaxed, flexible working environment. I can explore new ideas, develop innovative solutions to problems, and learn freely. Here, I’m able to plug in my headphones and work independently, or, when necessary, collaborate with my more experienced colleagues. They’ve always been available, and keen to discuss the progression of my project whenever I need reassurance or guidance.

Each summer working here has pushed my abilities. I’ve gained new skills and had the opportunity to improve upon things I had been introduced to at university. The enthusiasm of my colleagues has been contagious and inspiring, and I now feel confident in my ability to develop software as part of a team, to meet deadlines, and to create useful software.

I can’t thank Nonlinear Dynamics enough for this brilliant leg-up in my academic progression and future career as a software engineer. It’s an exciting, cutting-edge company and the learning and experiences I have gained here have been invaluable. If you’re a software developer looking for a new challenge, I can heartily recommend keeping an eye out for job vacancies with Nonlinear. Hint: at the time of writing, they’re currently recruiting — tell them I sent you! ;)

A busy summer for Nonlinear in Europe

The busy European summer conference season is almost upon us, so I’d just like to share with you all of the opportunities you’ll have to meet us and learn about the Progenesis range.

IMSC 2014

It all kicks off in the last week of August with the bi-annual International Mass Spectrometry Conference (IMSC), this year held in the beautiful city of Geneva at the Centre International De Conférences, Geneva (CICG). IMSC organisers are expecting around 1200 delegates to attend the week-long conference, so we’re anticipating another non-stop meeting. Once again, we’ll be busy demonstrating the new Progenesis QI range, as was the case at ASMS in Baltimore and then again at the Metabolomics 2014 conference in Japan.

Metabomeeting 2014

Shortly after IMSC, we are due to attend what’s being touted as “the metabolomics research conference of Europe”, Metabomeeting. This is being held at another prestigious venue: the Royal Institution in London. The organisers are expecting 300 attendees and are aiming to make the event as valuable as possible to both industry and academia.

Technology seminars

In September, we will be co-hosting a series of technology seminars with Waters, entitled Frontiers in metabolomics & lipidomics. These will take place at select European research hub locations, with the aim of the seminars focussed on discussing how new tools work together to provide superior workflows for lipidomics and metabolomics analyses.

Solutions will be presented from:

  • Small molecules research scientists talking about the workflows they are implementing to overcome challenges,
  • Waters separations science specialists discussing the novel analytical technologies to address these same challenges, and
  • Progenesis QI specialists showing off the objective and robust data analysis workflow designed to overcome small molecules research challenges

The first location of the workshop series has been confirmed, with details on more venues and speakers to follow:

On the 15th September, the technology roadshow will kick off at VIB department of Plant Systems in Ghent, Belgium – Geert Goeminne works in the Bioenergy group at VIB and will be delivering a talk on his work using LC-MS for metabolic profiling of plant energy systems.

We will have other meetings across Switzerland and Germany, with more information on dates and venues to come.

HUPO

The last conference of the summer/autumn for us will be the 13th Human Proteome Organisation World Congress (HUPO), in Madrid. HUPO really takes Progenesis back to its roots as this is the annual proteomics event which brings everyone in proteomics together. We attend every year as it is an ideal opportunity for us to catching up with a lot of old faces and to meet a few new ones too.

If you’re planning on attending any of these events or would like more information on how to book on to the technology seminars, please do get in touch. We hope to see you soon! :)

Progenesis QI: Big in Japan

At the end of June I headed across to Tsuruoka, Japan for the 10th Annual International Conference of the Metabolomics Society along with some of my colleagues from Nonlinear. With metabolomics still being a relatively new field for us, this was only the 2nd time we’d attended this conference so it was interesting to see how many more people are starting to associate the Progenesis name with not just proteomics, but small molecules analysis too.

IMG_20140624_103007

Interestingly for us there were a lot of talks focussed on software – something that came up was the importance of dedicated software testing, which some bespoke solutions lack; an example was made of an institute retracting 5 previously published papers due to software driven errors. One of the benefits of commercial software, such as Progenesis is the availability of a dedicated software testing team – in fact we’ve recently expanded ours.

Another recurrent theme was the challenges scientists face when identifying their metabolites, specifically sourcing suitable databases. It was therefore great to be able to talk people through the approach Progenesis QI takes, including the availability of theoretical fragmentation and the flexibility of what databases can be used.

One of the last talks of the conference was delivered by Matthew Lewis from the Phenome Centre who talked about how they’ve been putting Progenesis QI through its paces with thousands of LC-MS runs with emphasis on stabilisation for large scale experiments. One of the challenges faced is the ability for software to handle large data sets, and at Nonlinear we’ve been working hard to optimise use of the software for large scale experiments and have been pleased to work alongside the Phenome Centre to achieve this.

In the evenings we were free to explore the city of Tsuruoka and everything it had to offer, which was largely lots of games of darts(!), and as expected, some very tasty food.

A night out playing dartsEnjoying the local cuisine

In all the conference was a success, helping us to both spread the word of Progenesis QI and also a chance to listen to what the current challenges are to help guide the development of our next release. If you’d like to try Progenesis QI with your own data, get in touch and we’ll arrange a demo. Until then, sayōnara!

ASMS 2014: There and Back Again

At the end of last week, the Nonlinear team returned from ASMS 2014 in Baltimore, MD. Six of us attended: Mark and Jon from our US sales team, Ian and myself from the development team, plus Rob our Product Manager and Ronan our boss. With the exception of a spectacular thunderstorm on the evening of our arrival and a couple more on our final evening, we enjoyed fantastic weather and a great welcome in Baltimore.

It was my third ASMS after Minneapolis last year and Vancouver in 2012, but our first as part of the Waters Corporation. Being part of Waters meant a few changes: under ASMS rules, we weren’t allowed our own booth in the exhibition hall; and we attended the Waters Sales and User meetings before the conference properly started on Sunday evening. So for us, the conference started on the Saturday morning with the Users meeting which consisted of presentations by a mixture of Waters staff talking about new instruments and customers discussing their scientific endeavours. My personal highlight was a very entertaining and informative keynote talk by Jeremy Nicholson of Imperial College London on the topic of population scale phenomics and the iKnife.

On Sunday, the sales meeting gave us an opportunity to catch up with colleagues from the US and elsewhere to find out about the new developments in the wider Waters organisation whilst simultaneously informing the Waters field teams on what was going to be in the next versions of Progenesis QI and Progenesis QI for proteomics. After the sales meeting, Ian and I headed over to the hospitality suite to install our software and datasets onto the PCs, ready for Monday morning’s demos.

Our stands in the Waters hospitality suite

Monday was our first of three long days in the hospitality suite, demoing Progenesis QI and Progenesis QI for proteomics to existing and prospective customers. We had a shorter daytime session on the Monday to break early for the Waters press conference where we unveiled the exciting news of what’s coming in v2.0 of Progenesis QI for proteomics.

We also had our Scientific Advisory Board meeting during the break in the Waters hospitality meeting room in the Hilton hotel which was on the 15th floor with a fantastic view of the baseball stadium at Camden Yards:

Oriole Park at Camden Yards

The meeting was a great one, with members attending ASMS present in person and a couple of advisory board members dialling in from around the world. We had a very interesting discussion on present and future scientific advancements in the fields of proteomics and metabolomics and how Nonlinear products could help scientists with forthcoming challenges. After the Scientific Advisory Board meeting, half of us went to our customer dinner with some of the SAB members and a few other customers. Meanwhile, Jon and I went back to the hospitality suite for the evening shift, which involved a surprising number of customer demos, despite the lure of food and drink available at each of the suites!

Our three days in the hospitality suite were very busy, with a constant stream of interested scientists and existing customers wanting demonstrations of both the existing software and the new features that were coming in the next versions. On Wednesday morning, we also had a well-attended breakfast seminar on ‘Mass Spectrometry & Informatics for Molecular Phenotyping’ during which Rob and Mark talked about both Progenesis applications and how their results could be combined to give multi-omics pathways information in the next versions of the two products.

Now that we’re back in the office and over the worst of the jet lag, we need to crack on – working towards the Progenesis QI for proteomics v2 release and getting the new features we demonstrated into the hands of our customers. If you’d like to try Progenesis out for yourself, please get in touch.

Revealed: new features coming for Progenesis QI for proteomics v2.0

Yesterday, in Baltimore, Maryland, at the 62nd American Society of Mass Spectrometry (ASMS) conference we unveiled the exciting new features that we’ve been working on for the next release of Progenesis QI for proteomics. For those of you who missed the press release, here’s another chance to see what’s coming:

Pathway Analysis

Progenesis QI for proteomics v2.0 will include direct support for export to third party Pathway Analysis programs such as IMPaLA and PANTHER to allow you to put the protein differences in your data into a biological context in just a few clicks:

test

QC Metrics

No more wasting your valuable time performing data analysis on sub-optimal LC-MS data with the new QC Metrics, including chromatographic peak width, feature dynamic range, precursor mass error, missed cleavage count, peptide per fraction count, and many more:

image_5image_6

Automated Data Processing

With the new automation functionality you can set up your analysis to run right from the beginning at Import Data all the way to a completed peptide search without any user intervention – perfect for overnight processing.

“Hi-N” Quantification

The support of Hi-3 quantification for Waters MSe data has been extended to allow for absolute quantification of data from all the vendors we support as well as with the option to define the number of peptides used in quantification.

So if you’re at ASMS, come and see us in the Waters Hospitality suite, Ballroom 8 at the Hilton Baltimore where you can book a demonstration with some of the developers themselves. If you’re not at ASMS, get in touch and we can arrange a demonstration using your own data.

Talking LC/MS vs. NMR with the big cheeses of French metabolomics

standI’m pleased to report back after my first conference and first visit to France, having just attended The French Metabolomics and Fluxomics Network (RFMF) meeting. This is a key meeting for metabolomics research in France, bringing together both young and seasoned scientists to discuss methods and analysis in metabolomics research, which in France is heavily focussed on plant science.

There was a packed lecture schedule interspersed with short lunch breaks, and of course, special breaks to eat the plentiful amounts of cheese available. At the cocktail reception on the first evening my lovely colleague Agnès had great reason to laugh as she saw the look of horror on my face – the result of a combination of having just taken a bite of foreign fromage that exploded with insulting flavour and the perfect timing of Agnès commenting, “This is really special cheese, it’s 10 years old!”. There was yet more foodie fun to come…

From the research posters and chatting to delegates, it was easy to see a clear technology split between NMR and MS techniques. NMR is especially suited to analysis where sample availability is fairly generous. One of the senior scientists and organisers of the conference told me that there are around 200,000 variables (possible metabolites) in plant samples; being an NMR scientist, he mentioned that no one in his lab actually uses NMR, but rather they use LC-MS. In his opinion, other NMR metabolomics research scientists in France are going to have to look toward LC-MS if they are serious about doing research on plants, as metabolomic profiling of plants is too complex for NMR currently. Another challenge that was creeping up in my conversations with delegates was on the limited availability of suitable spectral libraries to study plants, so these researchers were pleased to hear about the capabilities of Progenesis QI being able to query existing databases, building up fragmentation databases and being able to do theoretical fragmentation analysis to gain confidence and to fill in the gaps.

On the final night, the conference committee organised a meal at Paul Bocuse’s restaurant, the prestigious L’Abbaye De Collonges. This was not a typical restaurant; in fact, it was closer to a carnival experience with musical puppets and waiters running around to a countdown… I’ll let the pictures tell the story, but it was fair to say that the food was opulent & highly calorific! We all left beyond the point of being satiated :)

Restaurant

cakeIMAG0299

The conference was a successful trip for us as we met researchers who were pleased to hear about Progenesis and how it could help them overcome some of the identification challenges they were facing in their metabolomics LC-MS analysis. If you are also finding many challenges with your LC-MS small molecule identifications, get in touch to find out whether Progenesis QI can help you too.

More new faces at Nonlinear

We’ve had a bit of a recruitment boom here at Nonlinear Dynamics over the past few months – firstly with the expansion of our software development team back in March – and now we’re pleased to announce the arrival of a new senior applications scientist and a new software tester.

DJ-2Dave Jackson has 14 years of experience in the proteomics sector, primarily in the 2D-gel field with emphases on methodological improvement, standardisation, analysis and biomarker discovery. Strictly speaking, this isn’t the first time we’ve shared an office with Dave as he used to work for Biosignatures Ltd, a spin-off company from Nonlinear who were fully separated following our acquisition by Waters, so it’s nice to have him back. :) He’ll be working in applications science with a particular focus on experimental design, analysis, and QC. When not found sampling the region’s ales, Dave likes to spend his time running, partly to work off the stress of being a Hull City supporter.

 

CM

Colin McKay has been a software tester since 1998 with experience in the electrical, financial, and health sectors. In fact, Colin is another person to fly the nest and come back, as he previously worked for us between 2002 and 2006 where he focussed on the testing of our 1D and 2D gel software applications, which are now developed and distributed by TotalLab. Nonlinear has a habit of luring people back! In his spare time Colin enjoys American football, watching films (particularly world cinema and horror) and amateur photography.

 

 

Interested in a career at Nonlinear Dynamics?

If Nonlinear Dynamics sounds like somewhere you’d like to work and you think you can strengthen our team, you might want to check out our recruitment page.