As of May 4, 2010, all new blog postings can be found on:

www.ludesi.com/blog

Please make sure to update your bookmarks, RSS feeds and Email subscriptions!

We look forward to welcoming you to the new site!

The Ludesi Team

Ángel García Alonso (Principal Investigator, Ramón y Cajal Research Fellow), in the center; Andrés Fernández Parguiña (PhD student), to the left; and Isaac Rosa Benito (Lab Technician), to the right.

This weeks customer profile introduces you to Dr. Ángel García’s, who is using 2D gel based proteomics to research new antithrombotic drugs and novel platelet-related biomarkers in acute coronary syndromes.

Dr. García is a Ramón y Cajal Research Fellow at the Department of Pharmacology at the University of Santiago de Compostela in Spain. He has many years experience in platelet proteomics, most recently also at the Glycobiology Institute at the University of Oxford, UK. Dr. García is successfully performing large-scale 2D-gel based proteomic analysis on clinical samples.

Read below for Dr. García’s answers to our seven customer profile questions:

How would you describe the overall goal of your research?

(i) Identify novel signalling proteins that could play a key role in platelet activation and serve as potential drug targets as antithrombotics; (ii) identify novel platelet-related biomarkers in acute coronary syndromes.

What is your goal specifically for your 2D gel based research?

Efficient and sensitive differential analyses to achieve the above goals.

What type of samples are you primarily working with?

Primarily, platelets and plasma.

What have you achieved by using 2D gels?

We have dissected platelet signalling cascades and identified novel platelet proteins. Furthermore, we have demonstrated that platelets from patients with acute coronary syndrome express a characteristic proteomic profile.

What type of Ludesi analysis do you primarily use (REDFIN Solo, Basic, or Pro)?

Primarily, Pro and Solo.

What made you choose Ludesi’s analysis service?

The versatility, time-savings and good service (with quick response to feedback and implementation of new tools)

How does using the analysis service help you in your work?

The analysis helps me to achieve my goals quickly and with reliable results. I am very pleased with Ludesi and the way they get in touch whenever I have doubts. I hope they will carry on implementing new tools (e.g. nonparametric statistics, landmarking) that will further help my research and the reliability of the results.

Read more about Dr. García’s work in some of his recent publications:

JR. High-resolution two-dimensional gel electrophoresis analysis of atrial tissue proteome reveals down-regulation of fibulin-1 in atrial fibrillation.
García A, Eiras S, Parguiña AF, Alonso J, Rosa I, Salgado-Somoza A, Rico TY, Teijeira-Fernández E, González-Juanatey JR.
Int J Cardiol, in press.

Proteomic analysis of integrin alphaIIbbeta3 outside-in signaling reveals Src-kinase-independent phosphorylation of Dok-1 and Dok-3 leading to SHIP-1 interactions.
Senis YA, Antrobus R, Severin S, Parguiña AF, Rosa I, Zitzmann N, Watson SP, García A.
J Thromb Haemost. 2009 Oct;7(10):1718-26. Epub 2009 Aug 11.
Pubmed Link

A global proteomics approach identifies novel phosphorylated signalling proteins in GPVI-activated platelets: involvement of G6f, a novel platelet Grb2-binding membrane adapter.
García A, Senis YA, Antrobus R, Hughes CE, Dwek RA, Watson SP, Zitzmann N.
Proteomics. 2006 Oct;6(19):5332-43.
Pubmed Link

Read more Ludesi customer profiles

To document some of the hard and exciting work that is going on in the field of 2D-gel based proteomics, we are going to present you with a close up of  six of our customers and introduce them to you as part of our new “Customer Profiles” series.

And first one up is Dr. Giulio Agnetti from the distinguished Van Eyk Lab:

Dr. Agnetti currently works as a post-doctoral Research Fellow at the prestigious Hopkins Bayview Proteomics Centre (Johns Hopkins University, Baltimore, MD) and is also affiliated with the Department of Biochemistry at the University of Bologna (Italy). He researches heart failure and cardiovascular proteomics together with Prof. Jennifer Van Eyk, who has a long-standing and successful track record within that area.

Read below for Dr. Agnetti’s answers to our seven customer profile questions:

How would you describe the overall goal of your research?

Understanding why the heart goes to failure. Heart failure is the clinical end-stage, common to a number of cardiac conditions (myocardial infarction, valve defect, hypertension, etc.). Being a muscle, the heart compensates for these pathological stimuli by increasing its size, a process called hypertrophy. In the short term this is beneficial but in the long run the heart tends to dilate and ends up failing to pump blood efficiently. The molecular reasons that underly this maladaptive trend are unknown. We believe that proteomics could help identifying the molecular players that drive the heart to failure.

What is your goal specifically for your 2D gel based research?

To monitor changes in post-translational modifications status of candidate cardiac proteins.

What type of samples are you primarily working with?

Cardiac tissue from both human and animal models and primary cultures of cardiac myocytes

What have you achieved by using 2D gels?

We believe that we identified a common change (candidate marker) to several types of failure. This protein may represent the key-stone of the processes that regulate the disease. We identified the first reported phosphorylations of a our candidate protein in vivo, using human and canine specimens. This may lead to the development of novel concepts and targeted pharmacological treatments for heart failure.

What type of Ludesi analysis do you primarily use (REDFIN Solo, Basic, or Pro)?

All of the above!

What made you choose Ludesi’s analysis service?

A friend and colleague compared different ways of analyzing 2D gels. Ludesi resulted in being the most reliable. Moreover, on top of saving a lot of time to researchers, the Ludesi analysis service is unbiased! This is particularly important when someone has a candidate marker.

How does using the analysis service help you in your work?

Saves me a lot of time, results are reliable, nicely organized, easily retrievable and editable. On top of resulting from an unbiased analysis!

Read more about Dr. Agnetti’s work in some of his recent publications:

Modulation of mitochondrial proteome and improved mitochondrial function by biventricular pacing of dyssynchronous failing hearts.
Agnetti G, Kaludercic N, Kane LA, Elliott ST, Guo Y, Chakir K, Samantapudi D, Paolocci N, Tomaselli GF, Kass DA, Van Eyk JE.
Circ Cardiovasc Genet. 2010 Feb 1;3(1):78-87. Epub 2009 Nov 17.
Pubmed Link

Proteomic profiling of endothelin-1-stimulated hypertrophic cardiomyocytes reveals the increase of four different desmin species and alpha-B-crystallin.
Agnetti G, Bezstarosti K, Dekkers DH, Verhoeven AJ, Giordano E, Guarnieri C, Caldarera CM, Van Eyk JE, Lamers JM.
Biochim Biophys Acta. 2008 Jul-Aug;1784(7-8):1068-76. Epub 2008 Apr 18.
Pubmed Link

Proteomic technologies in the study of kinases: novel tools for the investigation of PKC in the heart.
Agnetti G, Kane LA, Guarnieri C, Caldarera CM, Van Eyk JE.
Pharmacol Res. 2007 Jun;55(6):511-22. Epub 2007 May 3. Review.
Pubmed Link

Next week: Meet Dr. Àngel García who researches platelet proteomics at the University of Santiago de Compostela

Like for most of us, the biggest problem is always to prioritize what is most important to do right now, but nevertheless, we seem to manage a steady stream of new features and software updates coming out every few weeks. And instead of summarizing all features into yet another “new version” of the software, that we can charge you for, we instead just tell REDFIN to upgrade itself instantly at no extra cost to you.

And this time, when you start up REDFIN it will upgrade itself with the a new feature that allows you to easily customize expression profile graphs.

When we started mapping out how this feature should work, we consulted some of the leading journals in the field of proteomics to see what requirements they have for graphs and designed the tool accordingly. So now it is, for example, extremely easy for you export graphs in color or grey-scale and in exactly the right resolution needed for publication.

And this is how you do it:

First, click on the little “expand” icon above the expression profile graph in the Spot View.

The yellow circle indicates the "expand" icon.

Then, decide what style of graph you want to use.

Select your preferred style of graph

Then, click on any pre-defined label or white space within the plot to change or write your labels of choice.

Then, select if you want to display the graph in color or grey-scale.

The wobbly yellow circle indicates the display radio buttons.

Finally, click on the “Save graph” button and select what resolution you want to save the graph in.

Now all you need to do is decide under which folder you want to save the image file on your computer and you’re all done!

Do you plot your 2D gel image analysis data in other ways that we haven’t covered here? Let us know, and maybe your idea will be the next one we implement in REDFIN!

With the newest REDFIN feature of excluding individual gels at the end of an analysis, these days are over.

And, it give you even more flexibility when it comes to paying for your 2D gel image analysis.

This is why…

As you might know, you don’t have to purchase any analysis credits in REDFIN until you have reviewed and approved of the results. This already significantly trims down your costs as you only ever pay for usable data.

We have now also added the possibility for you to decide which gels you want to pay for after you have done the analysis. Nice!

For example:

You create a new analysis project in REDFIN containing 8 gels, but once reviewing the results you decide that only 6 of the gels are worth including in the overall analysis. Simply un-check the two “bad” gels, and thereby kick them out of the project before purchasing the necessary credits, without having to re-do the entire analysis!

Simply un-check the box during the review stage to exclude individual gels

Should you later decide that you would like to include those 2 gels after all, you can simply bring the project back into analysis, include the two gels you previously excluded and pay only 2 analysis credits. That’s it!

By having the option to tailor your analysis costs in this way, the average proteomics lab can keep their software costs for 2D gel image analysis below $ 3000 a year, which is only a tiny fraction of what most software package licenses cost…

Makes you think, doesn’t it?

Enjoy! (and treat the lab to something nice with the money you save… )

Here are two of our newest developments that you receive for free next time you start REDFIN:

1. Faster Spot Editing

Have you felt it takes too long time to save spot edits in REDFIN?

We for one did and have made some considerable improvements to the back-end saving of edited spot data. Basically, REDFIN is now much smarter in how it saves edited data making the whole process more efficient.

So, what does this mean for you?

Whenever you edit spot borders or matchings in REDFIN and apply the changes, after the save is done you can now immediately start editing again. No more waiting for data to be backed-up to the server.

It means that it is now twice as fast to apply edits to your REDFIN project!

2. Create Price Quotes Online

If you need a price quote for your purchasing department or grant application, you now don’t have to get in touch with a Ludesi representative but instead, you can simply create the price quote yourself online.

To create a price quote you either:

– Click on the “Buy Credits” link on your REDFIN start page and then click on the “Request price quote” link at the bottom of the window.

– Go directly to the online purchasing application by browsing to the “How to Buy” page on the Ludesi website and following the instructions on the right hand side.

Remember, that you don’t have to do anything to receive these improvements. REDFIN will automatically update itself when you next start it.

Enjoy!

And to put our money where or mouth is, we have now also created a nice step-by-step tutorial for the new Gel IQ software.

The tutorial consists of 9 short steps that guide you from software download, all the way to the results page, with appropriate links to more detailed material where needed.

And one last thing before we invite you to jump over to the tutorial itself: although not difficult to use interface-wise, the concepts behind Gel IQ can be difficult to understand at first glance, so we recommend to familiarize yourself with them before you get started. Especially the definitions of correctness are vital for a good and consistent evaluation using Gel IQ.

Click here for the Gel IQ tutorial

Hundreds of you have checked it out so far and many of you have re-tweeted about it in your personal Twitter threads – big thank you to those who did!

Just to recap – Proteomics Tweets is a free website created and maintained by us at Ludesi. It uses Twitters search widgets to pull together all proteomics-related tweets posted within the last day or so.

So in a sense it acts as a proteomics hub for Twitter, and for all of us working in the field, it is a pretty nice solution for keeping up-to-date with what is being talked about right now!

If you haven’t already checked it out, we invite you to do so by going to www.proteomics-tweets.com.

Do you feel that we are covering all relevant keywords?

A list of currently used keywords is shown on the About page and if you feel we are missing out on anything please let us know either by commenting on this blog post or by emailing support@ludesi.com.

Number 1: Quality control your 2D gel image analysis data

Gel IQ is the perfect tool to assess how correct and complete your image analysis really is.

Short of looking through and checking every single spot in the analysis, there has so far been no comprehensive way of ensuring your analysis was good.  Apart from being a clear scientific shortcoming, this inevitably results in time and money being spent on validating false hits, as well as missing out on potentially important expression changes.

By calculating a combined correctness score for your 2D gel image analysis using Gel IQ, you are able to quickly confirm that your quality is good enough or that either the spot detection or spot matching needs some tweaking.

How does it work? Simply download Gel IQ for free, upload your image analysis file, and score your analysis.

Our experience so far has shown, that anything above 70% can be deemed an acceptable analysis. If you are above 80% you should feel confident that you have a pretty complete and correct data set.

By logging the Combined Correctness scores you achieve for different sample types or stains, you will soon develop a feeling for when an analysis was good or not.

Number 2: Optimize your analysis protocol

In addition to using Gel IQ as a post-analysis quality check, you can use it to iteratively improve your analysis procedure.

Just as you would improve any other experimental protocol in your research, the image analysis process should undergo an equally systematic scrutiny. After all, a chain is only as strong as it’s weakest link!

You can start by running a more or less “automatic” analysis of your gels (for the record: we also do not believe in so-called fully automatic 2D gel image analysis…). Basically, set the parameters to what you consider to be most optimal for the gels in question, run the analysis, and score the outcome using Gel IQ.

Depending on the results, you will quickly see if either the spot detection or spot matching needs improvement. Go back and change the parameters, if possible, and re-run. If you are using a software package that doesn’t allow you to adapt detection or matching parameters, you will know what areas to focus on during manual editing.

Using Gel IQ, you will be able to confidently and systematically optimize the protocols you use for 2D gel image analysis.

After all, only what you are able to measure and quantify can also be systematically optimized…

Number 3: Evaluate the performance of different software packages

Ever stood before the decision of which software package to purchase for your analysis needs?

Now you no longer have to try and guess which software package are service works best on your data, you simply use Gel IQ to score the output you are able to achieve with them.

As always, you simply export the analysis files from the different software packages, score the quality using Gel IQ, and hey presto – you have some hard numbers on how good they really are!

Finally, we’d just like to point out again that Gel IQ is a non-commercial free-ware, based on the Combined Correctness metric, which was developed together with six proteomics research labs. We don’t want to charge for Gel IQ because we feel it is invaluable and necessary in helping you get the most out of your 2D gel experiments. And good science is something we all benefit from.

Jump over to the download page for Gel IQ

To make it easier for you to access them, we are now collecting these free tools in a new section of the Ludesi website, which we have aptly called “Free Tools“.

Depending on the nature of the tool, you will find links and instructions on how to either download the application for free, or simply access it directly through a website.

Here is a quick introduction to the tools we are currently listing:

1. Gel IQ – score the quality of your 2D gel image analysis data

Finally, an easy-to-use tool to create a quality score of how good your 2D gel image analysis really is. And most importantly – it is independent of what software package was used to perform the analysis.

Just download the free tool, upload your image analysis file (there are instructions on how to do this – don’t worry), and start scoring your quality.

Gel IQ is based on the Combined Correctness metric that was developed by Ludesi in collaboration with leading proteomic research labs.

Ludesi’s CEO Ola Forsstrom-Olsson has been presenting and discussing the issue of 2D gel image analysis data quality on numerous occasions, as part of the HUPO Industrial Advisory Board and most recently at the Quality Control in Proteomics Workshop in Hinxton. The extremely good reception that the Gel IQ tool has had so far underlines the importance of such a free tool to the community.

Try out Gel IQ on your own data by downloading it completely for free here.

2. Protein Relation Miner

Our MS data-mining tool is set out to answer one question:
How can my proteins be described and how are they related?

You simply upload a list of SwissProt accession numbers, and the application digs out information for each protein and finds similarities.

The information used today is GeneOntology terms, SwissProt keywords and SwissProt function description.

The results are displayed as tables showing which information matches among the proteins and what proteins are most similar.

Try the free Protein Relation Miner tool by clicking here

3. Image capture QC tool

Appropriate scanning of your 2D gels can have a greater impact on the outcome of your image analysis as you might think.

To help you check the quality of your image capture step, prior to committing time and money on the analysis, we have developed a small QC tool that scans your images for the following:

• image size
• image format
• greyscale
• bit depth
• dynamic range
• level of saturation
• intensity level
• grey-level span

If your images don’t pass any of these checks, you will be alerted to it, together with a message on possible consequences and a suggestion for how to improve it.

The image capture QC tool has been integrated into the REDFIN 2D gel image analysis software, but can be accessed by anyone for free simply by downloading REDFIN for free here.

More detailed instructions on how to check the quality of your scanning procedure can be found on Image Capture QC page.

As usual, if you have any questions simply send them to support@ludesi.com.