New guide to data interpretation

June 5, 2014
Effects of normalisation on false positives and false negatives when applying t-test for equality of the mean. (A) We consider responses to eight conditions with log-normal distributions with CV of 0.2 and means of the conditions from 1 to 8 equal to: 1, 2, 2, 4, 7, 7, 18, 18. A number n = 5 of sampled replicates are obtained from these distributions and normalised using the normalisations above. Using these replicates before and after normalisation, conditions are tested using a two-tailed t-test with threshold p-value of 0.05. We repeat this procedure a large number of times and estimate the percentage of false positives. (B) In analogy with (A), we estimate the number of false negatives considering means of the conditions from 1 to 8 equal to: 1, 2, 3, 4, 7, 10.5, 18, 27. Notice that for a fair comparison, when testing two conditions, one has a mean that is always 2/3 the mean of the other, e.g. Condition 5 has mean 7 and Condition 6 has mean 10.5, with 7/10.5 = 2/3. 

Western blotting is a widely used technique to detect specific proteins. Although considered a semi-quantitative method, the results are often interpreted quantitatively. Scientific articles often do not specify how researchers quantified these results and how they compared biological replicates for statistical testing.

Conway Fellow, Prof Boris Kholodenko and his team in Systems Biology Ireland have described how results of statistical testing applied to western blot data are affected by the choice of normalisation strategy applied to the data. They have created a step-by-step guide to help scientists choose the most appropriate normalisation strategy for their particular study.

Normalisation strategies transform or manipulate data to allow quantitative comparison of biological replicates. The Kholodenko group used mathematical models to simulate the effects of normalisation and actual experimental data to corroborate the results.

"We clarified the quantitative use of western blot data and how the normalisation strategy applied to the data affects the statistical testing, possibly increasing false positives or false negatives", explained Dr Andrea Degasperi from the Kholodenko group.

Western blotting provides experimental evidence that supports a given hypothesis and is also used to validate data obtained from other techniques. Clarifying its quantitative use for decision making or statistical testing is necessary.

The Kholodenko group hope their study findings will not only act as a reference for scientists but also encourage them to include this critical information about data interpretations in published articles.

Explore further: A new way of ranking universities

More information: A. Degasperi, M. R. Birtwistle, N. Volinsky, J. Rauch, W. Kolch, B. N. Kholodenko. "Evaluating Strategies to Normalise Biological Replicates of Western Blot Data." PLoS ONE, 9(1): e87293. DOI: 10.1371/journal.pone.0087293, 2014.

Related Stories

A new way of ranking universities

January 17, 2012

(PhysOrg.com) -- An academic at the University of Hertfordshire has challenged the way university league tables are calculated and presents a radically different way of formulating university rankings in the UK, in a paper ...

Researchers propose a better way to make sense of 'Big Data'

February 18, 2014

Big Data is everywhere, and we are constantly told that it holds the answers to almost any problem we want to solve. Companies collect information on how we shop, doctors and insurance companies gather our medical test results, ...

Drug sensitivity predicted computationally

June 3, 2014

With modern high-throughput technologies researchers can measure a multitude of molecular properties from cancer cells. A big question of precision medicine is how computational modelling can be used to predict sensitivity ...

Recommended for you

Just how good (or bad) is the fossil record of dinosaurs?

August 28, 2015

Everyone is excited by discoveries of new dinosaurs – or indeed any new fossil species. But a key question for palaeontologists is 'just how good is the fossil record?' Do we know fifty per cent of the species of dinosaurs ...

Fractals patterns in a drummer's music

August 28, 2015

Fractal patterns are profoundly human – at least in music. This is one of the findings of a team headed by researchers from the Max Planck Institute for Dynamics and Self-Organization in Göttingen and Harvard University ...

0 comments

Please sign in to add a comment. Registration is free, and takes less than a minute. Read more

Click here to reset your password.
Sign in to get notified via email when new comments are made.