Applications

Protein quality

Robust assessment of protein quality is critical in everything from routine protein analysis in basic and translational research through to the manufacture of protein based bio-pharmaceuticals.

Fluidic Analytics’ technology allows scientists to quickly check protein quality using tiny amounts of material, providing valuable insights into oligomeric state, stability, aggregation and folding. Measurements can be made without calibration, and are unaffected by the presence of detergents.

Learn how Fluidity One can help Learn why protein quality is importent

  • Publications and resources

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    • Video20 August 2019

      Application note video summaries

      On this page we collect short video summaries of our application notes

    • Blog04 July 2019

      The Protein Society Symposium 2019 roundup

      In this blog post we showcase videos of authors taking us through their poster presentations and highlight key papers from the talks from our favourite seven topics.

    • Application note04 April 2019

      Rapid quality control of commercially available antibodies by MDS for acceptance testing

      Size presents a simple metric to assess quality as an indicator of aggregation and degradation. Here MDS on the Fluidity One and SEC analysis are both used to check 6 commercially available antibodies. Both techniques sucessfully identify problem samples, but MDS does so with 80% less time and 90% less sample.

    • Infographic15 March 2019

      The cost of poor quality protein

      This infographic explores what is meant by "protein quality" - and where it can cause problems in research.

    • Blog25 February 2019

      Gatekeeping your protein analyses

      This blog explores how "gatekeeper" techniques can save time, cost and resources in your protein analyses and gives three golden rules to keep in mind when placing gatekeepers in your lab.

    • Blog18 February 2019

      Understanding the cost of poor quality protein preparations

      Poor quality protein preparations can dramatically impact downstream analyses, confounding results and wasting significant time and money. This blog explores just what protein quality means, and where poor quality can impact your research.

    • Documentation08 February 2019

      Fluidity One and One-W comparator

      This overview document quickly outlines the differences between our Fluidity One and Fluidity One-W instruments.

    • Blog20 December 2018

      The role of analytics in the discovery and development of biopharmaceuticals

      As the biopharmaceuticals market continues to grow, how do laboratory analysis methods support discovery and development? And what should be considered when selecting an analysis method?

    • Application note11 December 2018

      Quantitation of low Tryptophan and Tyrosine peptides using the Fluidity One

      UV absorbance is commonly used as a fast method for protein and peptide quantitation, however samples must contain tryptophan (Trp) or tyrosine (Tyr) residues to be detected. Here we demonstrate the ability of the Fluidity One to assess the concentration of a range of samples which have low or no Trp and Tyr content.

    • Application note31 October 2018

      Protein Size as an Indication of Structure

      Molecular weight (Mw) is a commonly used, and for many scientists a readily understood, parameter to describe the size of a protein or complex. Here we show how hydrodynamic radius (Rh) can be used in combination with Mw to provide insights into the shape and structure of proteins and illustrate how Mw alone may not always provide a complete picture.

    • Brochure24 August 2018

      Fluidity One brochure

      The Fluidity One brochure gives an overview of the capabilities, applications and full technical specifications.

    • Application note22 August 2018

      Interleukin-2 stability in changing buffer and temperature conditions

      The stability of interleukin-2 in different buffers and storage temperatures is evaluated using the Fluidity One. We find that IL-2 forms aggregates within 24 hours in some buffers, and that the Fluidity One provides a simple means to evaluate the stability of proteins across different conditions.

    • Blog23 April 2018

      What analytical laboratory methods are available for protein quantification? How do they compare?

      Quantification is an essential part of protein analysis and quality control. Here we look at the various approaches for protein quantification, their advantages, and their limitations.

    • Publication30 November 2017

      Microfluidic diffusion analysis of the sizes and interactions of proteins under native solution conditions.

      In this paper from Arosio et al microfluidic diffusional sizing (MDS) is used in combination with pre-labelled fluorescent biomolecules to size and quantify proteins in complex mixtures. Initially, MDS is compared to Dynamic Light Scatter (DLS) - and a good correlation is observed for monodispersed solutions.

  • FAQs

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Is your protein high quality?

Click here to learn how Fluidity One can help you determine whether your protein is high quality.

Why is protein quality important?

Poor quality protein samples can result in low quality data and wasted time in downstream applications. Protein quality is influenced by a number of factors, including age, storage conditions (including temperature, buffer formulation, number of freeze thaw cycles etc.), and the method of purification. Further to this, it is rarely possible to fully ascertain the nature of a protein using just a single technique - and researchers can end up working with incorrect assumptions about their protein.

For many biophysical/structural characterization experiments, correct interpretation relies on the assumption that:

  • The measured concentration of the protein is accurate
  • Protein samples are pure and homogeneous
  • All of the protein is solubilized and in a natively active state

In the 2014 paper “Quality assessment and optimization of purified protein samples: why and how?”, Raynal et al propose a series of steps to determine the quality of a purified protein sample, through which researchers can assess the purity, integrity, homogeneity, and activity of their protein.

The Fluidity One offers an additional approach to the characterisation techniques recommended in the paper and, due to it’s speed, minimal sample requirements, and ability to quantify proteins without aromatic residues it may in some cases offer a more appropriate alternative, particularly for time sensitive applications.

Protein quantification

Understanding of total protein concentration in a sample can be used to deduce the percentage of active protein (if active protein concentration has been determined), which is important for the interpretation of experiments. Commonly used protein quantification methods include UV-Vis spectroscopy or comparison to calibration standards in Bradford or Lowry assays.

In certain situations, the Fluidity One offers a number of advantages over UV-Vis spectroscopy and colorimetric assays for protein quantification:

  • Proteins or peptides with no aromatic residues can be detected - Fluidity One uses an amine reactive, fluorogenic dye that will react with the N-terminal amine and the lysine residues of a protein.
  • Low concentration requirements - the Fluidity One can quantify proteins down to 10 ng/µL.
  • Low sample volume requirements - samples can be quantified with a high level of accuracy using just 5 µL, preserving precious samples.

Homogeneity assessment

Provided the anticipated hydrodynamic radius of a full length, correctly folded protein is known, measurement of hydrodynamic radius on the Fluidity One can provide an indication of sample monodispersity. Critically this assessment can be made in 6 minutes with just 5 µL of sample - in contrast to SEC-MALS which can take ~30 minutes and >5 µL sample, often at high concentrations.

A lower than expected hydrodynamic radius can reflect degradation or truncation of the purified protein whereas a higher than expected hydrodynamic radius may be a consequence of oligomerisation or aggregation.

Aggregation analysis

As with Dynamic Light Scattering (DLS), the Fluidity One does not resolve multiple species in a single protein preparation. However, unlike DLS, the Fluidity One gives an average hydrodynamic radius of all proteins in solution.

DLS is an excellent tool for detecting even trace amounts of aggregates in a sample as a 60 nm radius particle scatters 1 million times more light than a 3 nm one. However, as even a few percent of larger aggregates in a sample can swamp the signal coming from small particles, this technique is ill-suited to characterising samples where particulate matter is common. In these instances, the Fluidity One allows researchers to understand the true heterogeneity of their protein prep.

Lot-to-lot consistency

To ensure experimental reproducibility, lot-to-lot consistency of protein preps should be routinely assessed. Raynal et al point out, however, that once the quality of a purified protein sample has been fully assessed and optimized, repetition of the whole quality control workflow is not necessarily required for each subsequent use.

In this context, the Fluidity One measurement of hydrodynamic radius and concentration can provide a rapid assessment of the consistency of a batch prior to commencing more time consuming or costly analysis.

Fluidity One applications

The speed, convenience and low sample consumption of the Fluidity One lends it to a number of routine lab applications

Learn more