Home » Archive

Protein Aggregation

Characterization of sub-visible protein aggregates in biotherapeutic formulations.

Differential Scanning Calorimetry, Dynamic Light Scattering, MicroCal, Molecular size, Molecular structure, Particle size, Protein Aggregation, Tech Talk, Zetasizer »

[16 Nov 2017 | ]

Measuring aggregation temperature using the Zetasizer

In dynamic light scattering (DLS), the hydrodynamic size and size distribution of particles in solution can be obtained.  It may be of interest to examine this measurement as a function of time and temperature, or as a ‘trend’, as it is called in the Malvern Zetasizer software.  For biological protein samples, this temperature trend is particularly interesting: although at low temperatures a protein may be stable and show repeatable size (and scattering intensity) measurements, typically at some elevated temperature (Tagg), protein molecules will show a tendency …

Read the full story »

Archimedes, Binding affinity and Kd, Chemical identification, Differential Scanning Calorimetry, Dynamic Light Scattering, Gel permeation chromatography, Image Analysis, Label-free analysis, MicroCal, Microcalorimetry, Morphologi G3-ID, Nanoparticle Tracking Analysis, Nanosight, Particle concentration, Particle shape, Particle size, Protein Aggregation, Raman spectroscopy, Resonant mass measurement, Rheology, Size Exclusion Chromatography, Taylor Dispersion Analysis, Tech Talk, Viscosizer TD, Viscotek, Zetasizer »

[21 Sep 2017 | ]

I know this sounds like the beginning of a bad joke, but in this case, it was actually the beginning of the 2017 Colorado Protein Stability Conference, held in July at Beaver Run Resort in Breckenridge, Colorado. In attendance was a range of scientists and researchers from all over the world, with one common subject in mind: Protein Stability.

One of the highlights of this conference was the pre-conference workshop. This was a free workshop for all attendees of the conference, which was sponsored by Malvern Panalytical. The workshop included a range …

Read the full story »

Differential Scanning Calorimetry, Dynamic Light Scattering, Meet the Experts, MicroCal, Molecular size, Molecular weight, OMNISEC, Particle concentration, Protein Aggregation, Size Exclusion Chromatography, Taylor Dispersion Analysis, Zetasizer »

[18 Jul 2017 | ]

Monoclonal Antibodies – or mAbs for short

Since many of these mAbs require refrigeration we had used dynamic light scattering (with a Zetasizer) to show the formation of aggregates after freeze-thaw cycles (for details see application note “Effect of storage conditions on Immunoglobulin“). These could be seen both in the increased polydispersity index (PDI) and the appearance of a large-size aggregate peak in the intensity size distribution as shown to the right. The area under the peak gives an indication of the presence of the large aggregates but is not suited for …

Read the full story »

Archimedes, Customer experience, Differential Scanning Calorimetry, Gel permeation chromatography, Image Analysis, MicroCal, Morphologi G3-ID, Nanoparticle Tracking Analysis, Nanosight, Particle size, Protein Aggregation, Raman spectroscopy, Resonant mass measurement, Viscotek »

[1 Jun 2017 | ]

Our key driver at Malvern Instruments is to create technologies that make a difference or solve a problem, and working hand-in-hand with our customers enables us to make sure that’s exactly what’s happening, out in the field.  We place a high value on our collaborations and partnerships with customers from a wide range of industries and organizations across the globe and love to see how our systems are being used in real-world scenarios.  This gives us vital feedback on what we’re doing well, and also any tweaks we need to make in order to optimize …

Read the full story »

Archimedes, Chemical identification, Label-free analysis, Malvern Panalytical Events, MicroCal, Microcalorimetry, Molecular size, Molecular structure, Molecular weight, Morphologi G3-ID, Nanosight, Particle concentration, Particle shape, Particle size, Protein Aggregation, Protein Mobility, Viscosizer TD, Zeta potential, Zetasizer »

[16 May 2017 | ]

With Pre-Conference Workshop on Biophysical Characterization and Particle Analysis
 

Date: July 17th – 20th.
We at Malvern know that protein stability is important in numerous disciplines, ranging from basic and medical biochemistry to pharmaceutical sciences. However, it is rare that researchers from all the relevant areas can join to discuss the critical issues in the field. The 2017 Colorado Protein Stability Conference provides a unique forum for this exchange of information. The topics to be covered include protein folding, dynamics and thermodynamic stability; amyloidosis and protein aggregation; and stability, formulation, analysis and processing …

Read the full story »

Dynamic Light Scattering, Molecular size, Particle concentration, Particle size, Protein Aggregation, Tech Talk, Zetasizer »

[7 Mar 2017 | ]

How can I use DLS for sample xyz?
What sample concentration / refractive index is needed for reliable DLS size measurements?

Has it been done before?
In Dynamic Light Scattering (DLS) intensity fluctuations in the sample are analyzed to obtain size information on the sample. The technique is applicable to a wide size range from sub-nanometer up to a few microns. In order to obtain a meaningful result, a minimum scattering signal strength must be present. Malvern’s Zetasizer Nano software contains a calculator to help estimate the required minimum concentration.
If anyone else has …

Read the full story »

Dynamic Light Scattering, Molecular size, Protein Aggregation, Tech Talk, Zetasizer »

[1 Nov 2016 | ]

α-synuclein promotes vesicle assembly – when size can support theory
 

What is a vesicle?
In cell biology, a vesicle is an (often spherical) assembly of amphiphilic molecules (usually proteins and phospholipids), and is typically sized between 30 nm – 100 nm in diameter. Vesicles are often classified by the type of interaction they are involved with. For extracellular vesicles (or exosomes), this is usually intercellular signal transmission. A particular example of exosomes is synaptic vesicles, which transmit signals between neurons (average size: 40 nm).
In a paper entitled “Structural basis of synaptic vesicle assembly promoted …

Read the full story »

Dynamic Light Scattering, Electrophoretic light scattering, Meet the Experts, Particle size, Protein Aggregation, Zeta potential, Zetasizer »

[11 Oct 2016 | ]

Adjuvants for vaccination – How size and zeta may be relevant
 
What is an adjuvant?
Adjuvants are substances which are added to medicines to trigger and enhance an immune response to an antigen. In the USA, the most commonly-used adjuvant is alum, and many vaccines in use today contain some aluminum components to help the vaccine work better. While the exact mechanisms of how adjuvants work is unclear to some extent, a typical adjuvant component is much larger than the antigen or antibody itself, often around 100 nm in radius.
In a paper entitled …

Read the full story »

Dynamic Light Scattering, Nanoparticle Tracking Analysis, Nanosight, Particle concentration, Particle size, Protein Aggregation, Tech Talk, Zetasizer »

[15 Sep 2016 | ]

Nanoparticle Tracking Analysis or Dynamic Light Scattering ?
After a recent joint NanoSight and Zetasizer demonstration, I received this set of questions that helps illustrate the core of the issue of these two techniques:

Since the Nanosight gives me particle concentration, it’s somewhat straightforward to calculate what % of my Nanoparticles are aggregated. Can I calculate % aggregated particles in the DLS as well?  Or does the varying amount of intensity, and the protein+nanoparticle mixture in the sample confound that calculation?

If I wanted to measure non-spherical particles (for example nanorods), which system …

Read the full story »

Nanoparticle Tracking Analysis, Nanosight, Particle concentration, Particle size, Protein Aggregation, Tech Talk »

[26 May 2016 | ]

We recently published our application note, “Adeno-Associated Virus titer and aggregation characterization“, which discusses a new NanoSight method that allows the visualization and therefore analysis of viruses smaller than what was previously possible to measure.  This represents an exciting extension of the Nanoparticle Tracking Analysis (NTA) technique in response to market demand for tools for better characterization of small virus types.
 
Adeno-Associated Virus (AAV) is commonly used in the gene therapy field as a delivery vector. Tools to characterize both the concentration and purity or aggregation of the samples are limited because the primary virus size is …

Read the full story »

Differential Scanning Calorimetry, Electrophoretic light scattering, Isothermal Titration Calorimetry, Label-free analysis, Malvern Panalytical Events, MicroCal, Microcalorimetry, Molecular structure, Protein Aggregation, Raman spectroscopy, Zetasizer »

[12 May 2016 | ]

I had the pleasure of participating in the 5th International Symposium on Higher Order Structure of Protein Therapeutics, also known as HOS2016, sponsored by CASSS. The meeting was at the Renaissance Hotel in Long Beach, California April 11-13, 2016. I was joined by my Malvern colleagues Rajib Ahmed and David Sanborn.
The HOS Symposium is a platform for open discussion of development and optimization of analytical techniques to study HOS of proteins and biopharmaceutical products (including monoclonal antibodies and antibody-drug conjugates), biocomparability, and biosimilars.
Protein higher order structure (HOS) includes the secondary, …

Read the full story »

Dynamic Light Scattering, Molecular size, Molecular weight, Protein Aggregation, Tech Talk, Zetasizer »

[21 Jan 2016 | ]

How do I know the minimum concentration needed for measuring the size of my protein?

We are asked this question all the time when we are demonstrating the Zetasizer.  Happily, if the Zetasizer software is installed (and it can be downloaded for free, by the way) there is a minimum concentration calculator integrated – so it’s simple! The calculator can be accessed from Tools – Calculators – Concentration Utilities – Minimum Concentration Calculator
Let’s look at a few examples to get an idea of some typical values for your samples. The left …

Read the full story »

Gel permeation chromatography, Protein Aggregation, Tech Talk, Viscotek, Zetasizer »

[15 Sep 2015 | ]

How can a quartz cuvette be properly cleaned?
The Zetasizer has a flow cell [ZEN0023] used in conjunction with either the titrator, the NanoSampler, or as a chromatography or field flow fractionation detector. A similar flow cell [ZMV1008] is also in use with the Zetasizer microV when used as a molecular weight detector in conjunction with the OmniSEC software. Advanced cuvettes may also be used in UV-Vis, fluorescence & circular dichroism spectrophotometers, MALLS as well as goniometer systems and other specialized optical detection. If the signal from the quartz cell is …

Read the full story »

Dynamic Light Scattering, Molecular size, Protein Aggregation, Tech Talk, Zetasizer »

[30 Jul 2015 | ]

Often, pharmaceutical formulations contain therapeutic proteins at relatively high concentration. Under such conditions, the stability of the protein as well as the formulation overall is of significant concern. Great effort is invested in optimizing stability, and particular attention goes to the choice of buffer components. Various components might form part of the ‘final’ recipe including cosolvents (such as polyethylene glycol (PEG), glycerol, ethanol) to reduce the dielectric constant of the buffer and influence electrostatic interactions, chelating agents (citric acid or citrate salt buffers, ethylene-diamine-tetra-acetic acid (EDTA), tris(hydroxymethyl)aminomethane (TRIS), tartaric acid) and …

Read the full story »

Molecular structure, Molecular weight, Protein Aggregation »

[21 May 2015 | ]

We recently published a new application note on protein characterization using our latest SEC system, OMNISEC, and I wanted to bring your attention to this elegant piece of work that beautifully shows the kind of information you can get from a multi-detector system and, perhaps more importantly, the kind of information you might be missing from a single-detector, UV-based system.The chromatogram below shows a typical chromatogram of a standard protein,    β-amylase:

It looks pretty typical with a monomer and a small amount of aggregates, right? Wrong!
Let’s looks again with a multi-detector …

Read the full story »

Dynamic Light Scattering, Particle size, Protein Aggregation, Tech Talk, Zetasizer »

[31 Mar 2015 | ]

Can one calculate a PDI value excluding a small aggregation peak? For example if the main particle species is ~100nm and there is a small contribution (2% by intensity) from a peak at 5 microns, is it possible to recalculate the z-average and the PDI ignoring the 5 micron peak. Alternatively, is it possible to determine a PDI for the smaller species only?
The z-average will be weighted more towards smaller components, because only the initial part of the correlation function is fit. Following the ISO method to determine the z-average …

Read the full story »

Dynamic Light Scattering, Meet the Experts, Nanoparticle Tracking Analysis, Protein Aggregation, Raman spectroscopy, Resonant mass measurement, Size Exclusion Chromatography, Tech Talk »

[24 Feb 2015 | ]

I recently took part in a question and answer session with BioPharm International about techniques for measuring and characterizing protein aggregates. The full discussion can be viewed here, however I thought I would also use this blog post to summarize the main points.
Essentially, there is a pressing need within the biopharmaceutical industry to understand aggregation pathways, and the risk factors that can induce aggregation, right from the start of the drug development process to align manufacturing processes with Quality by Design (QbD) principles. Given the link between protein aggregation and …

Read the full story »

Molecular weight, Protein Aggregation, Size Exclusion Chromatography, Tech Talk, Viscotek »

[28 Jan 2015 | ]

Light scattering in chromatography can sometimes be challenging: The scattering signal is proportional to the size to the 6th power, and that is why particles in the mobile phase can lead to noise in the light scattering detector. Now we have special light scattering columns available specifically designed for low shedding, making them particularly well suited for analytical chromatography of proteins and peptides. The Viscotek PLS-columns are silica-based columns for the separation of proteins for light scattering applications. Their excellent stability minimizes particle shedding to maintain light scattering baselines as …

Read the full story »

Dynamic Light Scattering, Molecular size, Protein Aggregation, Tech Talk, Zetasizer »

[14 Oct 2014 | ]

There is one, and it is called the NanoSampler.
The Zetasizer has been around for a few years, and the Zetasizer series now includes several models (90 degrees, backscattering, size and zeta, flowmode for chromatography detection, automated plate sampler). A very recent accessory which is compatible with any Zetasizer Nano in the field is the autosampler or with its full name the “Zetasizer NanoSampler“.
In dynamic light scattering (DLS) the size distribution of molecules or particles can be obtained in a relatively short time.  The technique is therefore ideal for automation, because preparing …

Read the full story »

Molecular weight, Protein Aggregation, Static Light Scattering, Tech Talk, Zetasizer »

[4 Sep 2014 | ]

Last week we offered a webex presentation titled “Speed your way through protein formulation screening by automating your measurements: A2/B22 with the Zetasizer APS”
The presentation had a focus on protein formulation, where often the objective is to assess protein stability as a function of various parameters. Here, light scattering can be used as a tool to assist in screening conditions:

Monitor: Aggregation temperature, Dynamic Light Scattering (DLS) vs temperature
Predict: Interaction parameter, DLS vs concentration
Predict: Second Virial Coefficient, Static Light Scattering (SLS) vs concentration

While the Zetasizer APS has previously been used for dynamic …

Read the full story »