Dynamic imaging particle analysis (DIPA) systems provide you with the in-depth characterization of particles in protein therapeutics as recommended by the FDA. Here are five important features that should be considered when exploring instrumentation that use this principle for protein aggregate characteriziation in your formulations.
#1. Image Quality
Image quality is extremely important when characterizing protein aggregates, and differentiating them from silicone oil and other contaminants in a drug formulation. A high quality image is essential for accurate sizing measurements. Because DIPA instruments take measurements directly from the images they collect, the precision and accuracy of the data is directly related to the quality, or sharpness, of the images.
Blurry images make it difficult to distinguish protein aggregates from other contaminants and to appreciate the morphological differences in aggregated proteins themselves. These differences may be an important indication of bacterial or viral contamination (e.g. aggregates containing round globules versus rods).
Poor image quality can also result in low concentrations or inaccurate characterization because smaller or faint particles (such as those with a low refractive index) are mischaracterized, or missed entirely. This results in increased measurement variation, and a lower statistical confidence.
The ability of a DIPA instrument to match patterns and classify particles is also affected by image quality. The true strength of the DIPA technique over other particle analysis techniques is the ability to differentiate particles based upon appearance. The better the overall image quality, the easier it is for the instrument’s software to recognize patterns, and in turn classify particles. Therefore, image quality is paramount to the value of a dynamic imaging particle analysis system. In short, fuzzy images lead to fuzzy measurements, which lead to fuzzy classifications.
Not all formulations are alike – particles vary significantly from one formulation to another. Depending on the formulation, these differences may necessitate a modification to capture settings to ensure proper measurement. It is important to choose a system that can be configured to suit the specific formulation you are testing and achieve optimum results when detecting protein aggregates. This is especially true if your formulations contain stabilizers, which can affect the refractive index of the matrix.
Also, if a wide range of particles sizes is expected, an instrument may need to support multiple magnifications. For example, the FlowCAM® VS provides interchangeable 2X, 4X, 10X, and 20X objectives, and can utilize both disposable flow cells and fixed field of view flow cells in a variety of sizes.
#3. Instrument Sensitivity
Many protein aggregates are translucent. This can make them difficult to detect with any particle analyzer because their refractive index is close to the matrix in which they are suspended.
In some systems, simple darker-only thresholding is used, which may cause transparent aggregates to be cut into smaller particle pieces or go undetected. This results in incorrect measurements and incorrect count or concentration calculations.
Having the ability to threshold on either darker pixels (relative to the background), lighter pixels or both simultaneously can ensure the proper protein aggregate characterization, especially when working with high concentration formulations or those with a large amount of stabilizer.
#4. Sample Volume Required
Since drug formulations can be expensive or in short supply, it is important to clearly understand the sample volumes a particle analyzer requires to provide accurate results. It is also important to understand how much sample will be needed (if any) to prime the system prior to measurement. Minimum sample volumes differ between types of imaging particle analyzers from 50 µL to 1 mL.
#5. Ease of Analysis and Data Processing
Data is one thing – actionable information is another. Once your data is captured using a dynamic imaging particle analysis system, it is important to have an easy way to analyze it. The quality of the software varies significantly from one type of DIPA system to another. Choose one that allows you to efficiently interact with your data and extract information quickly and easily.
Some key software features include:
Sorting and filtering particle images based upon criteria you supply, with immediate visual feedback.
Sophisticated pattern recognition capabilities that immediately find and display all similar-type particles in a heterogeneous sample.
Creation of user-defined particle-type libraries to instantly enumerate concentrations of specific particle types.
Satellite software for post-processing data at a remote location or sharing data with others.
The benefits of DIPA are examined in Protein Aggregation and Emerging Tools to Support Development and Characterization, a recent presentation by Gilead’s Danny K. Chou, PharmD, PhD.
Discover how you can improve protein aggregate characterization in parental drug formulations with DIPA in our informative eBook, The Ultimate Guide to Dynamic Imaging Particle Analysis.