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Framework to Biologics – Part Three

Part One: Biologics and the Problem with Residual DNA 

Part Two: Cell-Based Bioassays 

Part Three: Micro-Flow Imaging

In this final post of our biologics blog series, bioassay scientists will construct a guide to the components of micro-flow imaging: imaging, counting and sorting, plus sample requirements.

Biologics_Blog3.jpgImaging
Micro-Flow Imaging™, or MFI, couples microfluidics with digital microscopic imaging to determine particle size and concentration in biologics and injectables. The concept of Micro-Flow Imaging is very simple. If a particle moves through the flow cell, the particle is imaged and counted. The principles of flow cytometry, microfluidics, microscopy and digital imaging are combined into one instrument.

Sub-visible particles can be unintentionally present in parenterals as a byproduct of drug formulation and manufacturing. This type of analysis allows for the visual assessment of the types of particles present in a solution. Particles that can be present in biopharmaceutical preparations are silicone oil, air bubbles, protein aggregates, rubber pieces from closures and glass shards.

MFI can be used to evaluate stress, solubility, stability and mixing studies. For example, freeze-thaw studies can be performed to see if a drug product is stable over time. Additionally, shaking and mixing studies can be evaluated to determine if protein aggregation or particle formation occurs as result of product handling.

Counting
Alcami has a Protein Simple MFI 5200 that is capable of sizing particles from 1-70µm in sterile preparations with an analysis flow rate of 150 uL/minute. This instrument can count up to 900,000 particles/mL with low sample volume (as little as 500µL), which is beneficial in cases where sample is limited. For high throughput projects, automation is available. MFI can perform particle counts per USP chapter <788>, Particulate Matter for Injections.

Sorting
MFI comes equipped with analytical software that can sort and count particles based on a specific particle type. In addition to concentration, particles can be further characterized by their intensity and roundness for the sorting of particles based on size, shape and intensity. Protein aggregates can be sorted from non-proteinaceous particles as well as silicone oil from air bubbles. Translucent protein particles can be distinguished from darker particles, a feature that traditional light obscuration and microscopic methods may not be able to detect.

Sample Requirements
MFI relies on light obscuration so the solution tested must be optically clear. The instrument can analyze samples with some degree of viscosity. Additionally, a sealed, sterile product is required to ensure that extraneous particles are not introduced into the sample from the environment or container.

Thank you for your interest in this rapidly expanding market. This concludes our three part series delving into biologics. So what's next? If you think MFI analysis can benefit your drug development program, Alcami can perform a feasibility study to see if this method works with your product. After feasibility is established, the method can be validated for routine and stability studies.

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About the Authors

 

BryanTroxell.jpegBryan Troxell, Ph.D., is a Scientist within the Bioassay group of Alcami. He has 14 years of experience in the design and optimization of fluorescent-based PCR methods. Bryan's previous work focused on gene expression technologies and has published 18 scientific publications. He currently performs routine and non-routine testing in support of pharmaceutical product development and manufacturing.

 

 

AudreyONan-Photo.pngAudrey O'Nan, is a Scientist within the Bioassay group of Alcami.  She has over 15 years experience working in a genomics and proteomics lab with a focus on animal genomics and pharmacogenomics with an emphasis on gene expression, post-translational modifications, DNA and RNA sequencing, and real-time PCR. She has been with Alcami for over two years and performs residual DNA testing, Microflow Imaging, SDS-PAGE, routine enzymatic assays and mitogenic cell assays.  Audrey graduated from University of North Carolina at Wilmington with a B.S. in Biology and minor in Chemistry, and has a M.S. degree from the University of Tennessee.

 

Topics: Thought Leadership Biologics