Fluorescence

A-TEEM for Biotech and Biopharma

<< View all A-TEEM Spectroscopy Applications

A-TEEM is HORIBA’S proprietary fluorescence spectroscopy technology that simultaneously acquires, Absorbance, Transmission, and fluorescence Excitation and Emission Matrix measurements, correcting for the inner filter effect on the fly.

Fluorescence is insensitive to many common excipients, while being extremely sensitive to components containing conjugated rings. Proteins, small molecule APIs, aromatic amino acids, co-enzymes (NADH, Flavins) are easily characterized, whereas water, sugars, and other common excipients disappear.

A-TEEM can be used to characterize relevant components in complex matrices to sub-ppb levels. This makes it a natural fit for the characterization of many relevant biopharmaceutical products such as vaccines, insulin, viral vectors, anti-body drug conjugates, cell media, exosomes, CRISPr/CAS9, and more.  

As a spectroscopic tool, A-TEEM can be deployed for process analytics, and used to monitor bioprocesses or sample purity.

Putting a sample in a cuvette is often the only preparation needed for measurement. For some quantitative measurements, sample dilution may be required. The Aqualog can be configured to work with an autosampler for unattended operation of multiple samples.

The Aqualog software is fully equipped to facilitate both calibration and validation of all instrumental absorbance and fluorescence functions following the specifications describe in the USP General Chapters for Fluorescence Spectroscopy <USP 853> and UV Spectroscopy <USP857>. IQ/OQ protocols ensure proper install and operation according to requirements.

A-TEEM for Biotech and Biopharma

Biotech and Biopharma

Learn what A-TEEM Spectroscopy can do for pharmaceutical analysis.

Application Notes

Read the application notes created by our applications scientists using A-TEEM Molecular Fingerprinting.

eBooks

Our eBooks serve as a comprehensive survey of each application for A-TEEM. For water analysis, it presents scientific papers, case studies, and videos to better inform you of various applications for this category.

Webinars and Videos

Watch our on-demand water webinars and videos to gain a better foothold on current research and emerging techniques.

Case Studies

Read our articles on wine and spirits from the Science in Action series of case studies examining a variety of applications for wine, spirits, beer and tea characterization.

Bioprocess Products

Aqualog for wine

Find out which HORIBA products incorporate A-TEEM Spectroscopy.

A-TEEM for Biotech and Biopharma

A-TEEM is extremely sensitive to components containing conjugated rings. Molecules such as proteins, small molecule APIs, aromatic amino acids, co-enzymes (NADH, Flavins), vitamins, hydrolysates, by-products of bacterial contamination, and colored compounds are easily characterized.

By contrast, common excipients without conjugation, such as water, sugars, glycerin, polyethylene glycol, and others, are invisible to A-TEEM.

The high specificity and selectivity of A-TEEM fingerprints make it a natural fit for the characterization of biopharmaceutical products, enabling quantitative analysis at ppb levels of complex mixtures. 

A-TEEM has be used to determine the quality of basal and feed media, to monitor bioprocesses, for reaction monitoring, and the characterization of vaccines, insulin, viral vectors, exosomes, anti-body drug conjugates, exosomes, CRISPr/CAS9 compounds, and more. 

Samples are most often measured in a cuvette, which can be coupled with an auto-sampler for unattended measurements. As a spectroscopic tool, A-TEEM can be deployed for process analytics, used to monitor bioprocesses, or screen for batch-to-batch variance across a variety of relevant biomolecules.

Probe-based measurements in immersion or stand-off configurations are also possible, with many options for customization available.

 

A-TEEM Meets USP Specifications

The HORIBA Aqualog UV-800 facilitates both calibration and validation of all instrumental absorbance and fluorescence measurements, according to USP chapters <857> Ultraviolet-Visible Spectroscopy, and <853> Fluorescence Spectroscopy. The Calibration/Validation Toolbox is modular and intuitive, and along with HORIBA IQ/OQ documentation, guides users through the validation process. Individual Pass/Fail tests can be selected individually to minimize test redundance and facilitate more routine testing. The Aqualog is fully supported for pharmaceutical and industrial installations with a comprehensive IQ/OQ protocol.

Download the application note:
Validation of the Performance of the HORIBA Aqualog for Pharmaceutical and Industrial Analyses

Benefits of A-TEEM for Biotech and Biopharma

Benefit of Best-in-Class Molecular Sensitivity

  • Cell Culture Media: raw material QC screening
  • Vaccines: differentiate similar multicomponent vaccines, QC of finished products
  • Viral Vectors: Serotype differentiation in AAVs
     

Benefit of Quantification comparable to HPLC & GC

  • Bio-fermentation monitoring: track low level components to ppb limits
  • Viral Vectors: fully vs empty ratio determination for AAVs
  • Binding events: use tyrosine as environmental marker
     

Benefit of high selectivity and concentration independent measurements:

  • A-TEEM molecular fingerprint: create libraries of A-TEEM contours
  • Unknown and Contaminants: Rapid detection and ID of contaminants

A-TEEM for Cell Culture Media QC

The cost of cell culture media is a fraction of the value of the final product, therefore assessing its quality prior to the start of a bio-fermentation is a cost-effective way to ensure end-product quality and quantity. Standard approaches to perform this critical QC step (chromatography, mass spectrometry, test culture) are time consuming and expensive. A-TEEM provides a robust approach to assess the quality of cell culture media prior to the addition of cells, even for complex, non-chemically defined media using hydrolysate supplements.

A-TEEM for Cell Media characterization – boost upstream product yields

A-TEEM for Rapid Vaccine Characterization

Standard separations-based approaches for vaccine characterization are too slow to keep up with the industry demand for rapid analysis. However, typical vaccine formulations (very low protein concentration, aqueous solutions, with confounding excipients) make these very challenging samples for the “standard” spectroscopic toolbox (Raman, FT-IR, NIR). The capabilities of A-TEEM make it  highly suited for the analysis of vaccines. A-TEEM has been used to differentiate vaccine formulations, detect aggregation, identify amino acid substitutions and post-translational modifications, screen for batch-to-batch variation, and perform QC measurements for batch release.

A-TEEM for Vaccine Characterization – Differentiate similar formulations

A-TEEM for Vaccine Characterization – Differentiate similar formulations

Samples provided by Dr. Laura Sagle at the NIH Vaccine Research Center

A-TEEM for Vaccine QC – Detect batch-to-batch variation

A-TEEM for Vaccine QC – Detect batch-to-batch variation

A-TEEM for Viral Vector Characterization

The commercial deployment of biotherapeutics depends on advanced analytical tools for characterization, with a growing emphasis on speed and sustainability. A-TEEM has demonstrated the capacity to resolve AAV serotypes (AAV2 and AAV9), and to accurately quantify the payload filling percentage.  A-TEEM, mostly owing to the high sensitivity of fluorescence, could be a suitable rapid alternative to other methods such as TEM for characterizing AAV materials including these certified reference materials. 

A-TEEM for AAV characterization – differentiate serotypes, rapidly determine empty/full ratio

A-TEEM for AAV characterization – differentiate serotypes, rapidly determine empty/full ratio

A-TEEM for Exosomes

A-TEEM was evaluated for its potential to differentiate exosome types. Exosome standards (PC-3 and MCF7) and their mixtures were evaluated. The different exosomes were easily differentiated by A-TEEM, and a PLS model was readily able to predict the concentrations of the two components in samples comprised of the two exosomes mixed in known proportions. 

From these initial results, it is expected that A-TEEM could be used as a rapid and cost-effective tool for manufacturing to find contaminants, detect batch-to-batch variation, and quantify payloads.

A-TEEM for Exosomes – Differentiate and quantify exosome types

A-TEEM for Exosomes – Differentiate and quantify exosome types
Differentiate and quantify exosome types

A-TEEM for Protein Structure and Stability - Insulin

Changes to the local solvent environment of the fluorescent residues monitored by A-TEEM fingerprints provides insights into aggregation and protein secondary structure. Short-acting and long-acting insulin have unique A-TEEM molecular fingerprints based on the different protein environments between formulations.

Fully corrected EEMs and Absorbance spectra of human insulin

Source: “A-TEEM™, a new molecular fingerprinting technique: simultaneous absorbance-transmission and fluorescence excitation-emission matrix method,” published in Methods and Applications in Fluorescence, Volume 6, Number 2 https://doi.org/10.1088/2050-6120/aaa818. Acquired with Aqualog.

A-TEEM for Lipid Nanoparticles

A-TEEM has also been used to study the ratio of intact vs damaged Lipid Nanoparticles (LNPs) delivered by different nebulizer devices.  To quantify the integrity of the LNPs during the nebulization process, a fluorescent protein (calcein) was encapsulated in several LNP formulations. A-TEEM was used to quantify the relative amount of calcein released during this process. This measurement characterizes devices that are most effective in delivering intact LNPs to patients.

Read the publication:
Degradation of lipid based drug delivery formulations during nebulization

Application Notes

A-TEEM can be used to characterize relevant components in complex matrices to sub-ppb levels. This makes it a natural fit for the characterization of many relevant biopharmaceutical products such as vaccines, insulin, viral vectors, anti-body drug conjugates, cell media, exosomes, CRISPr/CAS9, and more. Use the filters below to find the application most suited for your needs.

Webinars and Videos

Watch ondemand webinars and videos and learn how A-TEEM spectroscopy is used in the Pharmaceutical indusrty.

Products

Aqualog - A-TEEM Industrial QC/QA Analyzer
Aqualog - A-TEEM Industrial QC/QA Analyzer

A Simple, Fast, “Column Free” Molecular Fingerprinting Technology

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