Biophysical Society 65th Annual Meeting Program Guide

Wednesday, February 24 11:30 AM – 12:00 PM Bruker

4:00 PM – 4:30 PM Horiba Scientific Rapid, Optical Technique For Sensitive Characterization and Differentiation of OTC Canine Vaccines The pharmaceutical industry increasingly relies on spectroscopy for quality assurance and has established, and conforms to, USP (United States Pharmacopeia) regulations. While some spectroscopic approach- es (NIR, FT-IR and Raman) are common, the adoption of fluorescence spectroscopy has lagged, even though it has high specificity and sen- sitivity in many analyses in demand, and is conveniently amenable to chemometrics analysis. We present identification and validation of “unknown” samples with 100% certainty based on A-TEEM fluorescence analysis of Solo- Jec brand canine vaccines from Boehringer Ingelheim VetMedica: Spectra-5, Spectra-6, Spectra-9, and Spectra-10, containing 5, 6, 9 and 10 vaccine combinations, respectively of coronavirus, hepatitis, adenovirus, parainfluenze, leptospirae, parvovirus, etc. A key con- sideration upon an established vaccine product release is specificity. Analytical techniques need to characterize the final product, and must also differentiate between it and all others made at the facility. The 3-D fluorescence molecular fingerprints of these vaccines were sub- jected to chemometric analysis through PARAFAC classification as well as XGBOOST discriminant analysis. The “most probable prediction” of unknown samples with 100% certainty was substantiated by the generated confusion matrix supporting the A-TEEM fluorescence claim to be a powerful addition to the arsenal of validation techniques. Two lots for each vaccine were measured on one instrument and were vali- dated with a different lot, instrument and operator. The data analysis approaches used were each able to differentiate between the vaccine products. Even Spectra-9 and Spectra-10, that differ ONLY by a coro- navirus component based on publicly available SDS data, were readily distinguished. Fluorescence EEMs (Excitation-Emission Matrix) solve the longstanding issue of imperfect quantification (a result of the Inner Filter Effect) by directly incorporating a UV spectrophotometer in the fluorometer. This allows the simultaneous acquisition in situ of a UV/VIS/NIR-absorbance spectrum for the real-time Inner Filter Effect (IFE) correction of the fluorescence spectrum, improving quantification accuracy and extend- ing the usable range of concentrations over which quantification can be performed. A-TEEM (Absorbance Transmission Excitation Emission Matrix) is fully validatable using United States Pharmacopeia mono- graph USP <853>, given that the novel aspect of simultaneous acquisi- tion of the UV/VIS/NIR absorbance spectrum for IFE correction is fully compatible with validation protocols. This spectroscopic approach provides a complete and traceable optical fingerprint for liquid samples that performs a similar role to chromatographic methods, and com- pared to other spectroscopic methodologies is faster, less expensive and can operate in production environments. Speaker Karoly Csatorday, Business Development Manager, Horiba Scientific

Bruker’s BioAFM Nano-Toolkit for Investigation of Me- chanics, Structures and Dynamic Processes in Life Science The ability of atomic force microscopy (AFM) to obtain three- dimensional topography images of biomolecules and complexes with nanometer resolution, and optical tweezers (OT) to measure sub-piconewton molecular forces under near-physiological condi- tions remains unmatched by other single molecule techniques. JPK BioAFM has developed the new NanoWizard® 4 XP and NanoRacer AFM’s, and Nanotracker2 (NT2) optical tweezers to set the bar even higher in these areas. The NW4XP is capable of high-speed study of the time-resolved dynamics associated with cellular processes; Nanoracer is designed for studying single molecule dynamics at 50 frames/sec; and NT2 can measure intra/inter molecular forces with multiple laser/trap options (up to 10 mW). With the latest scanner technologies inbuilt into NW4XP, NT2 and their compact design also allows full integration of AFM and OT into advanced commercially available light microscopy techniques. This seminar will focus on how the advances in Bruker’s latest BioAFM can be applied to study a wide-range of biological samples, from individual biomolecules to mammalian cells and tissues in real-time, in-situ experiments. We will present examples of how we are able to resolve the nanoscale struc- ture of individual biomolecules at high-speed scan rates (150 - 5000 Hz), follow the dynamic reorganization of the membrane-associated cytoskeleton of living cells at high-temporal and high-spatial resolu- tion, and automatically map the topography of cell cultures across the entire area of the microscope stage. We will also introduce our new scanning electrochemical microscopy (SECM) module and dis- cuss the full suite of BioAFM modes and accessories for studying the nanomechanical properties of cells and tissues, including direct cor- relation of multiparametric, quantitative AFM and super-resolution (STED) datasets. Speaker Samrat Dutta, Sales Applications Scientist, Bruker

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