[R01] Development of nanoscale infrared spectroscopic imaging for measuring cellular ultrastructure
Ente: National Cancer Institute
Scadenza: 2030-07-31
Importo max: 509.352 EUR
Paese: US
Descrizione
ABSTRACT
Electron and fluorescence microscopy now provide exquisite structural (~ nm) information using specific,
labeled molecular species. However, only a small fraction of cellular species can be illuminated in any
experiment and labeling techniques work primarily for large molecules and assemblies. Small molecules, such
as metabolites, remain largely hidden without significant effort (e.g., single cell mass spectrometry). Molecular
vibrational spectroscopies (infrared and Raman) offer a unique opportunity to study all molecules of life directly
but are typically limited in the far field by optical resolution, suffer from low sensitivity when sampling volumes
are reduced to nm scale or do not yet offer consistent, reproducible data for near-field imaging. Atomic Force
Microscopy (AFM) combined with Infrared (IR) spectroscopy provides a powerful tool for label-free chemical
imaging with nanoscale resolution. There were three main challenges that limited AFM-IR performance: (a)
mechanical artifacts coupled the morphology and chemical domains, preventing chemical analyses, (b) high
noise reduced analytical accuracy and precision and (c) low sensitivity precluded quantification of multiple
species. Consequently, recorded data of biological materials were restricted to studies using point spectra to
avoid image artifacts, used thick samples (~micron scale) for sufficient sensitivity or examined well-described
and simple domains like amyloid deposits. Recent scientific advances have paved a path to overcome these
limitations; however, legacy instruments cannot take advantage of these advances. Hence, in this proposal we
will develop a new AFM-IR technology, dubbed resonance enhance null-deflection IR (RENDIR) spectroscopic
imaging. RENDIR will include custom electronics for controlling the instrument based on our recent closed loop
null deflection technique, optimized data recording using theoretical understanding of the image formation
process, an optimized optical train with expanded range quantum cascade lasers, and novel optics integrated
into a custom designed AFM. These advances will enable high-quality AFM-IR signals for all types of samples
and by users who are not necessarily doctoral level experts. This innovative technology will be tested using
gold standard samples and its limits characterized using fabricated polymeric samples of known composition
and geometry as well as biomedical samples. Lithographically patterned polymer samples will first be used to
validate both spectral and spatial performance. An isogenic cancer progression cell line will be used as a
biological model, wherein all subcellular domains and local metabolites in cytoplasmic regions will be identified.
The end of this project period will yield an AFM-IR measurement technology (RENDIR) that significantly
surpasses state of the art by being free of mechanical artifacts, allowing ~5 nm (~64-fold smaller pixels)
resolution, images with ~25x lower noise (due to null defle
Istituzione: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
PI: Rohit Bhargava
Progetto: 5R01CA304244-02
Settori: National Cancer Institute
Vai al bando originale
Registrati gratis su Bandolo per trovare bandi compatibili con la tua azienda.