Mapping sub-cellular protein aggregates and lipid inclusions using synchrotron ATR-FTIR microspectroscopy

Area of Science:

• Biochemistry
• Cell Biology
• Spectroscopy

Background:

• Sub-cellular biochemical analysis presents visualization challenges.
• Existing microscopy methods indirectly detect biochemical markers.
• Innovation is needed to directly image diverse biochemical parameters in situ.

Purpose of the Study:

• To advance synchrotron radiation attenuated total reflectance Fourier transform infrared (SR-ATR-FTIR) microspectroscopy for sub-cellular biochemistry.
• To demonstrate direct imaging of sub-cellular protein aggregates and lipid inclusions.
• To optimize spectral acquisition parameters for improved speed and quality.

Main Methods:

• Application of SR-ATR-FTIR microspectroscopy.
• Imaging of protein aggregates in degenerating neurons.
• Mapping of lipid inclusions in bacterial cells.
• Characterization of spectral resolution and detector effects on acquisition time.

Main Results:

• Direct sub-cellular mapping of protein aggregates and lipid inclusions achieved.
• An 8 cm^-1 spectral resolution offers a balance between spectral quality and collection time, improving acquisition speed by ~30%.
• Coupling a focal plane array detector with SR-ATR-FTIR improved image acquisition time by a factor of 2.8.

Conclusions:

• SR-ATR-FTIR microspectroscopy enables direct visualization of key sub-cellular biochemical components.
• Optimized acquisition parameters enhance the practicality of SR-ATR-FTIR for biological studies.
• This technique provides a foundation for broader adoption in biological sciences for complex biochemical investigations.