Electric field standing wave artefacts in FTIR micro-spectroscopy of biological materials

DOI: 10.1039/c2an15995c
PMID: 22231204

Authors: Jacob Filik (Diamond Light Source) , Mark D. Frogley (Diamond Light Source) , Jacek K. Pijanka (Keele University) , Katia Wehbe (Diamond Light Source) , Gianfelice Cinque (Diamond Light Source)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Analyst

State: Published (Approved)
Published: January 2012

Abstract: FTIR absorption micro-spectroscopy is a widely used, powerful technique for analysing biological materials. In principle it is a straightforward linear absorption spectroscopy, but it can be affected by artefacts that complicate the interpretation of the data. In this article, artefacts produced by the electric-field standing-wave (EFSW) in micro-reflection-absorption (transflection) spectroscopy are investigated. An EFSW is present at reflective metallic surfaces due to the interference of incident and reflected light. The period of this standing wave is dependent on the wavelength of the radiation and can produce non-linear changes in absorbance with increasing sample thickness (non-Beer–Lambert like behaviour). A protein micro-structure was produced as a simple experimental model for a biological cell and used to evaluate the differences between FTIR spectra collected in transmission and transflection. By varying the thickness of the protein samples, the relationship between the absorbance and sample thickness in transflection was determined, and shown to be consistent with optical interference due to the EFSW coupled with internal reflection from the sample top surface. FTIR spectral image data from MCF 7 breast adenocarcinoma cells was then analysed to determine the severity of the EFSW artefact in data from a real sample. The results from these measurements confirmed that the EFSW artefact has a profound effect on transflection spectra, and in this case the main spectral variations were related to the sample thickness rather than any biochemical differences.

Diamond Keywords: Breast Cancer

Subject Areas: Technique Development, Biology and Bio-materials


Instruments: B22-Multimode InfraRed imaging And Microspectroscopy

Added On: 10/01/2012 11:31

Discipline Tags:

Non-Communicable Diseases Health & Wellbeing Cancer Technique Development - Life Sciences & Biotech Life Sciences & Biotech

Technical Tags:

Spectroscopy Infrared Spectroscopy Synchtron-based Fourier Transform Infrared Spectroscopy (SR-FTIR)