IR spectroscopy is an excellent method for biological analyses. It enables the nonperturbative, label-free extraction of biochemical information and images toward diagnosis and the assessment of cell functionality. Although not strictly microscopy in the conventional sense, it allows the construction of images of tissue or cell architecture by the passing of spectral data through a variety of computational algorithms. Because such images are constructed from fingerprint spectra, the notion is that they can be an objective reflection of the underlying health status of the analyzed sample. One of the major difficulties in the field has been determining a consensus on spectral pre-processing and data analysis. This manuscript brings together as coauthors some of the leaders in this field to allow the standardization of methods and procedures for adapting a multistage approach to a methodology that can be applied to a variety of cell biological questions or used within a clinical setting for disease screening or diagnosis. We describe a protocol for collecting IR spectra and images from biological samples (e.g., fixed cytology and tissue sections, live cells or biofluids) that assesses the instrumental options available, appropriate sample preparation, different sampling modes as well as important advances in spectral data acquisition. After acquisition, data processing consists of a sequence of steps including quality control, spectral pre-processing, feature extraction and classification of the supervised or unsupervised type. A typical experiment can be completed and analyzed within hours. Example results are presented on the use of IR spectra combined with multivariate data processing.

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Using Fourier transform IR spectroscopy to analyze biological materials

Surface infrared spectroscopy

Chemical imaging of biological tissue with synchrotron infrared light

In this study, we conducted synchrotron radiation Fourier transform infrared (IR) spectroscopy, potentiometric titration, and metal sorption experiments to characterize metal-cyanobacteria sorption reactions. Infrared spectra were collected with samples in solution for intact cyanobacterial filaments and separated exopolymeric sheath material to examine the deprotonation reactions of cell surface functional groups. The infrared spectra of intact cells sequentially titrated from pH 3.2 to 6.5 display an increase in peak intensity and area at 1400 cm(-1) corresponding to vibrational COO- frequencies from the formation of deprotonated carboxyl surface sites. Similarly, bulk acid-base titration of cyanobacterial filaments and sheath material indicates that the concentration of proton-active surface sites is higher on the cell wall compared to the overlying sheath. A three-site model provides an excellent fit to the titration curves of both intact cells and sheath material with corresponding pKa values of 4.7 +/- 0.4, 6.6 +/- 0.2, 9.2 +/- 0.3 and 4.8 +/- 0.3, 6.5 +/- 0.1, 8.7 +/- 0.2, respectively. Finally, Cu2+, Cd2+, and Pb2+ sorption experiments were conducted as a function of pH, and a site-specific surface complexation model was used to describe the metal sorption data. The modeling indicates that metal ions are partitioned between the exopolymer sheath and cell wall and that the carboxyl groups on the cyanobacterial cell wall are the dominant sink for metals at near neutral pH. These results demonstrate that the cyanobacterial surfaces are complex structures which contain distinct surface layers, each with unique molecular functional groups and metal binding properties.

http://homepages.see.leeds.ac.uk/~earlgb/Publications/Yee_et_al_EST_2004.pdf

Characterization of metal-cyanobacteria sorption reactions: a combined macroscopic and infrared spectroscopic investigation

Electromagnetic waves centred at a frequency of 1 THz lie between photonics on the one hand and electronics on the other, and are very hard to generate and detect. However, since the THz part of the spectrum is energetically equivalent to many important physical, chemical and biological processes including superconducting gaps and protein dynamical processes, it is of great interest to facilitate experimental research in this region. This has stimulated major steps in the past decade for filling this gap in the usable spectrum. In this review paper we describe the evolution of a new generation of sources that boost the average power available in the THz region by more than a million-fold, making this region routinely accessible for the first time. This is achieved using two enhancement factors, namely relativistic electrons and super-radiance. We will also point to the scientific potential for discovery that is now enabled in this region.

https://iopscience.iop.org/article/10.1088/0034-4885/69/2/R01/pdf

Filling the THz gap—high power sources and applications

Simple and useful approximations, valid at infrared wavelengths, to the equations for synchrotron radiation are presented and used to quantify the brightness and power advantage of current synchrotron radiation light sources over conventional infrared broadband laboratory sources. The Daresbury Synchrotron Radiation Source (SRS) and the Brookhaven National Synchrotron Light Source (vacuum ultraviolet) [NSLS(VUV)] storage rings are used as examples in the calculation of the properties of infrared synchrotron radiation. The pulsed nature of the emission is also discussed, and potential areas of application for the brightness, power, and time structure advantages are presented. The use of infrared free electron lasers and undulators on the next generation of storage ring light sources is briefly considered.

W. D. Duncan

Papers

Infrared synchrotron radiation from electron storage rings.