Accurate blood flow measurements during surgery can improve an operation's chance of success. We developed near-infrared spatio-temporal image spectroscopy (NIR-STICS), which has the potential to make blood flow measurements that are difficult to accomplish with existing methods. Specifically, we propose the technique and we show feasibility on phantom measurements. NIR-STICS has the potential of measuring the fluid velocity in small blood vessels (less than 1 mm in diameter) and of creating a map of blood flow rates over an area of approximately 1 cm(2). NIR-STICS employs near-infrared spectroscopy to probe inside blood vessel walls and spatiotemporal image correlation spectroscopy to directly-without the use of a model-extract fluid velocity from the fluctuations within an image. We present computer simulations and experiments on a phantom system that demonstrate the effectiveness of NIR-STICS.

One of the mayor complications during vascular surgery is the formation of blood clot. We tested an animal model and assembled a prototype device that shows the blood clot signature spectrum and its temporary growth. © 2006 OSA/BOSD, AOIMP, TLA 2006.

Scanning laser image correlation (SLIC) is an optical correlation technique for measuring the fluid velocity of particles suspended in a liquid. This technique combines laser scanning of an arbitrary pattern with pair cross-correlation between any two points in the pattern. SLIC overcomes many of the limitations of other optical correlation techniques for flow measurement, such as laser speckle, spatial temporal image correlation spectroscopy, and two-foci methods. One of the main advantages of SLIC is that the concept can be applied to measurements on a range of scales through simple zooming or modifications in the instrumentation. Additionally, SLIC is relatively insensitive to instrument noise through the use of correlation analysis and is insensitive to background. SLIC can provide detailed information about the direction and pattern of flow. SLIC has potential applications ranging from microfluidics to blood flow measurements.

The emergence of time-resolved fluorescence techniques has been very important to the study of biochemical, biophysical and biomedical research areas. Many of the advances in these fields have been technology driven. The Laboratory for Fluorescence Dynamics (LFD) has been actively involved in development of frequency domain technology for a variety of applications. Our recent work on microwave super heterodyning detectors, fluorescence lifetime resolved spectroscopy, frequency domain fluorescence imaging microscopy, fluorescence lifetime resolved stopped-flow kinetics and global methods of data analysis is summarized in this report. We provide examples of how these new technologies are applied.

Giant unilamellar vesicles (GUVs) have been widely used as a model membrane system to study membrane organization, dynamics, and protein-membrane interactions. Most recent studies have relied on imaging methods, which require good contrast for image resolution. Multiple sequential image processing only detects slow components of membrane dynamics. We have developed a new fluorescence correlation spectroscopy (FCS) technique, termed scanning FCS (i.e., SFCS), which performs multiple FCS measurements simultaneously by rapidly directing the excitation laser beam in a uniform (circular) scan across the bilayer of the GUVs in a repetitive fashion. The scan rate is fast compared to the diffusion of the membrane proteins and even small molecules in the GUVs. Scanning FCS outputs a "carpet" of timed fluorescence intensity fluctuations at specific points along the scan. In this study, GUVs were assembled from rat kidney brush border membranes, which included the integral membrane proteins. Scanning FCS measurements on GUVs allowed for a straightforward detection of spatial-temporal interactions between the protein and the membrane based on the diffusion rate of the protein. To test for protein incorporation into the bilayers of the GUVs, antibodies against one specific membrane protein (NaPi II cotransporter) were labeled with ALEXA-488. Fluorescence images of the GUVs in the presence of the labeled antibody showed marginal fluorescence enhancement on the GUV membrane bilayers (poor image contrast and resolution). With the application of scanning FCS, the binding of the antibody to the GUVs was detected directly from the analysis of diffusion rates of the fluorescent antibody. The diffusion coefficient of the antibody bound to NaPi II in the GUVs was approximately 200-fold smaller than that in solution. Scanning FCS provided a simple, quantitative, yet highly sensitive method to study protein-membrane interactions.

Frequency domain spectroscopy provides a quantitative measure of the optical properties, namely µa and µs' coefficient spectra, of multiply scattering, macroscopically homogeneous media or tissue. The diffusion model for photon transport provides the theoretical framework for the analytical expressions of the optical properties. Experimentally, we intensity modulated (60 MHz) a light emitting diode, which emits between 620-700 nm. From data sets of relative phase shifts and average intensity at different source-detector separations, we calculated on the basis of the analytical expressions a wavelength resolved absorption µa (?) and scattering µs' (?) coefficient spectrum. The test material was methylene blue, whose absorption spectrum (maximum 656 nm) closely matches the wavelength profile of the diode source. The multiply scattering, macroscopically homogeneous medium for dissolving the methylene blue was provided by a diluted fat emulsion, Liposyn III. The concentrations ofboth the absorbing and scattering materials were adjusted to correspond to ranges typical of µa and µs tissues. We obtained quantitative agreement between the measured µa (?) the scattering medium and a control solution measured in a spectrophotometer under non-scattering conditions.

Using a broadband spectral technique, we recently showed [J. Biomed. Opt. 10, 064009 (2005)] that during visual stimulation of the cat brain there were not only changes in oxy- and deoxyhemoglobin levels, reminiscent of the optical blood oxygenation level dependence (BOLD) effect reported in humans, but also the apparent water content of the tissue and the optical scattering contribution decreased during stimulation. These relatively fast changes (in seconds) in water tissue content are difficult to explain in physiological terms. We developed a simple model to explain how local vasodilation, which occurs as a result of the stimulation, could cause this apparent change in water content. We show that in a phantom model we can obtain spectral effects similar to those observed in the cat brain such as the apparent decrease of the water spectral component without changing the water content of the bath in which the phantom measurements were performed. Furthermore, using the phantom model, we show that the relative apparent changes in the spectral components due to vasodilation during stimulation are roughly comparable in magnitude to the changes in tissue chromophores due to the optical equivalent of the BOLD effect reported in the literature.

Introduction

Reports indicate that brain regulation of oxygenation is inhibited in patients with low baseline oxyhemoglobin concentrations and that brain oxyhemoglobin concentrations are decreased with aging. The purpose of this study was to determine if regulation of brain oxygenation to changes in blood pressure is inhibited by normal aging.

Methods

Brain oxyhemoglobin (OHb) and deoxyhemoglobin (HHb) concentrations were determined from the forehead using a frequency domain near infrared spectroscopy in 27 healthy volunteers. Subjects were separated into two groups by age (20-39, n=16; 40-60, n=11). Brain hemoglobin and non-invasive blood pressure were measured in (1) supine, (2) sitting, (3) supine and (4) sitting positions with 10-min equilibration intervals between each determination. Statistical differences were determined by two way repeated measures analysis of variance.

Results

Young subjects were 28+/-5 years (mean+/-S.D.) and older subjects were 48+/-6 years. In supine position, OHb and HHb were 28.4+/-8.3 and 15.4+/-2.4micromol/L, respectively, in young; 22.4+/-5.7 and 13.4+/-2.9micromol/L, respectively, in older subjects, both P<0.05 between groups. Changing position from supine to sitting decreased OHb 5% and increased HHb 5% with no difference between groups.

Conclusions

There was a small but significant decrease in OHb and an increase in HHb from supine to sitting position, and this effect was similar between young and older subjects. Regulation of brain oxygenation during modest decreases in blood pressure did not change in normal aging to 60 years compared to young adults.