Spectrum of the cavity-QED microlaser: strong coupling effects in the frequency pulling at off resonance.

We report the first experimental observation of the cavity-QED microlaser spectrum, specifically the unconventional frequency pulling brought by a strong atom-cavity coupling at off resonance. The pulling is enhanced quadratically by the atom-cavity coupling to result in a sensitive response to the number of pumping atoms (2.1 kHz per atom maximally). Periodic variation of the pulling due to the coherent Rabi oscillation is also observed as the number of pumping atoms is increased across multiple thresholds.

Imaging human epithelial properties with polarized light-scattering spectroscopy.

Biomedical imaging with light-scattering spectroscopy (LSS) is a novel optical technology developed to probe the structure of living epithelial cells in situ without need for tissue removal. LSS makes it possible to distinguish between single backscattering from epithelial-cell nuclei and multiply scattered light. The spectrum of the single backscattering component is further analyzed to provide quantitative information about the epithelial-cell nuclei such as nuclear size, degree of pleomorphism, degree of hyperchromasia and amount of chromatin. LSS imaging allows mapping these histological properties over wide areas of epithelial lining. Because nuclear enlargement, pleomorphism and hyperchromasia are principal features of nuclear atypia associated with precancerous and cancerous changes in virtually all epithelia, LSS imaging can be used to detect precancerous lesions in optically accessible organs.

Reversible molecular adsorption based on multiple-point interaction by shrinkable gels.

A general approach is presented for creating polymer gels that can recognize and capture a target molecule by multiple-point interaction and that can reversibly change their affinity to the target by more than one order of magnitude. The polymers consist of majority monomers that make the gel reversibly swell and shrink and minority monomers that constitute multiple-point adsorption centers for the target molecule. Multiple-point interaction is experimentally proven by power laws found between the affinity and the concentration of the adsorbing monomers within the gels.

Quantification of (7S,8R)-dihydroxy-(9R,10S)-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene adducts in human serum albumin by laser-induced fluorescence: implications for the in vivo metabolism of benzo[a]pyrene.

The ubiquitous environmental carcinogen benzo[a]pyrene (BaP) is metabolized in vivo in humans to its ultimate carcinogenic form of 7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE). Mouse skin tumorigenicity studies indicate that the (7R,8S,9S,10R) enantiomer of BPDE, (7R,8S)-dihydroxy-(9S,10R)-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(7R,8S,9S,10R)-BPDE], is a potent tumor initiator, whereas the (7S,8R,9R,10S) enantiomer of BPDE, (7S,8R)-dihydroxy-(9R,10S)-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(7S,8R,9R,10S)-BPDE], may act as a tumor promoter. In vitro experiments have shown that human liver microsomes are capable of metabolizing BaP to both the (7R,8S,9S,10R) and (7S,8R,9R,10S) enantiomers of BPDE. However, the metabolism of BaP to (7S,8R,9R,10S)-BPDE has not been demonstrated in humans in vivo. The adducts formed between human serum albumin (HSA) and the (7S,8R,9R,10R) and (7R,8S,9S,10R) enantiomers of BPDE have been described previously. (7S,8R,9R,10S)-BPDE forms a stable adduct at histidine146 of HSA, whereas (7R,8S,9R,10R)-BPDE forms a relatively unstable ester adduct at aspartate187 or glutamate188 of HSA. Using high-performance liquid chromatography with laser-induced fluorescence (LIF) detector, we quantified the level of (7S,8R,9R,10S)-BPDE adducts at histidine146 in HSA isolated from 63 healthy males who were population control subjects for an ongoing case-control study of bladder cancer. By design, roughly half of the participants were lifelong nonsmokers (n = 35), whereas the remaining 28 participants were current smokers of varying intensities. HP-BPDE adducts were detected in 60 of the 63 samples (95%) by HPLC-LIF. Adduct levels ranged from undetectable (<0.04 fmol/mg HSA) to 0.77 fmol/mg HSA. The samples had a mean and median (7S,8R,9R,10S)-BPDE-HSA adduct level of 0.22 and 0.16 fmol of adduct/mg albumin, respectively. Mean adduct levels did not differ between smokers and nonsmokers (P = 0.72). Occupational exposure to polycyclic aromatic hydrocarbons was unrelated to adduct level (P = 0.62). Intake frequencies of two food items showed statistically significant associations with adduct levels. Consumption of sweet potatoes was negatively related to adduct level (P = 0.029), whereas intake of grapefruit juice was positively related to adduct level (P = 0.045). None of the three indices of residential ambient air pollution under study showed a statistically significant association with adduct levels.

Biochemical analysis and mapping of atherosclerotic human artery using FT-IR microspectroscopy.

We report the application of FT-IR microspectroscopy for in situ spectroscopic characterization of molecular constituents of human atherosclerotic lesions. Since water content in tissue affects conformation-sensitive protein vibrational bands, tissue specimens were examined under moist conditions. In all measurements, vibrational bands from water were found to dominate the spectrum. By removing these water contributions, well resolved bands due to tissue components were readily observed. Utilizing the high sensitivity and good spatial resolution of IR microspectroscopy, spectra from a sample volume of 40 x 40 x 4 microns3 were collected using unstained cryostat sections mounted on a BaF2 flat in neutral isotonic saline. Microstructures were confirmed histologically by light microscopy in stained serial sections. In the spectrum of normal intima, major bands due to amide I (1656 cm-1), amide II (1556 cm-1), and CH bending (1457 cm-1) vibrations of the proteins collagen and elastin were observed. In the spectrum of the intima of noncalcified atherosclerotic plaque, major bands due to both proteins and lipids were observed. The lipid bands at 1734, 1468, 1171 and 1058 cm-1 were assigned to the C = O (ester) stretch, CH2 bend, C--O (ester) stretch and C--O stretch, respectively. At a more detailed level, bands specific to free cholesterol, and cholesterol esters were identified. A plot of the integrated intensity ratio of these bands to the protein amide II mode versus depth from the luminal surface confirmed a heterogeneous distribution of these constituents in the atheromatous core. In the spectra of calcified atherosclerotic plaque, bands were attributed to three types of biochemical microstructures: proteins (1657, 1555, 1243 cm-1), lipids (1735, 1466, 1170, 1085, 1055 cm-1) and calcium minerals such as hydroxyapatite (1094, 1040, 962 cm-1), and carbonated apatite (1463, 1412, 872 cm-1). The results demonstrate that IR microspectroscopy can be used for in situ characterization of molecular constituents in human unstained arterial sections. The molecular information obtained from these studies could be important in understanding the pathogenesis of atherosclerosis.

Feasibility of field-based light scattering spectroscopy.

Light scattering spectroscopy (LSS) is a new technique capable of accurately measuring the features of nuclei and other cellular organelles in situ. We present the considerations required to implement and interpret field-based detection in LSS, where the scattered electric field is detected interferometrically, and demonstrate that the technique is experimentally feasible. A theoretical formalism for modeling field-based LSS signals based on Mie scattering is presented. Phase-front uniformity is shown to play an important and novel role. Results of heterodyne experiments with polystyrene microspheres that localize LSS signals to a region about 30 microns in axial extent are reported. In addition, differences between field-based LSS and the earlier intensity-based LSS are discussed.

Optical computed tomography in a turbid medium using early arriving photons.

We employ photon migration to image absorbing objects embedded in a turbid medium. For improved resolution, we use early arriving photons (a few hundred picoseconds in excess of the time of flight), a regime in which the diffusion approximation breaks down. Our image reconstruction method is based on extension of x-ray computed tomography (CT) to the optical regime. The CT algorithm must be generalized to take into account the distributions of photon paths. We express the point spread function (PSF) in terms of the Green's function for the transport equation. This PSF then provides weighting functions for use in a generalized series expansion method of x-ray CT. Experiments were performed on a turbid medium with scattering and absorption properties similar to those of human breast tissue. Multiple absorbers were embedded into the medium to mimic tumors. Coaxial transmission scans were collected in two projections, and the early-time portions were analyzed. Through accurate modeling, we could remove the blurring associated with multiple scattering and obtain high-resolution images. Our results show that the diffusion approximation PSF is inadequate to describe the early arriving photons. A PSF incorporating causality is required to reconstruct accurate images of turbid media.

Instrumentation for multi-modal spectroscopic diagnosis of epithelial dysplasia.

Reflectance and fluorescence spectroscopies have shown great promise for early detection of epithelial dysplasia. We have developed a clinical reflectance spectrofluorimeter for multimodal spectroscopic diagnosis of epithelial dysplasia. This clinical instrument, the FastEEM, collects white light reflectance and fluorescence excitation-emission matrices (EEM's) within a fraction of a second. In this paper we describe the FastEEM instrumentation, designed for collection of multi-modal spectroscopic data. We illustrate its performance using tissue phantoms with well defined optical properties and biochemicals of known fluorescence properties. In addition, we discuss our plans to develop a system that combines a multi-spectral imaging device for wide area surveillance with this contact probe device.

Fluorescence line-narrowing studies of antibody-benzo[a]pyrene tetrol complexes.

Benzo[a]pyrene tetrol (BPT) was used as a fluorescent probe to investigate the nature of antigen binding by two different monoclonal antibodies (MAb) that recognize a variety of derivatives of anti-7,8-dihydroxy-9,10-epoxy-7,8,9,10- tetrahydrobenzo[a]pyrenes (BPDE). Fluorescence line-narrowed spectra of the physical complexes of BPT formed with antibodies 8E11 and 3C3 were recorded at 4 K by employing vibronic excitation into the S1 electronic state. The frequencies of the vibrational modes of the S1 state were only marginally affected, though changes in relative intensities of some bands were observed. Fluorescence spectra recorded at 77 K by excitation into the S2 state showed that the (0,0) fluorescence emission of BPT was shifted to red on complex formation. Intensity ratios of the (0,0) band and the main vibrational band at 1300 cm-1 were used to assess the degree of interior binding of the chromophore. Quenching studies with acrylamide were employed to designate the complexes as type I, solvent inaccessible, or type II, solvent accessible. These studies also indicated that antibody 3C3 complexes tend to be more heterogeneous compared to the 8E11 complex. Deuterated BPT-d-12 also formed complexes with both antibodies, however, with different quenching behavior.

Raman spectroscopy and fluorescence photon migration for breast cancer diagnosis and imaging.

We are developing optical methods based on near infrared Raman spectroscopy and fluorescence photon migration for diagnosis and localization of breast cancer. We demonstrate the ability of Raman spectroscopy to classify accurately normal, benign and malignant breast tissues, an important step in developing Raman spectroscopic needle probes as a tool for improving the accuracy of needle biopsy. We also show that photon migration imaging can be used to localize accurately small fluorescent objects imbedded in a thick turbid medium with realistic optical properties, thus demonstrating the potential of this technique for optical imaging.

Near-infrared fluorescence spectroscopy detects Alzheimer's disease in vitro.

The purpose of this study was to investigate whether near-infrared (NIR) fluorescence spectroscopy could be used to detect Alzheimer's disease (AD) by brain tissue autofluorescence. Unfixed temporal cortex specimens from AD cases and age-matched, non-AD controls were frozen at autopsy and then thawed just prior to spectral measurement. Spectra of intrinsic tissue fluorescence induced by 647 nm light were recorded from 650 to 850 nm. We used principal component analysis of the tissue spectra from 17 AD cases and 5 non-AD control cases in a calibration study to establish a diagnostic algorithm. Retrospectively applied to the calibration set, the algorithm correctly classified 23 of 24 specimens. In a prospective study of 19 specimens from 5 AD brains and 2 non-AD control brains, 3 of the 4 control specimens and all AD specimens were correctly diagnosed. Both the excitation light used and the measured brain tissue autofluorescence are at NIR wavelengths that can propagate through skull and overlying tissue. Therefore, our results demonstrate an optical spectroscopic technique that carries direct molecular level information about disease. This is the first step toward a clinical tool that has the potential to be applied to the noninvasive diagnosis of AD in living patients.

Prospects for in vivo Raman spectroscopy.

Raman spectroscopy is a potentially important clinical tool for real-time diagnosis of disease and in situ evaluation of living tissue. The purpose of this article is to review the biological and physical basis of Raman spectroscopy of tissue, to assess the current status of the field and to explore future directions. The principles of Raman spectroscopy and the molecular level information it provides are explained. An overview of the evolution of Raman spectroscopic techniques in biology and medicine, from early investigations using visible laser excitation to present-day technology based on near-infrared laser excitation and charge-coupled device array detection, is presented. State-of-the-art Raman spectrometer systems for research laboratory and clinical settings are described. Modern methods of multivariate spectral analysis for extracting diagnostic, chemical and morphological information are reviewed. Several in-depth applications are presented to illustrate the methods of collecting, processing and analysing data, as well as the range of medical applications under study. Finally, the issues to be addressed in implementing Raman spectroscopy in various clinical applications, as well as some long-term directions for future study, are discussed.

Intensities of calcium dipicolinate and Bacillus subtilis spore Raman spectra excited with 244 nm light.

Ultraviolet (UV) resonance Raman spectra of Bacillus subtilis endospores have been excited at 244 nm. Spectra can be interpreted in terms of contributions from calcium dipicolinate and nucleic acid components. Differences between spectra of spores and vegetative cells are very large and are due to the dominance of the dipicolinate features in the spore spectra. Because the DNA and RNA composition of B. subtilis spores is known and because the cross-sections of Raman bands belonging to DNA and RNA bases are known, it is possible to calculate resonance Raman spectral cross-sections for the spore Raman peaks associated with the nucleic acids. The cross-sections of peaks associated with calcium dipicolinate have been measured from aqueous solutions. Cross-section values of the dominant 1017 cm(-1) calcium dipicolinate peak measured from the Bacillus spores have been shown to be consistent with a calcium dipicolinate composition of ten percent or less by weight in the spores. It is suggested that spectral cross-sections of endospores excited at 244 nm can be estimated to be the sum of the cross-sections of the calcium dipicolinate, DNA, and RNA components of the spore. It appears that the peaks due to DNA and RNA can be used as an internal standard in the calculation of spore Raman peak cross-sections, and potentially the amount of calcium dipicolinate in spores. It is estimated on the basis of known nucleic acid base cross-sections that the most intense Raman band of the Bacillus subtilis spore spectra has a cross-section of no more than 4 x 10(-18) cm(2)/mol-sr.

An overview of activities at the Laser Biomedical Research Center.

Research projects on laser heating and ablation and on spectroscopy of biological tissues are described. The discussion focuses on studies regarding microscopic laser light scattering of biological cells and structures, ablation of calcific tissue using pulsed HF laser radiation, and fluorescence and its use in diagnosing atherosclerosis. A specifically designed multifiber laser catheter constructed to collect tissue fluorescence spectra using fiber optics is described.

Effect of maternal low dose aspirin on neonatal platelet function.

OBJECTIVE: To evaluate the effect of maternal low dose aspirin ingestion in platelet function of newborn. DESIGN: Prospective randomized placebo controlled study. METHODS: 25 neonates born to mothers receiving low dose aspirin and 25 matched neonates with no maternal exposure to aspirin were studied. 2 ml of EDTA and 4.5 ml of citrate blood was collected from umbilical vein using double clamped umbilical stump for hemogram, coagulation profile and platelet functions. RESULTS: The platelet counts (10(9)/l) of study and control groups were 186.4 +/- 22.76 (116-225) and 205.28 +/- 17.34 (176-225), respectively. There was no significant difference in coagulation parameters. Prothrombin time index (PTI) was 86.24 +/- 6.623 and 87 +/- 6.43, respectively in the study and control group while PTTK (sec) was 55.88 +/- 20.54 and 52.12 +/- 11.82 in study and control subjects, respectively. The platelet aggregation studies (platelet function) with various platelet agonists in study and control group did not show any significant difference. Clinically, none of the babies had bleeding. CONCLUSIONS: Use of low dose aspirin in pregnant women was found to be safe and had no adverse effects on platelet functions of newborn.

Raman spectroscopy: a real-time tool for identifying microcalcifications during stereotactic breast core needle biopsies.

Microcalcifications are an early mammographic sign of breast cancer and a target for stereotactic breast needle biopsy. We present here a Raman spectroscopic tool for detecting microcalcifications in breast tissue based on their chemical composition. We collected ex vivo Raman spectra from 159 tissue sites in fresh stereotactic breast needle biopsies from 33 patients, including 54 normal sites, 75 lesions with microcalcifications and 30 lesions without microcalcifications. Application of our Raman technique resulted in a positive predictive value of 97% for detecting microcalcifications. This study shows that Raman spectroscopy has the potential to detect microcalcifications during stereotactic breast core biopsies and provide real-time feedback to radiologists, thus reducing non-diagnostic and false negative biopsies.

Observation of sub-poisson photon statistics in the cavity-QED microlaser.

We have measured the second-order correlation function of the cavity-QED microlaser output and observed a transition from photon bunching to antibunching with increasing average number of intracavity atoms. The observed correlation times and the transition from super- to sub-Poisson photon statistics can be well described by gain-loss feedback or enhanced-reduced restoring action against fluctuations in photon number in the context of a quantum microlaser theory and a photon rate equation picture. However, the theory predicts a degree of antibunching several times larger than that observed, which may indicate the inadequacy of its treatment of atomic velocity distributions.

Traveling-wave atom cavity interaction in the single-atom microlaser.

We demonstrated traveling-wave atom-cavity interaction in the single-atom microlaser by tilting the atomic beam from its usual orientation of normal incidence with respect to the cavity mode. Laser-tuning curves, measured for various excitation pulse areas, are in good agreement with one-atom microlaser-maser theory.