Emerging Applications Endoscopic Guided Light Reflectance Absorption Analysis

One of the problems associated with direct visualization via endoscopy when attempting to identify desiccated tissue or organ types is that desiccated tissue or organ morphology and anatomical orientation are typically altered, making identification difficult if not impossible. The application of light reflectance theory in this setting rests in the construct that all tissues and organs are chemically different and may absorb and reflect different wavelengths of the spectra. If the theory is correct, we hypothesized that various organs and tissues may have unique wavelength absorption or reflectance signatures.

Mummified tissues and organs are subject to complete or partial desiccation, resulting in a loss of morphological characteristics. In addition, different embalming methods result in varied states of decomposition. Many cultures also practiced some form of tissue or organ removal and at times replaced those organs within body cavities. The impact of desiccation, state of decomposition, and organ removal or replacement all lead to a loss of anatomical landmarks often used to identify those organs (Aufderheide and Rodriguez-Martin 1998). Differentiation among organs through direct observation is greatly challenged by these variables.

Current analytical methods used to differentiate tissue or organ remains include tissue and organ sampling or sectioning, rehydration for histological analysis, elemental and chemical analysis, and pathological and cellular analyses (Cockburn et al. 1998). Each of these methods alters any remaining internal anatomical context and is therefore a destructive technique.

We conducted an initial project (Beckett et al. 2007a) whose objective was to determine if tissue- or organ-type differentiation could be made from nondestructive light reflectance or absorption methodologies when applied to tissues or organs from varied mummification and preservation methods.

We established the following research questions:

1. Do different tissue types absorb or reflect light of varied wavelengths?

2. Do different organ systems absorb or reflect light of varied wavelengths?

3. If different tissue types or organ systems absorb or reflect light at varied wavelengths, is there variance in light reflectance signatures among mummified samples?

Our methods, materials, and background research regarding light wave reflectance included an examination of the theory of light reflectance and absorption, which is grounded in color theory.

Color theory demonstrates that the human eye is imperfect, in that it cannot differentiate among varied shades, and there are areas of the light spectrum that cannot be detected. What is absorbed and what is reflected is determined by the chemistry of the material. The application of light wave absorption and reflection is used in living tissue in medical science and analytical chemistry. One medical example is the varied light wave characteristics of hemoglobin (Hb) when compared to oxy-Hb (oxygen attached to the

Hb molecule). Each state of Hb has different chemical characteristics. This is also true for alternate states of Hb such as carboxy-Hb, Met-Hb, Sulf-Hb, and deoxy-Hb (Ruppel 2005). Each substance can be differentiated from the others by using its unique light wave absorption and reflection signatures. This is accomplished by using an instrument known in medicine as a Co-Oximeter, which employs the spectrophotometry theory of light wave absorption. Light of varied wavelengths is sent through the sample. Those light waves that are not absorbed by the substance are measured by a photomultiplier and quantified. The absorbed wavelengths represent the specific absorption signature based on the chemistry of the substance under study.

Another common application using reflectance is found in the reflectance pulse oxi-meter used to determine the degree to which Hb is saturated with oxygen. This instrument is placed on the forehead of newborns or infants. Light waves specific for the Hb molecule and for saturated Hb molecules are shown through the surface of the skin. The light waves that are not absorbed are reflected back to the photomultiplier from the frontal bone, and the determination of the percentage of Hb saturation is derived.

As an integral part of our methods, we established our experimental design. A reflectance probe was used to both emit light waves and to collect the reflected nonabsorbed light waves. We first applied the reflectance probe to a preserved laboratory feline and examined the results. After examining the results, we then applied the reflectance probe to mummified tissues and organs under endoscopic guidance with radiographic positional correlation. Our subjects for this stage of the experimental design were a 100-year-old North American sideshow mummy and a 2500-year-old Egyptian mummy. We developed a plan to combine the technologies of the reflectance probe with endoscopy, which was used for reflectance probe guidance into internal body cavities, and radiography, which was used for probe positional documentation. We then collected and compared data derived from various organs and tissue types including cardiac, vascular, skeletal, renal, pulmonary, hepatic, gastrointestinal, and dental. We developed a logical uniform reporting and data comparison method as related to the reflectance data. We recorded the nanometer value at peak percent reflectance reading. We then refined our methods and resolved any technological or data collection problems. Finally, we repeated the experiment to assess the reproducibility of observed data and, ultimately, determine if there were any significant variations or correlations among and between subjects.

In order to collect the required data, we employed three major technologies (Figure 4.40) in the course of this experiment:

Figure 4.40 combined paleoimaging technologies. shown here, from left to right, are standard radiography, reflectance instrumentation, and videoendoscopy system.

1. Videoendoscopy was used for reflectance probe placement and location. Also, it was used to document the target organ or tissue and correlate the location with existing anatomical landmarks. We employed a fiber-optic VE, light source, camera control unit, and digital video data collection system.

2. Radiography was used to document probe location and correlate its position with anatomical landmarks using standard or instant film imaging and advanced imaging as needed.

3. Reflectance instrumentation was employed to collect reflectance data using a light guide or receiver probe, light source, and a spectrometer with a computer interface. The reflectance system was calibrated to absolute white and absolute black.

As we began our experimentation, several application problems arose that required minor technological modifications. The first problem was keeping a standard and consistent distance between the probe and the tissue or organ target, thereby controlling for distance as a variable. To eliminate this variable, a 4 mm section on nonreflective quartz was affixed to the probe tip, ensuring a constant distance from probe tip to the target tissue or organ. Unfortunately, this modification using the quartz tip allowed ambient light contamination to enter the probe tip from the lateral aspect of the quartz, altering the reflectance data. To correct for this problem, we protected the lateral aspects of the quartz tip by using a thin rubber sheath, thus shielding the circumferential aspect of the nonreflective quartz and eliminating the ambient light contamination.

The results from four separate experiments were promising.

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