Figure 1: Representative NIR spectra of vein blood on gauze aged over a period of one month.

Blood samples from human vein and capillary vessels were placed on different substrates and monitored in the NIR region over a one-month period. The composition of blood varies between veins and capillaries. The dissimilarities depend upon how a vessel stores and transports blood through the body as well as the distribution of cells and plasma in the vessels. The components of blood that are suitable for determining blood degradation as a function of time must be understood, and their effects on optical properties verified.

Materials and Methods

Human blood from vein and capillary vessels was used for this investigation. The blood samples originated from people of various age groups and both genders. Acquired blood samples were prepared and measured by a person trained and certified by OSHA for blood-borne pathogens.

Figure 2: Representative NIR spectra of capillary blood on gauze aged over a period of one month.

The bloodstain samples were prepared by depositing approximately six drops of blood on solid or porous substrates. A watch glass was used as a solid substrate and the strip of gauze placed on a glass microscope slide was used as a porous substrate. NIR spectra of 3-mm diameter samples were measured periodically at ambient conditions over a period of one month. The samples were stored at room temperature of 20 °C between measurements. An NIRSystems (FOSS, Inc., Laurel, Maryland) Model 5000 Rapid Content Analyzer was used to measure the diffuse reflection spectra of the samples. A total of 64 scans was measured over the spectral range of 1100–2500 nm. These scans were averaged to provide the final spectrum. Reference spectra of each of the substrates were measured through the same aperture with a TiO₂ diffusion plate placed behind the substrate. Spectra of lyophilized crystalline hemoglobin (Sigma-Aldrich, St. Louis, Missouri) were measured as a reference. This sample was reported to contain ~70% methemoglobin.

Results and Discussion

Figure 3: Representative NIR spectra of vein blood aged on glass aged over a period of one month.

Samples of vein and capillary blood from four volunteers were aged over a period of one month. Typical spectra of blood aging on a gauze substrate are shown in Figures 1 and 2, whereas blood on a glass substrate is shown in Figures 3 and 4. As is immediately obvious from the four figures, water is the major contributor to the spectra before aging. The experiments were performed under ambient conditions and the humidity was not controlled. In the case of the blood samples on glass, the bands due to water disappeared after 1 h of aging. Spectra of the blood on the gauze substrate exhibited evidence of water after 1 h. This is most evident in Figure 1, where the H₂O bands at ~1400 and ~1900 nm are still present in the 1-h spectrum. The band at ~1400 nm disappears into the background in most of the spectra shown in the figures. However, the band at 1900 nm exhibits some variation as the samples age. For example, in Figure 1 the band at 1900 nm is rather pronounced in the 1-h spectra, appears only as a shoulder at 25 h, but reappears at 189 h. The water bands in spectra of samples on the glass substrate disappeared rather quickly; however, the band at 1900 nm continued to vary slightly during the entire study.