**3.3 Hair analysis**

Hair can prove to be a useful evidence in crimes in relation to determine the history of drug intake and abuse as well as exposure to toxins as the chemical composition of hair does not change by the external environment. Hair is the most encountered evidence in a forensic investigation and can act as a good source of DNA. The mitochondrial DNA present in the hair shaft and nuclear DNA is mostly within

*Atomic Force Microscope in Forensic Examination DOI: http://dx.doi.org/10.5772/intechopen.104704*

#### **Figure 5.**

*Duplicator paper (1), copper printing paper (2), glassine paper (3) and kraft paper (4); (a) topographic image, (b) amplitude image.*

the root sheath paly important role in DNA examination [35]. AFM offers unique advantages for analysis of hair surface, primarily due to the high image resolution as well as an ease of sample preparation. Durkan and Wang employed atomic force microscopy in a forensic approach to distinguish between different hair care products on the basis of the deposits left behind. They studied AFM techniques on hair samples that which were washed/treated with a number of different shampoos/conditioners and 2-in-1 products as shown in **Figure 6**. They found that the exocuticle carries a negative charge and gets deposits on unwashed hair with a mean roughness of up to 50 nm. Further they found that washing hair with shampoos reduces the roughness of hair + deposits to typically below 10 nm also the 2-in-1 products, conditioners or shampoos shows deposits that cover the entire surface, with roughness up to 30 nm. They concluded that the measurement of surface roughness combined with images of the resulting surface deposits can prove effective to distinguish between the effects of different hair care products [17].

The surface topography of human hair is defined by the cuticles which helps in cosmetic properties determination of the hair. The cuticles condition has the potential to aid in the medical diagnosis and forensic sciences. AFM offer unique advantages in hair surface analysis as it provides high resolution image and the simplicity of sample preparation. Gurden used an algorithm for the automatic examination of AFM images of human hair. By using a series of descriptors such as tilt angle, step height and cuticle density, the cuticular structure of hair was characterized and quantitatively investigated. They studied 38 AFM images consisting of hair samples untreated and bleached hair samples along with examination of the root and distal ends of the hair fiber. The multivariate classification technique partial least squares discriminant analysis was used to test the capability of the algorithm for further characterization of the images according to the hair properties. They were able to classify 86% hair images correctly. They study the classification of hair properties based on several cuticular descriptors by calculating it form the height images of various hair parts. The cuticular descriptors provided information on hair surface properties which can be correlate between the hair structure characteristics and environmental conditions

#### *Electron Microscopy*

#### **Figure 6.**

*(a) AFM image of unwashed human hair. Scale bar is 5 lm. (b) The same image after flattening, where the debris on the hair is now more prominent. (c) Side-lit 3D representation of an area of hair revealing that particulates and deposits are relatively uniformly distributed across the hair surface. Scale bar is 3 lm. (d) Smaller scale image showing deposits on hair, away from particulates and cuticle edges. Scale bar is 200 nm. (e) Cross-section through topography of a 35 lm long section of hair with four cuticle edges in the range 200– 500 nm thick with a lateral spacing approximately 7 lm, and where the deposits can be seen as ripples/bumps. (f) AFM topography image revealing woodgrain striations, characteristic of the exocuticle. (g) Cross-section through a clean hair devoid of deposits, for comparison to Figure 3e.*

the hairs are exposed to. Though the direct forensic relevance of this work was not established but the study do created extensive database of hair image along its mechanical properties [36].

Jeong et al. [18] have given an interesting contribution by studying the effects of aging on normal Korean hair diameter and surface features using AFM. They

#### *Atomic Force Microscope in Forensic Examination DOI: http://dx.doi.org/10.5772/intechopen.104704*

examine 60 Korean volunteers of various ages who had no hair diseases and studied hair diameter, hair surface, cuticular descriptors and micro-scale mechanical properties to determine their associations with aging. They found that hair diameter increases for the first 20–30 years of life and later showed decrease. AFM images of most of the younger subjects showed clear scale edges of hair while of older subjects revealed dilapidated structures, poorly defined scale edges and undulated surfaces. The cuticular descriptors, surface roughness showed increase significantly with age. Also the force to distance analysis confirmed its dependence on age. They concluded that aging causes changes in hair diameter and surface structure. These work done by Jeong et al. do contribute in estimating the age from forensic trace evidences like hair. The hair surface area studies were done by Tomes et al. using both SEM and AFM which showed little difference in quality of surface profiles obtained. For forensic hair imaging, the minimally invasive AFM technique can be preferred over SEM [37].

AFM is also used to investigate the effects of ethnicity, fatigue and water absorption on the tensile strength of hair and found in different ethnic hair types namely Caucasian, Asian and African and the results indicated that they have different mechanical properties [38]. Seshadri did the similar study on the tensile strength of hair by imaging the cuticular structure of hair. They found that hair shows stress-strain curve for keratinous fiber. Also the chemical, mechanical damage and conditioner treatment does not have any effect on the stress-strain curve or its tensile properties [39]. DelRio and Cook [19] provided interesting data of hair samples untreated virgin hairs and conditioned and bleached hairs. They stated an indentation modulus of 2.4 ± 1.1 GPa and 1.8 ± 0.9 GPa respectively for virgin and the bleached hairs samples while for the conditioned hairs, the indentation modulus varied between 0.05 and 0.5 GPa. They performed all the measurements on a 5 by 5 μm area.

#### **3.4 Diatom test**

Diatoms are a group of algae found in oceans and fresh waters possessing tough silica wall (SiO2) which is resistant to decay. Diatoms plays very crucial role in cases of drowning to determine whether it is antimortem or postmortem drowning, hence proving useful in forensic investigation. The recovery of diatoms from different organs, their quantitative and qualitative composition examination prove to be very trustworthy proof to determine the place and time of drowning in many cases [40]. They are studied in forensic geoscience in relation to transfer from different environments to clothing to obtain information of the crime scene and the perpetrator [41]. Newer techniques namely nuclear magnetic resonance, AFM, inductively coupled plasma (ICP) hyphenated technologies, fluorimetry and automatic diatom identification and classification are also been used for diatom study. AFM is used to study the diatoms morphological characteristics which can act as an indicator of its location, its growth cycle henceforward demonstrating its usefulness in forensic application. AFM has the potential to differentiate diatoms on the basis of its feature and can individualized atoms by scanning the objects that are 8″ long and having a diameter of 0.5″. Even the largest diatoms can be scanned in this range also the technique has the additional advantage of scanning the object in vertical and horizontal axis [42].

Almqvist et al. explored the possibilities of AFM to study diatoms in relation to its biomineralization and micromechanical properties. They studied the silica shell of the diatom Naviculapelliculosa (Bréb.) Hilse. The structure was imaged and the shell's micromechanical properties were studied in semi-quantitatively manner. The results indicated that the diatom's overall hardness and elasticity are same as that of silicas.

**Figure 7** shows the separated epitheca and hypotheca of one cell. They also showed that certain areas of the shell were significantly harder or more elastic which can be detect in different crystalline phases [43].
