METHOD FOR FINGERPRINTS AGE DETERMINATION

Gheorghe Popa1, Romic Potorac2, Nicolae Preda3 1 General Inspectorate of Romanian Police, Bucharest, Romania 2 DNA Departament, Forensic Institute, Bucharest, Romania 3 Forensic Service, Giurgiu, Romania
Abstract: The aim of this study was to establish an estimation relationship of the age of fingerprints left on surfaces, by morphological, structural, macro- and microscopic examinations, together with biochemical and titration DNA tests in order to confirm the rate of biological degradation during a certain period. The capacity of counting the age of a fingerprint lead to the possibility to place it in time and to correlate it with the time of doing the criminal act, bringing us information about the presence of a person in a certain place and period. As research methods we used forensic techniques for fingerprints, as well as cytology and molecular biological methods (DNA analysis, DNA quantification with TaqMan using Real Time PCR). The estimation of the age of fingerprints using these methods offers us the advantages of standardization based on relationships between morphological or/and biochemical characteristics depending on time, as well as the possibility to assign as a rough guide a blood type to an individual.

General overview The present research study has had, as a starting point, the necessity of solving the complex problems resulting from the practical forensic daily work, having the purpose of combating or sustaining certain statements made by subjects, regarding their own traces left within the perimeters representing criminal areas. The study was carried out during two years and its main objectives were the following ones: Establishing and quantifying the evolution of fingerprints aging process, from the point of view of: - The ridge thickness; - The distance between ridges/ the valleys; - The number of dactiloscopical macroscopic elements ; - The number of pores; - The number of epithelial cells; - Quantifying the total DNA; Quantifying the evolution of the temporal degradation of fingerprints coming from various persons, specific for the four human blood groups. Selecting the necessary criteria in order to establish the chronology of a fingerprint. Establishing a calibration curve, on the basis of the criteria modifications in time, in order to identify the fingerprint age and the blood group. Materials and Methods The descriptive method, the method of laboratory morphological analyses, the method of physical chemical determination, and the method of reporting the degrading biochemical processes have been the methodological research basis for the investigated material. The groups of subjects, used for the study, were students from Campina Police School, being homogenous from a social and educational point of view and representative for the investigated characteristics. The research study was extended and

repeated every 180 days, covering a period of time of over 2 years and using over 800 fingerprints. The fingerprints were lifted on sterile glass. Lucia Forensic application was used for examining each fingerprint, allowing us to take over the images taken by the microscope video camera or by a digital photo camera, to process them at the same scale, to automatically make the necessary measures, to mark the specific elements, and to locate and orient two images and compare them. In order to carry out the microscopic examination of fingerprints, a microscope with a 20x objective, having artificial lighting (white color filter, 70W), was used. The microscopic preparation was done with Nuclear Fast Red (5g of Aluminiumsulphate), 0,1g Nuclear Fast Red, 100ml distilled water. Quantifying the human DNA existing in fingerprints was carried out by using the 7500 Fast Real-Time PCR Systems and the following reactions: TaqMan Fast Univ PCR Master Mix, Human DNAQuant Assay, positive control, de-ionized water. POLILIGHT instrument was used for noticing fingerprints, as well as a lighting source with a variable wave length between 420 460 nanometers, at an incidence angle of 45%. In order to establish the age determination criteria, the fingerprints were examined by using the microscope Nikon SMZ 800, having a Plan 1x-X-1,5x objective and a lighting source Intralux 6000-1 unde incidental light. The comparative results were centralized, being the basis of expressing the morphological differences Figure 1.
70 j 58 g 62 59 60 40 61 h 41 26 42 43 d 28 44 27 12 a 25 6 7 8 5 9 10 11 3 17 13 14 15 1 2 16 45 47 46 63 4 18 29 64 39 37 38 c 23 24 22 21 20 19 30 48 51 31 50 49 57 i 56 55 36 35 34 b 33 32 52 f e 65 54 53 66 69 68 67 71

68 67

51

44

Figure 1. Comparative images of the same fingerprint, immediately after it was created and then after 180 days.

Determining the action of the environmental and intrinsic factors on the degradation in time of the latent

biological material existing in fingerprints was carried out indirectly by studying the degradation stage and/or the epithelial cells disappearance. These determinations were carried out by examining the cytological preparations, directly on fingerprint lifting material, colored with Nuclear Fast Red (NFR). The estimative number of incorporated cells in each fingerprint was determined by mathematical calculus, the values being obtained through the arithmetical rates of three observations, from random areas, where cells were counted. The constant element was the 20x objective surface of the Leica DMLS2 microscope which delimited the surface area where the cells were counted. [1-3] The biological material, transferred on sterile cotton and washed, was concentrated by centrifugation. The DNA was extracted by using Chelex 5% solution, in the presence of Proteinase K, at 56 degrees Celsius. After lizing the epithelial cells to pH 11, the DNA macromolecules were purified by centrifugation and then by boiling in the presence of the ion changing resin. The DNA extract was put under the process of an advanced purification and concentration, by using Microcon YM100 filters. The obtained deposit was suspended in 40 l buffer solution Tris EDTA, pH 8,0. [4] The DNA extract preparation with the view to be quantified was carried out by mixing each sample with 5l TaqMan Fast Univ PCR Master Mix, 1 l Human DNA Quant Assay and 4l of extract. The samples, being prepared that way, were put into the quantification system Fast Real Time PCR 7500 (Applied Biosystems, USA), including the positive and negative control and the standard calibration solutions. The data analysis was performed by using the Sequence Detection Software, version 1.3.1., 7500 Fast System SDS. The result analysis and interpretation was carried out by using the device standard settings, including the software. The threshold was maintained at a constant value of 0.08730054, for all analyses, and the baseline between 3 -15 cycles. [5] The extracted DNA amplification was carried out by using the Identifilter kit (Applied Biosystems, USA) with a PE 9700 thermocycler, at 34 cycles. The reaction mixture, for each sample, consisted in 5 l PCR Reaction Mix, 2,5 l Primer set, 0,25l AmplitaqGold DNA polymerase and 4,75 DNA extract. The capillary electrophoresis was carried out by using the ABI Prism 3100 (ABI PRISM 3100 Genetic Analyzer, a multi-color fluorescence-based DNA analysis system with 16 capillaries operating in parallel). [6 - 9] Results and Discussions The ridge thickness and the valley width were measured by using Lucia Forensic application, every 15 days, during a period of time of 180 days. The following aspects were noticed, on the basis of the microscopic study of the shape and location of pores:

The pores, under observation, after one day from being created, have a density higher than 10/ centimeter of ridge, the pores being located at a short distance fro each other; The position of pores can be both in the center of the papillary ridge and at the ridge periphery. The ones being at the periphery of the papillary ridge can be closed or open, while the ones from within the ridge can only be closed and well-defined; After 5 days, the pores start modifying their shape, especially the marginal ones which could become open pores. Because of the papillary ridges degradation, some pores disappear; The distance between pores gets shorter, and the near ones unite, forming a chain. The ridge thickness after 1 day: The papillary fingerprints, being kept in conditions of an indoor environment: the ridge thickness varies between 0,30mm and 0,34mm; The papillary fingerprints, being kept in conditions of an outdoor environment: the thickness varies between 0,28mm and 0,32mm. The ridge thickness after 180 days: diminishes in time, varying between 0,24,, and 0,28mm for the indoor ones and between 0,22mm and 0,26mm, for the outdoor ones.

The width of initial valleys: For the latent papillary fingerprints left indoors, the valleys vary between 0,24mm and 0,28mm; For the latent papillary fingerprints left outdoors, the valleys vary between 0,26mm and 0,28mm. The valley width after 180 days: As for the papillary fingerprints left indoors, the valleys width gets larger, varying between 0,28mm and 0,30mm; As for the papillary fingerprints left outdoors, the width vary between 0,26mm and 0,28mm. The analysis of the number of macroscopic dactiloscopical elements, during a period of time of 150 days, is shown in the chart below: Examination Period (days)
1 15 30 45 60 75 90 105 120 135 150

No. of Macroscopic Elements

41 41 33 27 25 23 19 18 15 13 12

The biochemical degradation rate, shown by the degradation/ disappearance of the latent epithelial cells from papillary fingerprints, after 180 days, is of

93.43% for blood group O, and of 99% for blood group B.

Blood Group Percentage of degraded/ disappeared cells O
93.43%

A
95.9%

B
99.85%

AB
97.92%

An example of a macroscopic and microscopic degradation of a papillary fingerprint is shown by the figures 2,3 and 4 from below:

Figure 2. Macro and microscopic images of a fingerprint, right after it was created.

Figure 3. Macro and microscopic images of a fingerprint, after 90 days.

Figure 4. Macro and microscopic images of a fingerprint, after 180 days.

The minimization, in percents, of the quantity of DNA, expressed in pg/l, obtained as a result of the quantification, is shown in the chart below, and the graphic representation is shown by Figure 5.
O A 292.12 88.55 33.6 B 284.58 83.56 28.02 AB 287.55 85.12 30.08

Initially After 90 days After 180 days

286.32 88.6 36.4

A slightly increased stability can be noticed at the epithelial cells that belong to blood group O, being followed by A, AB and B, in the end. The study of the degradation in time of the fingerprint incorporated epithelial cells, depending on environmental conditions, material support and persons blood group, led to the following statements: The degradation rate of the biochemical cell constituents existing in fingerprints depends on the exposure to environmental factors; No major differences can be identified between 8 groups of fingerprints kept in the same conditions, that belong to persons having the same blood group; There is a direct cause-effect relation between the variables under study, for the group of persons under study, having the fingerprints being kept in the same conditions and temporal differences in biochemical degradation between different blood groups; By comparing the biochemical degradation phenomenon of the papillary fingerprints that belong to persons, for all 4 blood groups, the conclusion that one may draw is that the epithelial cells have a pre-disposition for biological degradation, depending on the persons blood group, if the environmental factors are the same. The observations, the determinations and the dosages lead to the following classification of the biological material resistance, incorporated in fingerprints, depending on the blood groups: O A AB B; The variation in time of the aging process of fingerprints that belong to persons with secretory or non-secretory character was not quantified. Since the observations were made on groups of fingerprints taken from groups of persons, and, in order to express the results the values of the arithmetical rates of the determinations were used, this aspect was not of an influence on the final results; The fingerprint degradation rate, from the point of view of the ridges thickness, the valleys width, the number of macroscopic dactiloscopic elements, the number of pores, the number of epithelial cells, and the total DNA quantification, is shown in Figure 6.

Figure 5. Graphic representation of the biological material degradation, as a result of DNA Quantification, remaining from fingerprints.

Figure 6. Graphic representation of the 6 types of degradation taking place in a fingerprint.

Conclusions The topic approached in this research study is of high interest and it has been chosen because of the necessity of knowing the behavior in time of the papillary fingerprints, with the view to establish their chronology. The fingerprint age determination is a source of information regarding the study of morphological, physical, chemical and biochemical transformations and it provides important material for the relational interpretative terms between the traces existing at the crime scene, the temporal space and the group of individuals. Gathering the morphological determinations, the physical chemical ones, and the determination of biochemical degradation of fingerprint traces, belonging to persons, for all 4 blood groups (O, A, AB and B), the result is that the biological material and, implicitly, the fingerprints of persons having different blood groups, degrades differently in time. In time, the fingerprints of persons having blood group BIII and ABIV are sensibly more exposed to the aging process comparing to persons having blood group OI and AII. If the environmental factors are identical, the fingerprint decreasing degradation order, depending on blood groups, is B, AB, A and O. The research results have led to certain relational graphics showing the time elapsed between the moment of creation and the morphological, physical, chemical and biochemical elements, which are to be used for estimating the age of this type of traces, used in forensic techniques. The new elements shown by this research study are related to the location in time of the papillary fingerprints by interpreting the temporal degradation stages and to the manner in which the blood group may influence the fingerprint aging process. Therefore, the fingerprint age can be estimated and a certain blood group could be identified. The study results are important for the information gathering, being useful in the investigation process, in creating a smaller group of suspects by establishing a certain blood group, and not for persons identification.

The research results have been used for setting up certain relational graphics showing the time elapsed between the moment of creation and the morphological, physical, chemical and biochemical elements, which are to be used for estimating the age of these types of fingerprints, in forensic techniques.

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