Use of RGB values in the Periodic Acid-Schiff color test to determine the presence of vaginal fluid.

This study demonstrates how RGB color values from microscopic smears stained with the Periodic Acid-Schiff reagent under standardized microscopy conditions can be used to indicate the presence of vaginal secretions. Based on data obtained in the study, a numeric threshold determined from the sum of separate values for red, blue and green was determined to differentiate vaginal-based samples with other body fluids. Using this threshold, 55 of 57 vaginal-based samples tested positive for the presence of vaginal secretion. Conversely, 27 of 29 smears prepared from other body fluids yielded negative results. However, when graphing RGB sum values against a calculated RGB integer no overlap in data was obtained between all vaginal-based samples and other body fluid samples, clearly differentiating them. One-way ANOVA testing with a 95% confidence interval indicated that vaginal samples from different age groups showed no difference in RGB sum values. Similarly, the location that vaginal swabs were collected (from the outside of a condom or a vaginal swab) also showed no statistical difference using one-way ANOVA at 95% confidence. Furthermore, refrigerated test swabs aged up to 15 months showed no demonstrable differences. Pair-wise t-testing using RGB sum values, however, did show significant differences between vaginal samples and all other body fluids tested. Finally, the method successfully differentiated between pre-and post-coital penile swabs and finger swabs taken before and after digital vaginal penetration in anecdotal comparisons using the method.

Identifying PCP and four PCP analogs using the gold chloride microcrystalline test followed by raman microspectroscopy and chemometrics.

Identifying drug analogs can be a vexing problem for forensic scientists particularly in today's evolving drug market. This study proposes a method that utilizes microcrystalline tests, Raman microspectroscopy, and chemometrics to help solve this problem. In the present case, the method described was used to clearly differentiate and identify phencyclidine (PCP) and four of its analogs, namely tenocyclidine (TCP), rolicyclidine (PCPy), 3-methoxy phencyclidine (3-MeO PCP), and 4-methoxy phencyclidine (4-MeO PCP). Microcrystals were grown from each drug with gold chloride and examined using polarized light microscopy. Morphological and optical properties such as shape, habit, time of growth, color, retardation colors, type/angle of extinction, and sign of elongation were observed and documented to characterize each microcrystal. Analysis with a Raman microscope was able to provide structural information on the microcrystals. Objective analysis of the microcrystal spectra was done by employing chemometrics. A training set of Raman shifts was compiled and transformed with principal component analysis (PCA) followed by linear discriminant analysis (LDA). The training set was validated by leave-one-out cross validation (LOOCV) and subsequently ran against a separately-compiled test set. Mahalanobis distances between test samples and the clusters of training samples in LDA space were calculated to empirically demonstrate the applicability of this drug analysis technique. From the results of this study, a drug analysis protocol was developed for analysts to use for the identification of PCP, TCP, PCPy, 3-MeO PCP, and 4-MeO PCP and to serve as a model for drug analogs in general.

Development of a multiplex, PCR-based genotyping assay for African and Asian elephants for forensic purposes.

Wildlife crimes and the threats they present to elephant populations raise the need to develop and implement DNA-based methodology as an aid for wildlife forensic investigations and conservation efforts. This study describes the development of a tetra-nucleotide repeat STR multiplex, genotyping assay that will identify Asian elephant (Elephas maximus) and African elephant (Loxodonta africana) DNA. The assay targets six tetra-nucleotide STRs and two sex-typing markers simultaneously in both genera of elephants, a first for elephant genotyping assays. The developed assay has potential application in wildlife investigations to associate a biological sample to a particular individual elephant and additionally in conservation science for population management.

Detection of prostate specific antigen and salivary amylase in vaginal swabs using SERATEC® immunochromatographic assays.

Immunochromatographic assays are used by crime laboratories to conduct simple and quick analyses of bodily fluids. These streamlined tests are ideal for decreasing the sexual assault kit backlog in the United States. A large-scale analysis of the frequency of positive results of amylase and prostate specific antigen (PSA) endogenously found in the vaginal cavity was conducted using the SERATEC PSA Semiquant and Amylase tests. Vaginal swabs were self-collected by participants after 7-10 days of no oral contact or male ejaculation. In this study of 50 participants, 98% were negative for PSA and 92% were negative for amylase. Positive results were confirmed to contain no exogenous DNA by male-specific quantitation, short tandem repeat (STR) typing, and Y-STR typing. These results can be used by crime laboratories to help guide interpretation of immunochromatographic test results from vaginal swabs and aid in decision-making in downstream DNA testing.

Mandatory certification of forensic science practitioners in the United States: A supportive perspective.

The article presented is supportive of mandatory certification of forensic scientists and believes that such a mandate can help establish a threshold for competency in the profession, provide a universal standard for ethical professional conduct, and enhance the credibility of forensic science in users of the profession and the general societal public. After examining the history of professional certification in the United States across a spectrum of professions including forensic science, results of surveys sent to forensic science practitioners to gauge their views on mandatory certification is discussed. Seventy-three surveys were received with most surveyors being in support of mandatory certification related to their discipline. Reasons why many practitioners have chosen not to engage in voluntary certification were also provided on the surveys and discussed. Finally, the article discusses possible ways of implementing mandatory certification and concludes that the mechanisms to achieve this goal may already be in place.

High resolution melt curve analysis based on methylation status for human semen identification.

A high resolution melt curve assay to differentiate semen from blood, saliva, urine, and vaginal fluid based on methylation status at the Dapper Isoform 1 (DACT1) gene was developed. Stains made from blood, saliva, urine, semen, and vaginal fluid were obtained from volunteers and DNA was isolated using either organic extraction (saliva, urine, and vaginal fluid) or Chelex® 100 extraction (blood and semen). Extracts were then subjected to bisulfite modification in order to convert unmethylated cytosines to uracil, consequently creating sequences whose amplicons have melt curves that vary depending on their initial methylation status. When primers designed to amplify the promoter region of the DACT1 gene were used, DNA from semen samples was distinguishable from other fluids by a having a statistically significant lower melting temperature. The assay was found to be sperm-significant since semen from a vasectomized man produced a melting temperature similar to the non-semen body fluids. Blood and semen stains stored up to 5 months and tested at various intervals showed little variation in melt temperature indicating the methylation status was stable during the course of the study. The assay is a more viable method for forensic science practice than most molecular-based methods for body fluid stain identification since it is time efficient and utilizes instrumentation common to forensic biology laboratories. In addition, the assay is advantageous over traditional presumptive chemical methods for body fluid identification since results are confirmatory and the assay offers the possibility of multiplexing which may test for multiple body fluids simultaneously.

Detection of male DNA in the vaginal cavity after digital penetration using Y-chromosome short tandem repeats.

In this study, useful genetic information from male donors was obtained on vaginal swabs taken from female volunteers after male digital vaginal penetration in a time frame relevant to a sexual assault investigation. Vaginal swabs were collected from eight volunteers at intervals of 1, 6, 12, 24, and 72 hours after digital vaginal penetration. DNA was extracted from collected swabs and subsequently genotyped using a commercially available Y-chromosome short tandem repeats (Y-STR) multiplex kit. Fifty-eight vaginal swabs were collected and analyzed in the study. Composite Y-STR profiles from all combined volunteers showed that 85% of all possible alleles were detected at the 1-hour interval, 77% of all possible alleles were detected at the 6-hour interval, 73% of all possible alleles were detected at the 12-hour interval, 66% of all possible alleles were detected at the 24-hour time interval, and 71% of all possible alleles were detected at 72 hours after digital vaginal penetration. Results indicate that a viable possibility exists that probative Y-STR profiles, useful for investigative purposes, can be obtained from vaginal swabs taken from subjects exposed to digital penetration at time intervals up to 72 hours postpenetration.

Comparative forensic soil analysis of New Jersey state parks using a combination of simple techniques with multivariate statistics.

This study has shown that the combination of simple techniques with the use of multivariate statistics offers the potential for the comparative analysis of soil samples. Five samples were obtained from each of twelve state parks across New Jersey in both the summer and fall seasons. Each sample was examined using particle-size distribution, pH analysis in both water and 1 M CaCl2 , and a loss on ignition technique. Data from each of the techniques were combined, and principal component analysis (PCA) and canonical discriminant analysis (CDA) were used for multivariate data transformation. Samples from different locations could be visually differentiated from one another using these multivariate plots. Hold-one-out cross-validation analysis showed error rates as low as 3.33%. Ten blind study samples were analyzed resulting in no misclassifications using Mahalanobis distance calculations and visual examinations of multivariate plots. Seasonal variation was minimal between corresponding samples, suggesting potential success in forensic applications.

The utility of Y-STR profiling in four-, six- and eight-day postcoital vaginal swabs.

Y-STR profiles using the Promega PowerPlex(®) Y system were attempted on multiple vaginal swabs collected at four, six and eight days after intercourse from female partners of 11 couples. At four days postcoitus, full composite profiles (where swabs yielded confirmed alleles at all 11 loci) were obtained for five of the 11 couples and 78% of all possible alleles summed for all couples were confirmed (able to be duplicated in different swabs). Results for composite profiles for all couples taken at six days after intercourse showed that 53% of all alleles summed for all couples were confirmed. Only one couple yielded a full composite profile at six days after intercourse. Composite profiles from swabs taken at eight days after intercourse for all couples confirmed only 44% of all possible alleles summed for all couples. At eight days postcoitus, no couple yielded a full composite profile and the largest number of confirmed alleles for any couple was eight. However, one of 44 individual swabs taken from all couples combined at eight days postcoitus yielded a 10-locus profile. Composite partial profiles from the eight-day postcoital set with confirmed results at a minimum of five loci (8 of 11 couples) yielded haplotype frequencies from 0.000323 to 0.125862 using the Y chromosome haplotype reference database, suggesting that meaningful Y-STR information can still be obtained at much extended postcoital intervals.

The utility of polyester and cotton as swabbing substrates for the removal of cellular material from surfaces.

Various types of cotton and polyester fabrics were tested to ascertain the optimal physical and chemical characteristics of fabrics needed for the removal of cellular material from surfaces. DNA quantitation values obtained on dried saliva stains showed no difference between cotton and polyester across all constructions and solvent conditions. Fabrics used dry and with water yielded higher quantitation values than those used with isopropanol. Quantitation values were also higher for wovens and nonwovens than knits across all solvent conditions. Low thread count fabrics used with water yielded higher quantitation values, but no correlation between thread count and quantitation values was observed with dry fabrics. A low thread count woven fabric, however, outperformed other tested fabrics when swabbing object surfaces in a highly used room. Full DNA profiles from fingerprints on glass surfaces were obtained with low thread count woven and nonwoven fabrics but not with the knit fabric tested.

Differentiation of twenty-one glitter lip-glosses by pyrolysis gas chromatography/mass spectroscopy*.

Differentiation of 21 glitter lip-glosses from seven manufacturers was attempted by pyrolysis gas chromatography/mass spectroscopy. Samples were pyrolyzed on a ribbon probe at 800 degrees C for 20 sec and analyzed with an Agilent 6890N Network GC System and Agilent 5973 Network Mass Selective Detector with MSD Productivity ChemStation Data Analysis software. The total ion chromatograms obtained were examined and differences in the presence or absence of certain chromatographic peaks corresponding to certain pyrolysis products (e.g., styrene, cyclohexane) noted. In cases where the total ion chromatograms between lip-glosses were similar, select ion profiling was performed. Of the 21 lip-glosses, 15 were differentiated by either the total ion chromatograms alone or through select ion profiling. Considering that lip-glosses are typically worn by young women (who are disproportionately victims of sexual assault), this study offers the potential of being able to provide investigative leads in sexual assault investigations with evidentiary samples of this kind.

Measurement of substance P and met-enkephalin in the serum of violent death victims.

Very often the allocation of putative damages for wrongful death and the determination of aggravating factors in the sentencing of an individual convicted of homicide by a jury is based on a subjective determination of the amount of pain suffered by the victim. This study was designed to determine whether the quantitative determination of peptides involved in nociception and inflammation offer the potential to provide an objective basis for an assessment of pain prior to death. Two peptides. substance P and met-enkephalin, were quantitated using radioimmunoassay (RIA) in the serum of 131 autopsy subjects. Cases were selected that presented decedents who underwent a violent death resulting in extensive trauma to tissue. Only decedents with no known prior clinical manifestation and no indication of prior drug use were selected. Of 131 cases selected, 59 died from blunt trauma deaths, 47 from gunshot deaths, and 25 from stabbing deaths. Cases were selected without regard to whether the death was accidental, or by homicide or suicide. Values from cases having similar incident-death time intervals were pooled and then compared. Results show that an observable pattern exists between the concentrations of substance P and met-enkephalin and the incident-death time interval. Data showed that the concentrations of substance P and met-enkephalin vary with the incident-death time interval. The amount of serum substance P initially increases with increasing incident-death time interval but begins to decrease at longer incident-death time intervals. In contrast, the serum concentration of met-enkephalin continues to show increased concentration as the incident-death time intervals become greater. The Kruskal-Wallis test was performed to determine the level of significance of the variation in both peptide concentrations within four consecutive time intervals. Variation in substance P concentration was statistically significant in all comparisons performed with 0.01 being the lowest level of significance of any four consecutive groups tested. Conversely, intervals encompassing incident-death time intervals of 1-2 hours to 5-10 days did not demonstrate significant variation in met-enkephalin concentration. However, groups with smaller and larger time intervals than the nonsignificant groups did show statistical variation. Although owing to a number of variables, a direct correlation between peptide concentrations and the level of pain may not be possible, the results of the study indicate that a presumption of antemortem pain may be possible with future study.

An ELISA method for the identification of salivary amylase.

An ELISA method for the detection of salivary amylase in dried stains using a monoclonal anti-human salivary amylase antibody was developed. Studies demonstrated the assay to be sensitive down to 0.0002 Sigma units and showed a linear response between absorbance and salivary amylase activity between 0.002 and 0.2 units. The assay showed no cross reactivity with either commercially purchased human pancreatic or bacterial amylase. Sample studies utilizing swabs from several human body fluids showed that 100% of all saliva containing swabs (sixteen of sixteen) and 13% of non-saliva human body fluid swabs (eight of sixty-three) showed a net absorbance with the method. Of these eight non-saliva swabs yielding a net absorbance, none exceeded a salivary amylase activity of 0.003 units. In contrast, only three of the sixteen saliva-containing swabs (swabs produced from saliva diluted 1:5, 1:6, and 1:10, respectively) showed an activity below 0.2 units. Of these swabs, the 1:100 dilution showed the lowest activity (0.048). This value is still more than ten times that of the non-saliva containing swab with the highest activity.