Robert C. Shaler

Analysis of restriction fragment length polymorphisms in deoxyribonucleic acid (DNA) recovered from dried bloodstains.

Deoxyribonucleic acid (DNA) was recovered from dried bloodstains aged up to three years and shown to be of high molecular weight. DNA was digested with restriction endonucleases and fractionated by agarose gel electrophoresis. Following transfer to a filter, DNA was hybridized with two different radioactively labeled recombinant probes which recognize highly polymorphic regions in human DNA. The autoradiographic pattern observed was not altered by sample age, and the size of the alleles was consistent with those observed in the general population. Therefore, DNA of high molecular weight prepared from dried blood samples can be used for identification.

Evaluation of Deoxyribonucleic Acid (DNA) Isolated from Human Bloodstains Exposed to Ultraviolet Light, Heat, Humidity, and Soil Contamination

This study was designed to analyze the effects of common environmental insults on the ability to obtain deoxyribonucleic acid (DNA) restriction fragment-length polymorphisms (RFLP) patterns from laboratory prepared specimens. The environmental conditions studied include the exposure of dried bloodstains to varying amounts of relative humidity (0, 33, 67, and 98%), heat (37 degrees C), and ultraviolet light for periods of up to five days. In addition, the effect of drying over a four-day period in whole blood collected with and without ethylenediaminetetraacetate (EDTA) was examined. The results of the study showed that, under the conditions studied, the integrity of DNA is not altered such that false RFLP patterns are obtained. The only effect observed was that the overall RFLP pattern becomes weaker, but individual RFLP fragments are neither created nor destroyed.

Validation and casework application of a Y chromosome specific STR multiplex

A series of validation experiments was performed for a Y chromosome specific STR multiplex system following the suggestions made by the Technical Working Group DNA Analysis Methods (TWGDAM). The multiplex PCR products were detected on Perkin-Elmer 373 and 377 automated sequencers using two labeling colors. No problems regarding the stability, robustness and sensitivity of the Y STR multiplex were observed. Mixture studies revealed a cut off rate similar to autosomal STRs for mixtures of male DNAs and no interference of any female admixture. The comparison of the Y STR results to the autosomal typing results for 56 nonprobative semen stains and swabs, showed a slightly higher success rate in detecting the semen donors alleles for the Y STR multiplex. Two examples are shown to illustrate the usefulness of Y STR typing for DNA mixtures. In one case the Y STR results confirmed an isolated exclusion; in the other case, the interpretation of a mixture was clarified since the Y STR results proved the presence of DNA from at least two semen donors. Y STR typing is a valuable addition to the forensic DNA testing panel.

Bioinformatics and human identification in mass fatality incidents : The world trade center disaster

Abstract:  Victim identification initiatives undertaken in the wake of Mass Fatality Incidents (MFIs) where high‐body fragmentation has been sustained are often dependent on DNA typing technologies to complete their mandate. The success of these endeavors is linked to the choice of DNA typing methods and the bioinformatic tools required to make the necessary associations. Several bioinformatic tools were developed to assist with the identification of the victims of the World Trade Center attacks, one of the most complex incidents to date. This report describes one of these tools, the Mass Disaster Kinship Analysis Program (MDKAP), a pair‐wise comparison software designed to handle large numbers of complete or partial Short Tandem Repeats (STR) genotypes, and infer identity of, or biological relationships between tested samples. The software performs all functions required to take full advantage of the information content of processed genotypic data sets from large‐scale MFIs, including the collapse of victims data sets, remains re‐association, virtual genotype generation through gap‐filling, parentage trio searching, and a consistency check of reported/inferred biological relationships within families. Although very few WTC victims were genetically related, the software can detect parentage trios from within a victim’s genotype data set through a nontriangulated approach that screens all possible parentage trios. All software‐inferred relationships from WTC data were confirmed by independent statistical analysis. With a 13 STR loci complement, a fortuitous parentage trio (FPT) involving nonrelated individuals was detected. Additional STR loci would be required to reduce the risk of an FPT going undetected in large‐scale MFIs involving related individuals among the victims. Kinship analysis has proven successful in this incident but its continued success in larger scale MFIs is contingent on the use of a sufficient number of STR loci to reduce the risk of undetected FPTs, the use of mtDNA and Y‐STRs to confirm parentage and of bioinformatics that can support large‐scale comparative genotyping schemes capable of detecting parentage trios from within a group of related victims.

The Effects of Environment and Substrata on Deoxyribonucleic Acid (DNA): The Use of Casework Samples from New York City

This study was designed to analyze the effects of the environment and substrata on the quality of deoxyribonucleic acid (DNA) isolated from evidentiary specimens. The quality of DNA isolated from actual casework specimens was determined by measuring its size by agarose gel electrophoresis. The information obtained could be used to predict the suitability of the DNA in the samples for restriction fragment length polymorphism (RFLP) analysis. The evidentiary specimens chosen for DNA were classified according to substrate (scrapings, plastic bags, synthetics, denim, and carpet) and according to a subjective evaluation of the condition of the stain (soiled, damp, or putrefied) and to its size (small or large). The results show that DNA of sufficient quality and high molecular weight (HMW) can be reliably isolated from bloodstains deposited on evidentiary items which have an unknown environmental history and which have dried onto a variety of substrata. Subsequent RFLP analysis of a selected number of these samples verified that the DNA was suitable for this type of analysis.

The Evaluation of Fatty Acid Ratios in Latent Fingermarks by Gas Chromatography/Mass Spectrometry (GC/MS) Analysis†

Despite advances in DNA, fingermarks remain one the best forms of evidence available. While fingermarks are routinely analyzed in terms of their patterns, it may be possible to obtain additional information in terms of their chemical composition. If successful, a chemical analysis of the constituents of a fingermark may give scientists additional information that may help in the identification of a person. The results presented herein describe the initial investigation into the analytical determination of some of these compounds, specifically the fatty acids. This study was specifically aimed at identifying possible fatty acids, which could aid in profiling or perhaps uniquely identifying an individual. Preliminary data obtained in this study suggests that this may in fact be possible, though additional research is certainly necessary. Utilizing gas chromatography–mass spectrometry, significant differences in the ratios of several fatty acid methyl esters were found when comparing individuals of varying race and gender. In addition, large intervariability and intravariability was discovered for some compounds, suggesting the possibility of being able to individualize based on chemical profile. Follow‐up investigations will continue to determine whether this continues to be the case as greater numbers of individuals are sampled and more extensive control and information on the subjects is obtained.

A Physical Method for Separating Spermatozoa from Epithelial Cells in Sexual Assault Evidence

The analysis of genetic markers for the purpose of individualization of semen specimens is extremely important in cases of sexual abuse and assault. The serological analysis of sexual assault evidence can sometimes be complicated because stains are often composed of a mixture of spermatozoa, vaginal epithelial cells and white and red blood cells. A filtration method has been developed to cleanly separate spermatozoa from epithelial cells based upon differences in size and shape. Nylon mesh filters of the appropriate pore size can be used to separate the smaller oval shaped spermatozoal cells from the larger and flatter epithelial cells. The former pass freely through the membrane while the latter are retained on the filter. In this study, cell separation was demonstrated by (a) microscopic observation of stained cells, (b) amplified fragment length polymorphism analysis of DNA obtained from separated cells. The results of these analyses indicate that: (1) Approximately 70% of spermatozoa in the mixed cell sample will penetrate the 10 microns pore size filter, (2) Only about 1-2% of intact epithelial cells will do so, and (3) A small number of nuclei from spontaneously lysed epithelial cells will cross the filter. Experimental results using mixtures of spermatozoa and vaginal epithelial cells prepared in different ratios support the conclusion that the filtration process is an efficient and reliable method to separate spermatozoa from epithelial cells in casework specimens for subsequent DNA analysis.

An Assessment of DNA Contamination Risks in New York City Medical Examiner Facilities

DNA evidence holds an important position in criminal investigations and proceedings. The polymerase chain reaction (PCR) is often utilized to amplify polymorphic regions of DNA which are subsequently typed to produce distinct genotypes. The sensitivity of PCR-based techniques provides a major advantage over other DNA or conventional serological typing systems. Samples containing quantities of DNA in the picogram range are often typed. However, the unprecedented sensitivity of PCR is often cited as a criticism. One concern is that the interpretation of PCR typing can be affected by DNA contaminants from foreign sources. In this report, the level of DNA contamination in New York City Medical Examiner facilities and its potential affects on HLA-DQA1 typing were assessed. Two related studies conducted over a five week period measured and typed HLA-DQA1 from accumulated DNA on autopsy room and Forensic DNA Laboratory structures. The potential for DNA contamination from airborne sources was also evaluated in the autopsy suites. This study demonstrated the presence of small amounts of DNA on structural surfaces, but little evidence of airborne DNA contamination.

Solid-State Acquisition of Fingermark Topology using Dense Columnar Thin Films

Abstract:  Various vacuum techniques are employed to develop fingermarks on evidentiary items. In this work, a vacuum was used to deposit columnar thin films (CTFs) on untreated, cyanoacrylate‐fumed or dusted fingermarks on a limited selection of nonporous surfaces (microscope glass slides and evidence tape). CTF deposition was not attempted on fingermarks deposited on porous surfaces. The fingermarks were placed in a vacuum chamber with the fingermark side facing an evaporating source boat containing either chalcogenide glass or MgF2. Thermal evaporation of chalcogenide glass or MgF2 under a 1 μTorr vacuum for 30 min formed dense CTFs on fingermark ridges, capturing the topographical features. The results show that it is possible to capture fingermark topology using CTFs on selected untreated, vacuumed cyanoacrylate‐fumed or black powder–dusted nonporous surfaces. Additionally, the results suggested this might be a mechanism to help elucidate the sequence of deposition.

Columnar-thin-film acquisition of fingerprint topology

Fingerprint visualization obtained from physical evidence taken from crime scenes for subsequent comparison typically requires the use of physical and chemical techniques. One physical technique to visualize or develop sebaceous fingerprints on various surfaces employs the deposition of metals such as gold and zinc thereon. We have developed a different vacuum technology: the conformal-evaporated-film-by-rotation technique to deposit dense columnar thin films (CTFs) on latent fingerprints on different types of surfaces. Sample fingerprints, acting as nonplanar substrates, deposited on different surfaces were placed in a vacuum chamber with the fingerprint side facing a boat containing an evaporant material such as chalcogenide glass. Thermal evaporation of the solid material led to the formation of a dense CTF on the fingerprint, thereby capturing the topographical texture with high resolution. Our results show that it is possible to acquire the topology of latent fingerprints on nonporous surfaces. Additionally, deposition of CTFs on overlapping fingerprints suggested ours may be a technique for elucidating the sequence of deposition of the fingerprints at the scene.