105th SEMI-ANNUAL SEMINAR (Spring 2005)
CALIFORNIA ASSOCIATION OF CRIMINALISTS
May 9-13, 2005
Oakland Airport Hilton, California

A NOVEL METHOD OF SCREENING FOR HUMAN MALE DNA IN FORENSIC SAMPLES
Jaiprakash G. Shewale, Elaine Schneida(1), Jerilyn A. Walker(2), Mark A. Batzer(2) and Sudhir K. Sinha*(1)

(1)ReliaGene Technologies, Inc.; (2)Department of Biological Sciences, Biological Computation and Visualization Center, Louisiana State University

Screening of sexual assault evidence samples for the presence of sperm or semen is generally the first step in forensic DNA analysis. For this study, a total of 887 sexual assault cases were screened by using p30, AP and microscopy methods. The failure rate for obtaining a male profile from p30 positive, microscopic positive and p30 and microscopic positive cases was 58, 45 and 17%, respectively. Thus, currently used methods of screening provide false positive results. In addition, these screening tools are not targeted to detect the presence of male DNA. As a result many evidence samples containing tissue or body fluids other than semen cannot be screened using these methods. Thus, there is a need for a sensitive, reliable and high throughput screening method for the detection of male DNA in forensic samples. We have developed a novel screening system, Y-Screen, for the detection of male DNA in forensic samples. The method is based on PCR amplification of Alu insertions fixed within the Y chromosome. The Alu family of interspersed repeats is the most successful of the mobile genetic elements within primate genomes, having amplified to a copy number of greater than 1,000,000 per haploid genome. Alu repeats are unique nuclear markers that are ideally suited for human identity testing. Individual Alu repeats are approximately 300 bp in length and are thought to be derived from the 7SL RNA gene. This approach eliminates false positive results obtained from degraded DNA, a drawback of currently available DNA quantitation methods based on real-time PCR. Using the YScreen assay, it is possible to detect as little as 5 pg of male DNA. Avian DNA is incorporated as an internal control to monitor the presence of PCR inhibitors in the extract. The protocol is designed to consume less than 10% of the evidence sample. Screening assay can be performed using a 96 well format to facilitate high-throughput screening. The utility of the Y-Screen assay in forensic DNA analysis will be discussed.


MITOCHONDRIAL DNA ANALYSIS OF AGED SIBLINGS AND LONGITUDINAL SAMPLES
Jessica Wijbenga, BSc.*(1,2) and Cassandra D. Calloway, MSc.(1,3)

(1) Roche Molecular Systems, Alameda, California; (2) Utrecht University, Utrecht, the Netherlands; (3) University of California, Berkeley, California

When nuclear DNA analysis fails or is not appropriate, mitochondrial DNA (mtDNA) can be analyzed to obtain forensic information for crime scene cases. High copy number and high degree of sequence variation allow for the analysis of even degraded DNA samples. Also since mtDNA is maternally inherited, it allows for an expanded number of reference samples in missing persons cases. However, heteroplasmy (presence of both wild-type and mutant mtDNA's within a cell) can occur during life and result in differences between maternally related individuals. Heteroplasmy has been reported to increase with age and to occur at certain positions more frequently. Heteroplasmy may be inherited maternally or result from a somatic mutation.

To further characterize heteroplasmy, we typed the hypervariable regions I (HVI) and II (HVII) in lymphoblasts (n=45 samples) collected from aged sib-pairs (SP) and fibroblasts (n=89 samples) from the National Institute of Aging Cell Repository Longitudinal Study (LS). The SP study consisted of 15 families with an individual and at least one sibling. For the LS study, biopsies were collected from individuals two or more times throughout life.

Samples were typed using the Linear Array Mitochondrial DNA HVI/II Region-Sequence Typing Kit from Roche Applied Science. This linear array allowed us to rapidly screen 134DNAsamples from78 individuals. In addition, we screened the samples for heteroplasmy at position 414 in HVII using a SSO-probe. Previous studies showed that this mutation occurs more frequent in older individuals. Since many forensic laboratories use a reverse HVII primer that contains the mutation site (T414G), we investigated this site to determine whether the mutation could influence the PCR yield and thus the typing/ sequencing results. Heteroplasmy was observed in two of the LS individuals for region HVI (at position 16093) and in four of the LS individuals for region HVII (position 146, 152, 189 and 195). Three individuals showed an occurrence of the heteroplasmy at an older age, but this heteroplasmy could not be detected at a younger age. These data suggest that the heteroplasmy is a result of a mutational event rather than inherited. Interestingly, one individual was heteroplasmic at position 146 at age 36 but not at age 51. Heteroplasmy was observed at position 16304 in one individual of the SP study, but was not observed in the sibling's sample. This finding suggests that the mutated mtDNA is not inherited from the mother.

Heteroplasmy was detected at position 414 in eight LS individuals. The presence of mutant mtDNA reached up to 75% of mtDNA. Another individual had a complete transversion from a T to G at the age of 79 but this mutation was not detected at ages 73 and 85. Of note, there was no detectable heteroplasmy of the 414 position in lymphoblasts. In total, there were 19 occurrences of heteroplasmy in 89 fibroblast samples versus one occurrence in 45 lymphoblast samples. This data suggests that the occurrence of heteroplasmy is more prominent in skin cells. Heteroplasmy at position 414 or even the T414G mutation does not affect binding of the HVII reverse primer.

The analysis of mtDNA can be very useful in forensic cases. Important is that criminalists realize that heteroplasmy can have an impact on mtDNA typing or sequencing, especially in cases when older individuals or skin cells are involved.


WRONGFULLY CONVICTED-THE CLUE "SAVING RAY KRONE"
Christopher J. Plourd, J.D.

The goal of this presentation is to demonstrate that innocent people can be convicted of serious crimes because of crime laboratory errors and omissions. This is a serious problem. The educational objective of this presentation is to identify common errors in forensic scientific investigations and suggest strategies for improving objectivity in crime laboratory analysis. The attendee will be able to understand the need for caution in reaching a conclusion. The problem of innocent people being convicted and unjustly imprisoned for crimes they did not commit is a growing national concern which has been receiving public acknowledgment by politicians and is catching the attention of the general public. Advances in DNA identity testing have exonerated a number of innocent people. Some exoneration cases involve crime laboratory errors relating to trace and biological evidence. Ray Krone was the 100th person convicted of and sentenced to death for a capital murder to walk free from prison since the reinstatement of the death penalty in the United States. Ray Krone had maintained his innocence throughout his incarceration. Ray Krone was sentenced to death in 1992 for the brutal murder of Kim Ancona, a Phoenix bar manager. Krone spent three years on Arizona's death row before his death sentence and conviction was overturned. Krone was then retried and convicted a second time and sentenced to life in prison in 1996. Ray Krone, who had been branded as the "snaggletooth killer," was proved innocent of the murder of Kim Ancona by Post Conviction STR DNA testing in 2002. After being cleared by DNA, Ray Krone walked out of an Arizona State Prison a free man after 10 years.

The murdered bar manager, Kim Ancona, had been cleaning the CBS Lounge in Phoenix, Arizona on the evening of December 28, 1991. Her naked body was found in the men's restroom the following morning. She had been stabbed eleven times. An examination of the body revealed that she had been bitten on the left breast. There were unidentified shoe impressions, fingerprints, and hairs. Other evidence indicated she had been sexually assaulted. There was blood at the crime scene and on the victims clothing. The blood was typed as ABO Type O, the same as Ancona, Krone, and some 43% of the population. Forensic DNA technology available at the time of the 1992 prosecution (DQ alpha) did not identify the blood or saliva of the perpetrator. Crime Laboratory errors occurred in 1992 that caused a misinterpretation of the blood and saliva evidence. This same evidence, with use of STR PCR DNA testing, would expose these errors in 2002. Ray Krone was a United States postal letter carrier who had no criminal record and had been honorably discharged from the U.S. Air Force. He knew the victim, as he had socialized with her and had been a customer of the CBS Lounge. There was little evidence that tied Krone to the killing except for evidence of a bitemark on the victim's breast, which an American Board of Forensic Odontology (ABFO), Board Certified Forensic Odontologist said positively, (better than a fingerprint) matched the dentition of Ray Krone. Despite evidence of his innocence presented at both of Krone's trials, the State's circumstantial evidence bolstered by the forensic bitemark evidence convicted Krone. This bitemark evidence was controversial and disputed by other ABFO Board Certified forensic experts.

After Krone's appeals were exhausted following his second conviction Krone sought post-conviction DNA testing. Krone's lawyers asked that the victim's tank top, through which the bitemark may have been inflicted, be examined for saliva DNA analysis. Not only was saliva found, but the results of testing showed that neither Krone nor the victim Ancona could have been the genetic source of the saliva. Comparison of the genetic profile of the saliva donor against the FBI Combined DNA Index System (CODIS) database associated the DNA evidence with a 36-year old inmate of the Florence, Arizona State prison. The inmate was Kenneth Phillips, who had been convicted of child molestation after the date of the Ancona murder. The Krone case is clear proof, again, of the power of DNA. Not only did the DNA test show that Ray Krone was excluded as the perpetrator, it also identified a different individual who was already incarcerated in the penitentiary for an unrelated sex crime. The odds were 1.3 quadrillion to one that Kenneth Phillips was the contributor of the saliva DNA found on Kim Anconas' tank top. After the saliva DNA matched Phillips, his hair was found to be consistent with evidence hairs found on the victim's body. Phillips confessed to being present at the time of the murder of Ancona in a tape recorded interview. Phillips' blood was genetically identified on the inside and outside of the victim's jeans and underwear. Phillips' fingerprints were found in the men's room of the CBS lounge where Kim Ancona's body was found.

The Krone case is another in a growing number of cases where crime laboratory testing has been shown to be erroneous. Lessons can be learned from the Krone case. An independent scientific technical working group of forensic scientists should be formed to objectively study exoneration cases.


DISCLOSURE: A LEGAL OBLIGATION, A PROFESSIONAL REQUIREMENT, AND A SCIENTIFIC RESPONSIBILITY
Peter D. Barnett, Forensic Science Associates

The product of a scientific inquiry is an opinion. The expression of a scientific opinion, most often made in a written communication of some type, must include the scientific facts (data) upon which the opinion is based, it must be capable of review by other knowledgeable scientists, and must explicitly answer a question that is of interest in the inquiry being made. Neither the facts nor the opinion, standing alone, are of much value. It is only in their combination, and their ability to provide an answer to a relevant question, that the work of the scientist provides information on which decisions can be based. The communications of forensic scientists frequently fall short of accepted legal requirements, professional responsibilities, and accepted scientific practice. To the extent that accepted laboratory practices fail to satisfy the legal, professional or scientific requirements for disclosure, we must examine our practices and change them to conform our practices to what is legally required, professionally mandated, and scientifically acceptable.

Requirements for disclosure do not simply mean that a forensic scientist has to provide, when requested, bench notes, proficiency test results, laboratory procedure manuals, and the like. The disclosure requirements mandate that the scientist disclose what was known to him before any work started, what information was obtained, from any source, as the investigation progressed, what questions were being addressed by the work that was being done, the thought process that resulted in the particular work that was undertaken, , and the conclusions and opinions of the scientist. How those opinions will be presented in court, what visual aides may be used, and what exhibits might be presented to the jury must also be part of the disclosure obligation.

Legal requirements for disclosure are set forth in the Federal Rule of Civil Procedure Rule 26, California Penal Code Section 1054 (which codifies Proposition 115 which was enacted by California voters), and various judicial rulings. Professional responsibilities are set forth in the CAC Code of Ethics, the ABC Rules of Professional Conduct, ASTM Standard Practice for Reporting Opinions of Technical Experts (E620-97), and a current ASCLD/LAB proposed accreditation requirement ("Proposal 22"). Scientific practices are set forth in books (e.g., Robert A. Day, How to Write and Publish a Scientific Paper), authors' instructions for various journals, and established practice. The legal, professional and scientific disclosure obligations will be discussed in the context of situations in which one or more of these obligations was not met by the work that was done, the report that was published, or the testimony that was given. Repercussions of the failure to satisfy disclosure obligations will be described.


SOMETHING FROM NOTHING [S&N]: THE DNA ANALYSIS OF HANDGUN SWABS AT CELLMARK IN U.S. VS. WILLIAMS
Edward T. Blake* and Alan Keel, Forensic Science Associates

During the investigation of a shooting in Washington, D.C., a Sig Sauer handgun was recovered. Five swabs were collected from different areas on the handgun by Metropolitan Police crime scene technicians. These swabs were taken from the handgun [1] left hand grip, [2] right hand grip, [3] backstrap, [4] trigger, and [5] finger grip. These handgun swabs and other evidence [baseball cap and vehicle airbag] were submitted to Cellmark for DNA analysis by the U.S. Attorney's Office. This report describes the Cellmark analysis of the five handgun swabs.

The analysis of the handgun swabs at Cellmark proceeded according to the following steps: First all five swabs were bisected and extracted for DNA without microscopic examination for cell debris. A human DNA slot blot assay failed to detect human DNA. Next, DNA was extracted from the five remaining combined half swabs and assayed for human DNA. Like the first combined swab DNA extract, no human DNA was detected in the second combined swab DNA extract. Next, all of the first handgun swab extract [20μl] and all of the second handgun swab extract [30μl] were combined and concentrated to 10 μl using Microcon filters. All of the concentrated handgun swab DNA extract [10μl] was subjected to a PCR based analysis of the nine Profiler Plus STR genes and amelogenin. The result of this analysis revealed an amelogenin X allele at ca. 6800 rfu and a Y allele at ca. 4500 rfu. Five alleles were detected at the D8S1179 locus [10, 12, 13, 14, and 15] that ranged from 651 rfu [12] to 104 rfu [13]. One other possible allele was detected at the D21S11 locus [31.2] at 57 rfu. No alleles were detected at the remaining seven STR loci. Unsatisfied with this result, the Cellmark technician then rinsed the tube originally containing the concentrated handgun swab extract with 10 μl of TE buffer and subjected this tube rinse to a second PCR based analysis of the Profiler Plus genes. The tube rinse analysis also produced a mixture with an amelogenin X allele at 2359 rfu and Y allele at 1077 rfu. Two or more alleles were produced at all nine STR loci where the major alleles are shared by defendant, Williams. Many of these alleles were in the rfu range of the Y allele. Thus, the rinsed out tube preparation appeared to be significantly less degraded than the original concentrated and consumed handgun swab DNA preparation and contained significantly more genetic information.

The Cellmark report authored by Rachel Cline and Lewis Maddox contained only the result from the rinsed out tube analysis and failed to reveal the result from the concentrated combined handgun swab extract that was consumed. The Cellmark report also failed to reveal that the published result from the handgun swabs was based on the analysis of a rinsed out tube previously containing the handgun swab concentrated DNA extract. The result obtained by Cellmark appears to violate well understood scientific principles such as conservation of mass, reproducibility of the same or similar sample, and irreversible direction of degradation.

This report presents the record and testimony from this analysis and queries whether or not Cellmark scientists have discovered new and previously unknown properties of nature.


DISCRIMINATION OF BEIGE CARPET FIBERS
Catherine Beitia, Sacramento County District Attorney, Laboratory of Forensic Services

The analytical scheme a forensic scientist uses to identify and discriminate among fibers in evidence cases was used to calculate the discriminating power of forensic analytical fiber protocols used currently in forensic laboratories. This project attempts to do this by looking at the individual beige to colorless carpet fibers found in common residential carpets within the Sacramento, California area. Two hundred eighteen carpet yarns were collected from a carpet retailer. Analytical tests performed on the carpet fibers included visual stereomicroscopy, polarized light microscopy, transmitted and reflected fluorescent microscopy, Fourier Transform Infrared Microspectrophotometry (FTIR), and UV/ Visible Microspectrophotometry. In addition, fibers were cross-sectioned, and modification ratios measured (an indication of circularity). The presence or absence of delusterant was also noted. Results of the study will be discussed.


DEVELOPMENT OF AN IN-HOUSE DNA LABORATORY INFORMATION MANAGEMENT SYSTEM (DNA-LIMS) TO INCREASE WORKFLOW EFFICIENCY
Patrick Paton, BS and Matthew Gabriel*, MFS, San Francisco Police Department Criminalistics Laboratory

Due to recent state and federal grant awards, the San Francisco Police Department Criminalistics Laboratory DNA Unit has had the opportunity to analyze >500 backlogged suspectless cases (e.g. homicides, sexual assaults, robberies and burglaries) in addition to active cases for the generation of DNA profiles for submission to the Combined DNA Index System (CODIS). The DNA Unit has addressed the need to expand casework testing capabilities and streamline the analysis process by implementing higher throughput laboratory instrumentation for DNA extraction, quantitation and genotyping. With this increase in the amount of casework data generated, the DNA Unit has also developed an in-house Microsoft Access based DNA Laboratory Information Management System (DNA-LIMS) to accommodate the data associated with these cases.

Over the past two years, the SFPD DNA-LIMS has been developed to meet the custom demands of the Forensic Biology Unit and serve as a fully integrated data management system for case and evidence tracking. Further functionality of the DNA-LIMS was designed to offer an effective analysis tool for data management relating to each step in the DNA analysis process (e.g. DNA extraction, quantitation with QuantiBlot™ or Quantifiler™, PCR amplification, ABI Prism 310 STR run, export of GenoTyper® data, DNA STR profile management for CODIS uploads and creation of genotype tables for laboratory notes). Since this system was developed with the primary objective of allowing the analyst to minimize time spent performing data management and administrative tasks for batches of case evidence samples (e.g. duplicate data entry and handwritten case notes), DNA-LIMS offers a streamlined approach to benchwork. Other useful administrative features of the DNA-LIMS include: on-line technical and administrative review, generation of Reports of Laboratory Examination, monitoring of casework statistics for each analyst or entire unit for a given time frame, managing casework photo documentation, recording communications, court dates and requests for discovery materials, tracking and maintaining quality control of reagents, reagent kits, pipette calibrations, supplies inventory, external proficiency tests, and so on.

The Microsoft Access based DNA-LIMS offers integration with current DNA analysis instrumentation such as the ABI Prism® 7000 Sequence Detection System for DNA quantitation and GenoTyper® software for creating and exporting genotype tables. Future implementation of a high throughput liquid handling platform such as the Bio-Mek® 2000 for DNA extraction, sample concentration normalization and PCR setup as well as the ABI Prism 3100 Genetic Analyzer / GeneMapper ID® for data analysis will offer additional data migration and management capabilities for even more efficient DNA testing. The flexibility of a system such as the DNA-LIMS also offers adaptability to other forensic science disciplines that require similar case management, evidence and sample tracking and laboratory data analysis needs.


THE ROLE OF THE CRIMINALIST WORKING FOR THE DEFENSE IN CRIMINAL CASES
Chuck Morton, Forensic Analytical

This is a personal review of 43 years of criminalistics experience working primarily for defendants in criminal cases. Since the large majority of criminalists work in public laboratories where the objective is to assist law enforcement and prosecutors in solving and prosecuting crimes it should be instructive to the members of this organization to get a sense of the similarities and differences in the roles we play in the American system of criminal justice. The CAC has historically been a professional home for a significant number of criminalists who do work for defendants. This has resulted in a greater appreciation, in California, of the defense criminalist's role than in much of the country. It has also contributed to the practice of a higher level of forensic science than encountered in much of the country. Although we all like to think of ourselves as, and presumably attempt to be, neutral scientists in carrying out our respective assignments, the context in which we work imparts specific responsibilities and limitations in our interaction with the criminal justice system and each other. A discussion of the practical implications of responsibilities and limitations influencing the work of the criminalist hired by the defense and examples of casework encountered over the years should provide the membership with an even greater appreciation of the role of the defense criminalist.


THE SEARCH FOR A SERIAL KILLER-THE ROLE OF DNA DATABASES
Mary M. Hong, Orange County Sheriff-Coroner

In 1995, the Orange County Crime Lab began to re-examine evidence in unsolved homicide cases with the intention of obtaining DNA profiles to enter into the DNA database (CODIS). This project was begun at this time due the availability of casework and offender databases in the State of California, and the routine use of DNA typing technologies, including RFLP and PCR-based methods.

Among these cases, three separate cases, which included a total of four homicide victims, were determined by DNA analysis to have been committed by a serial killer. Each of these involved a rape/murder of a female victim; in one of the cases the husband was also murdered. The initial analysis linking these cases was PCR-based DNA typing using CTT, DQalpha and D1S80. RFLP analysis was also performed on two of the cases to enable the DNA profile to be searched in the CA database and nationally through the then operating fax network. An investigative task force was formed which found a fourth case, from Ventura County. This case involved a female rape homicide victim and her male companion. DNA analysis demonstrated that this double homicide was committed by the same individual who had committed the previous crimes. After Profiler Plus and COfiler typing became available in 1999, the thirteen locus profile was developed and submitted to CODIS. In 2001, the Orange County Crime Lab was contacted by the Contra Costa Crime Lab with a request to compare a DNA profile obtained from three of their cases that were part of a series of rapes occurring in the late 1970's. The Contra Costa County profile was found to match that of the Orange County cases, thus linking a homicide series with six victims to a rape series with more than 50 victims. These cases are still unsolved. It is hoped that with the increased number of samples introduced to the offender databases with the passage of Proposition 69, this serial killer will be found.


PUMA PANEL: STR MARKER DEVELOPMENT FOR FORENSIC AND ECOLOGICAL APPLICATIONS
Julia A. Collins*(1, 3); Jay A. Well(1) & Holly B. Ernest(1,2,3,4);
(1) Wildlife and Ecology Unit, Veterinary Genetics Laboratory, University of California; (2) Wildlife Health Center, University of California; (3) Forensic Science Graduate Group, University of California; (4) Department of Population Health and Reproduction, School of Veterinary Medicine, University of California

Polymorphic microsatellite DNA markers (commonly called Short Tandem Repeats, STRs, in forensic literature) have become important tools of wildlife forensics and molecular ecology by providing accurate means of identifying individuals and characterizing the genetic structure and variation of wildlife populations. Microsatellite markers are particularly useful for studying threatened wildlife species such as puma (Puma concolor, also called mountain lion and cougar), which are highly secretive and dangerous to handle. An accurate method of identification is essential in cases of livestock predation, public safety incidents, poaching, and illegal capture. Accurate census and genotype data are needed to properly manage and conserve this protected species. The aim of this study was to test and optimize tetranucleotide STRs originally developed for the domestic cat (Felis catus) by the Laboratory of Genetic Diversity, National Cancer Institute, Frederick, MD, in contribution toward a polymerase chain reaction (PCR) multiplex of puma markers capable of providing discriminatory forensic match probabilities. Sex and species informative markers were investigated to augment the multiplex. The multiplex assembled in this study does not provide sufficient match probabilities, however additional puma-specific markers are in development.


JUSTICE AT LAST: WHO REALLY KILLED STEPHANIE CROWE
Connie Milton, Criminalist, San Diego Sheriff's Crime Lab

On the morning of January 21, 1998 twelve year old Stephanie Anne Crowe was found murdered in her bedroom. Her entire family was home at the time of the murder. Witnessed in the area the night before was a schizophrenic transient. Initially charged with her murder were Stephanie's older brother, 14 year old Michael, and two of his teenaged friends. When Stephanie's blood was later found on the transient's clothing, charges against the boys were dropped. But how did the blood get there? Was the transient a killer, or just the unlucky subject of police contamination? Examination of the physical evidence led to the eventual resolution of this case and justice for the murder of Stephanie Crowe.


A COMPARISON OF THE EFFECTIVENESS OF SWABS AND FLOSS AS A MEANS OF RECOVERING SPERMATOZOA FROM THE ORAL CAVITY
Sherille L. Cruz, M.S.(1,2); Katherine A. Roberts, Ph.D.(1,2); Donald J. Johnson, M.S.(1,2); Barry A. J. Fisher(1,3)
(1) California Forensic Science Institute; (2) School of Criminal Justice and Criminalistics, California State University, Los Angeles; (3) Los Angeles Sheriff's Department, Scientific Services Bureau

The investigation of cases involving allegations of sexual assault can prove to be problematic. One compounding issue is the fact that there are usually no witnesses to the crime. This can mean that accounts of the alleged incident often conflict. While physical evidence, such as semen, saliva, and physical trauma establishes an association between the victim and the suspect, this is contingent on the successful recovery of evidence. The particular focus of this study was to evaluate several factors that influence the successful recovery of spermatozoa from the oral cavity. A total of 86 oral cavity samples were collected from 9 individuals. The samples were sub-characterized according to the collection method used: 43 were collected using floss and 43 were collected by swab. Successful recovery of spermatozoa was assessed as a function of three variables: The collection method; the time elapsed, post-copulation; and, the effect of oral activity post-copulation.

All of the samples were subjected to a differential extraction procedure prior to microscopic evaluation of the extracted pellet using hematoxylin and eosin staining. The microscope slides were examined at X200 and X400 magnification and the concentration of spermatozoa per microscopic field of view was scored. Spermatozoa were successfully recovered in 47% of the 86 samples included in this study. Further, the results demonstrate differences in recovery of spermatozoa as a function of the collection method: 54% of the swabs were successful in recovering spermatozoa compared to 40% of the floss samples. As a general trend, the average concentration recovered (combining the floss and swab data) decreases as the post-copulation time interval increases; the greatest decline in average concentration recovered occurs within 1.5 to 6 hours. However, spermatozoa were successfully recovered from 4 samples even at the 24-hour post-copulation time interval. Finally, the data from this study suggest that spermatozoa recovery from the oral cavity decreases as oral activity increases. The floss collection method has been shown to recover spermatozoa in cases where the swabs were negative. Further, in cases where both the floss and the swabs are successful in recovering spermatozoa, the extract can be combined for analysis purposes. Therefore, the inclusion of floss in California's standardized sexual assault kit is recommended to facilitate, not replace, swabs in the collection of spermatozoa from the oral cavity.


A NEW RAPID AND EASY-TO-USE DIFFERENTIAL EXTRACTION METHOD
Yasser Daoudi*, Kimberly Huston, Allan Tereba, Laura Flanagan, Paraj Mandrekar, and Ryan Olson, Promega Corporation

One of the main reasons for the large backlog of sexual assault samples is the difficultly in working with the evidentiary material. Typical vaginal swabs contain a mixture of victim epithelial cells in large excess over sperm cells. Unprocessed, these samples can only be analyzed using male specific markers that provide important evidence but are of limited use in searching national databases due to the inheritance and nonrecombinatorial nature of the Y chromosome.

In 1985, Gill et al. developed a method to enrich for sperm cells in the presence of an excess of epithelial cells. After a controlled proteolysis in the absence of a reducing agent, the sample is centrifuged in a spin basket to remove from the solid matrix intact sperm and solution containing the DNA from lysed epithelial cells. Because the resulting sperm pellet contains loose cell debris a considerable amount of contaminating solution is left and must be diluted out with serial washings and centrifugations. This process is time consuming and results in loss of sperm and variability between examiners. We have developed a new differential extraction method that takes advantage of the nearly two decades of experience using the standard differential extraction. After a standard Proteinase K digestion of the sample, the solid support and DNA-containing solution are centrifuged through a special material that effectively separates the sperm from soluble DNA and cell debris. The samples are washed once without centrifugation to remove any remaining soluble DNA in the sperm fraction. DNAIQ™ Lysis Buffer containing DTT is then added to the epithelial and sperm fractions. This buffer effectively lyses the sperm without need for further Proteinase K digestion. The total time for separating the sperm from epithelial cells following addition of the sample to the Proteinase K Digestion Solution is approximately 1 hour 20 minutes which includes the 1-hour Proteinase K digestion. The purification of the DNA requires 40 minutes so the total separation and purification can be accomplished in 2 hours.

Because the same standard Proteinase K digestion and initial centrifugation is used to help remove the sperm from the solid support and to lyse the epithelial cells, the efficiency of these steps will be identical to what is currently available. However, only one centrifugation is required for efficient separation so the sperm recovery is better. In addition, the hands on time as well as the overall time needed to do the separation has been greatly reduced from the current method. Data will be presented on the sensitivity and successful processing of old samples.


SUMMARY OF THE APRIL 2005 FORENSIC SCIENCE SOCIETY CONFERENCE
Adam Dutra, San Diego Police Department

I was honored to receive the CAC's Paul Kirk and President's Awards, which include a paid trip to a Forensic Science Society Conference. I attended the April 2005 conference in Leeds entitled "Homicide Investigations". To thank the CAC for selecting me for the award, I would like to provide a summary of the meeting and highlight a few of the more interesting papers.


HOW "UNIQUE" ARE IMPRESSED TOOLMARKS? - AN EMPIRICAL STUDY OF 20 WORN HAMMER FACES
Eric R. Collins

The examination of impression evidence has long been based upon the assumptions that a particular impressed mark may show uniqueness due to its physical characteristics and position relative to other marks or some point of reference. Traditionally, firearm and toolmark examiners have had to rely upon the criteria of whether or not the appearance of an impressed toolmark exceeds the "best known non-match" when drawing conclusions of identity or non-identity. Until now, no study has been performed or attempted to quantify the uniqueness of impressed marks based upon their observable physical characteristics. This has presumably been due to the difficulty with which impressed (or compressed) marks lend themselves to the characterization of their individuality. Rocky Stone recently authored an article in the AFTE Journal in which he provided some theoretical probabilities associated with "idealized" impressed toolmarks that might be found on a hypothetical hammer face. Stone's work also established a model on which an empirical study could be based. This research project represents such a study, the purpose of which was to test the applicability of Stone's conclusions to real toolmarks by examining the nature of actual impressed marks found on the faces of twenty (20) hammers that had been subjected to various degrees of wear and abuse through normal use. In addition, this presentation will establish a foundation for evaluating the practical statistical uniqueness of impressed toolmarks on not only hammer faces, but any given surface.


HISTORY OF MICROCRYSTAL TESTS IN FORENSIC SCIENCE
Hiram K. Evans, M.Sc., F-ABC, San Bernardino Co Sheriff's Department Scientific Investigations Division

The early history of microcrystal tests is the history of chemistry and microscopy. By the mid-1830's toxicologists needed something besides the drastic chemical treatments applied to heavy metal poisons for application to alkaloidal poisons. While any history moves forward in small steps, microcrystal tests in forensic science have a series of watershed dates. 1865 brought Helwig's Das Mikroskop in Der Toxicologie and Wormley's Microchemistry of Poisons. By 1921 and the publication of Behrens-Kley's Organische Mikrochemische Analyse and Stephenson's Some Microchemical Tests for Alkaloids forensic science had expanded to include the identification of controlled drugs. 1934 and 1935 saw the publication, respectively, of Amelink's Schema zur Mikrochemischen Identifikation von Alkaloiden and Rosenthaler's Toxicologische Mikroanalyse. From the 1920's through the 1960's, frequent collaborative work was performed and published in JOAC, expanding application and introducing acid reagent media. 1969 was probably the greatest year with publication of E. G. C. Clarke's Isolation and Identification of Drugs and Charles C. Fulton's Modern Microcrystal Tests for Drugs. Publications on microcrystal tests have decreased in number, concentrating on determination of isomeric forms, but the tests remain part of some training programs, are included in new ASTM Standard Guides, and are accepted by ASCLD/LAB for use in accredited laboratories.


OVERLOOKED CLUES, DOWNRIGHT CONSPIRACIES, OR JUST MONDAY- MORNING QUARTERBACKING: YOU BE THE JUDGE: AN INTERACTIVE PRESENTATION
Alan Keel, Forensic Science Associates

Pre-trial and post-conviction peer reviews/re-examinations protect an accused person's constitutional right to confront the evidence used against them at trial. By default, that process also provides the opportunity to assess the work of the prosecution experts who proffered that evidence on behalf of the People. All too often, when shoddy, incompetent, or even purposefully misrepresented work for the government is revealed, that appraisal is dismissed by peers, supervisors, and bureaucrats as unfortunate misadventure rather than as demonstration of systemic training and oversight inadequacies, and criminal justice system corruption. These deficiencies are clearly demonstrated in numerous cases revealed during defense examination by evidence that was:

Due to time constraints, this presentation will focus on evidence that was discovered pre-trial and mishandled at the lab bench, along with the resultant fallout. Cases from California, Nevada, Michigan, Illinois, Maryland, and others will be presented as time allows. The facts and appropriate visual displays of evidence in each case will be provided in an anecdotal summary of the case background and its progress through our criminal justice system. Those in attendance will have the opportunity to recognize the pertinent evidence or clues in each case that is not overtly addressed and voice their appraisal at any time. Causative factors for each "misadventure" will be elicited from the audience as well. This interaction should be educational and entertaining, and hopefully, all in attendance will gain a new perspective for personal introspection regarding one's own impact upon our profession, and the effectiveness of current in-house and legislated oversight methods in our criminal justice system.


FORENSIC GLASS ANALYSIS BY LA-ICP-MS: ASSESSING THE FEASIBILITY OF CORRELATING WINDSHIELD COMPOSITION AND SUPPLIER
Abbegayle J. Dodds* (1,2); Donald P. Land(1); And Edward M. "Chip" Pollock(2)

(1) University of California Davis, Department of Chemistry and Graduate Group in Forensic Science; (2) Sacramento County District Attorney's Laboratory of Forensic Services

Recently, much attention has turned to the use of elemental analysis in comparing glass fragments of different sources, which may appear similar by refractive index measurement. Trace elemental analysis appears to be one of the most distinguishing techniques available and has been established as a forensically valid tool for glass examination. Preliminary results are presented that evaluate the potential utility of associating automotive windshields and manufacturers by trace elemental composition. To our knowledge, a systematic study addressing the variability of trace elements in windshield glass has not been published. This is an important first step in determining if manufacturers and windshields can be associated by composition alone.

A three-part study was conducted. First, windshield glass homogeneity was investigated. Ten windshields, representing both domestic and international manufacture, were multiply sampled across the length of the windshield. Samples were taken such that both layers of glass could be examined. Where possible, windshields suspected to be produced within the same batch were included in this set. Second, the resulting elemental profiles of these individual windshields were then compared to each other. This provided preliminary data regarding what variation in elemental profile to expect among different sources. Finally, batch samples of float glass obtained from a single production line were examined. These batch samples were taken from the left, center and right sides of the glass ribbon, multiple times a day for thirty days.

These analytical results provided invaluable information regarding windshield homogeneity and population variation. We present this data to offer practitioners a basis for assigning significance to elemental profiling of windshield glass.


POLICE OFFICER FATALITY SHOOTING RECONSTRUCTION
Mark Bennett, Criminalist II, Oakland Police Department

While responding to a home invasion call, a sergeant with the Tallahassee Police Department was fatally shot. The suspect was apprehended a short time later in possession of a Smith &Wesson .357 Magnum revolver, containing six fired cartridge cases. There were no witnesses to the incident and the sequence of events became key to the capital offence trial in which the defense counsel claimed the defendant "saw an unknown figure in the distance and, fearful for his life, shot his gun while running away." Evaluation of evidence, including examination of the victim's gunshot wounds, uniform clothing, location and characteristics of recovered bullets, bullet comparisons, police radio audio tape, and the sequence of cartridges in the suspect's revolver provided a high confidence reconstruction of the sequence of events. The forensic reconstruction was illustrated in court using computer 3D rendering software that graphically showed the likely relative positions of the gunman and officer for each shot.


THE MAYFIELD CASE-ANATOMY OF ERROR
Kenneth Moses, Director of Forensic Identification Services in San Francisco

In May of 2004, Brendan Mayfield was released from custody as a material witness when the F.B.I. disclosed that it had made an erroneous identification of his fingerprint. This presentation will examine how and why the error occurred as well as future implications to the field of forensic identification.


Y-STRS COME OF AGE-A DISPUTED INTERPRETATION
Norah Rudin

A case in which DNA was typed using Y-STRs will be discussed. The interpretation of the primary analyst was disputed. Issues include: partial profiles, expectations for duplicated loci, detection of homologous sequences in female DNA, laboratory contamination, and non-concordant amplification results. Working in the military justice system will also be discussed if time permits.


PREPARING FOR AN ACCREDITATION UNDER ISO 17025 STANDARDS
Scott R. Oulton

Accreditation is becoming a de facto requirement for the recognition of a laboratory's operation under documented policies and procedures. As such, the objective of this program is to synopsize the procedures undertaken by the Drug Enforcement Administration (DEA) to prepare analysts and laboratories for accreditation under ISO/IEC 17025 standards. An important part of this program will involve discussions of those accreditation requirements which caused a significant degree of concern and fell outside of the "comfort zone" of familiarity with previous accreditation inspections. The program will also describe in detail how laboratory program standards were enhanced to meet the requirements for accreditation. It is intended that the discussions in this program will be candid and the goal is to answer questions and present suggestions for achieving accreditation under ISO.


IDENTIFICATION OF AN INOPERABLE FIREARM TO BULLETS AND CARTRIDGE CASES FROM A HOMICIDE FOURTEEN YEARS PRIOR
Ron Nichols, Bureau of Alcohol, Tobacco, Firearms and Explosives, Forensic Science Laboratory

A recent case involved the comparison of bullets and cartridge cases from a homicide in 1990 to a firearm recovered from a drainage ditch, approximately 12 years later. The firearm was received in an inoperable condition. The firearm was dismantled and the various parts of the firearm that may be expected to come into contact with either the cartridge case or the bullet were cleaned with gun solvent and rust dissolver. The breechface of the Raven pistol was cleaned sufficiently to reveal concentric circular tool marks from the finishing process. The firing pin was removed from the pistol and cleaned sufficiently to reveal the surface of the firing pin before it became encrusted with foreign debris. The cast of the breechface was compared with the cartridge cases that were recovered from the scene of the shooting and the firing pin was compared with casts of the firing pin impressions present on the same five cartridge cases. There was sufficient agreement to conclude that the cartridge cases were fired in the submitted Raven pistol. The barrel of the Raven was cleaned. A cast of the barrel showed significant deterioration of the rifling through most of the barrel exclusive of the muzzle end. I obtained test marks from the muzzle end by swaging a piece of lead into that area and then punching it out of the muzzle end, simulating the exit of a bullet. The lead swage was compared with the bullets recovered from the victim and a significant amount of corresponding striations was observed to permit a conclusion that the bullets were likely fired from the Raven pistol. Despite the deteriorated, inoperable condition of the pistol, it could be dismantled and adequate test marks produced to compare with submitted bullets and cartridge cases. Sufficient individual markings were present to permit a positive association between the recovered pistol and the submitted bullets and cartridge cases.


FINGERPRINTS AND DNA ON CARTRIDGES AND CARTRIDGE CASES: HOW LIKELY?
Terry Spear *, Jeanne Clark, Mike Giusto, Neda Khoshkebari, Michael Murphy and John Rush, California Department of Justice, Bureau of Forensic Services

This study examined the likelihood of: (1) obtaining fingerprints on fired and unfired cartridges and cartridge cases [cartridges/cases] and (2) obtaining DNA typing results from any of the cartridges/cases processed for fingerprints. Three types of fingerprints (bloody prints, sweat prints and oily prints) were placed on brass, nickel-plated and aluminum cartridges. Half of the test cartridges were fired after receiving the fingerprints. The cartridges and cartridge cases were then processed for fingerprints using either amido black or cyanoacrylate fuming/ dye staining. After the cartridges and cartridge cases were processed for fingerprints, they were swabbed to collect DNA. No usable prints were observed on any of the twelve smaller (22LR) cartridge cases (N=12) regardless of the type of fingerprint or fingerprint processing method used. In contrast, 6 useable prints were developed on the 9mmP and 45ACP cartridges (N=36). Thus, only 6 prints were classified as useable or identifiable on the 48 cartridge/cases examined. The swabs used to collect the DNA from the cartridges/ cases were organically extracted for DNA. Since these samples were not expected to yield very much DNA, the extracts were not quantified but were directly amplified for STR markers using Applied Biosystems's Profiler Plus reagent kit. Only 3 STR profiles were obtained from the 48 cartridges/cases that were tested: two profiles from 2 unfired cartridges and one profile from a fired cartridge case. Since fingerprint processing results in the loss of DNA, it is likely that more STR profiles might have been obtained if these cartridges/cases had not been first processed for fingerprints.


SUBJECT'S WALLET FOUND AT SCENE- HOW DO THE CLUELESS LEAVE SO MANY CLUES?
Susan Morton

How is it that criminals who have nary a clue to start with strew so many for the discerning investigator to find? This is a philosophical question in need of much pondering. We will examine a number of cases in which it is clear that the perpetrator would lose a debate with your average root vegetable. After hearing this presentation, the audience will come to the humbling realization that we don't catch the smart ones.


SPME VS. CHARCOAL ADSORPTION: A COMPARATIVE STUDY OF EXTRACTION METHODS FOR VOLATILE ACCELERANTS PRESENT ON HUMAN SKIN, OR, COLONEL MUSTARD WITH GASOLINE IN THE BEDROOM- CAN WE PROVE IT?
Katherine Hutches*(1), John D. DeHaan(2) and Don P. Land(3)

(1) Graduate Group in Forensic Science, University of California at Davis, (2) Fire-Ex Forensics, (3) Department of Chemistry, University of California at Davis

When suspects are interviewed during arson investigations, the odor of gasoline or other volatile accelerants is sometimes detected on their hands. Attempts to remove these residues for identification have, in the past, relied on sampling of the hands with either dry or solvent-wetted swabs. These techniques have almost invariably failed due to the low concentration of volatile absorbed into the skin. Methods of non-invasive extraction of fire debris that are widely used in general arson investigation include: charcoal adsorption/elution and solid phase microextraction (SPME). This study sought to determine which, if either, of these methods is suitable for use as a test for the presence of volatile hydrocarbons on the skin and whether there is selective absorption of components by the skin that affect the ratios of extractable volatiles in mixtures such as gasoline. While these methods are currently applied to the extraction of volatile compounds present in fire debris, these methods may show promise in extracting volatiles from skin. Preliminary results demonstrate differences in the utility of each of these two methods in terms of accuracy and viability for use in short-term samples.