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Serological Evidence
SEROLOGICAL EVIDENCE
IN SEXUAL ASSAULT INVESTIGATIONS
By
Robert R.J. Grispino, M.A.
Special Agent
Serology Unit, Laboratory Division
FBI Headquarters
NOTE: The following article presents a purely scientific
approach to sexual assault evidence collection. The
scientific step-by-step procedures that are explained here
should always be accompanied by supportive treatment of the
victim. It should also be noted that the investigating
officer will be responsible for both overseeing the
execution of the medical procedures described and managing
the collection of the physical evidence.
Police officers throughout the United States routinely
handle and oversee sexual assault investigations. Yet, these
officers rarely receive training on the proper methods to be used
for sexual assault evidence collection and preservation. As a
result, valuable physical evidence may either be overlooked or
inadvertently allowed to deteriorate biologically. This article
establishes proper evidence collection and preservation protocol
in sexual assault matters and demonstrates how modern forensic
serology can aid in the eventual successful prosecution of the
assailant.
AFTER THE ASSAULT
A sexual assault has been committed. Upon arriving at the
scene, an officer usually encounters a distraught, possibly
hysterical, victim in the first psychological coping stages
after an intrusively brutal assault. At this time, the victim
is not usually receptive to examination, even though the ideal
time to collect body fluid evidence is immediately following the
assault. This best evidence of the crime is present, but
unfortunately, it is degrading with every elapsed minute. It is
crucial that the officer be aware of the need for immediate
examination in order to ensure that evidence is collected
properly.
The victim should be transported expeditiously to a local
hospital or rape trauma center where trained nurses or
physicians can gather the appropriate physical evidence. The
investigating officer is not only responsible for accompanying
the victim through the phases of the examination but must also
oversee the collection and preservation of medical evidence. By
being fully aware of the procedures that should take place and
the evidence that can be collected, the officer will be able to
ensure that the case is as complete as possible.
During this examination period, health professionals should
accumulate and catalog physical evidence. The completeness of
the physical examination depends on the care, consideration, and
thoroughness of the examining physician. As much evidence as
possible should be obtained from the victim. However, if the
health professional does not gather all of the available
background data, the investigating officer can and should step in
to make sure that it is elicited entirely from the victim.
INTERVIEWING THE VICTIM
The initial victim interview is notoriously the most
incomplete in sexual assault matters. The excuses are numerous.
The victim is hysterical and has to be sedated. The doctor or
nurse is not willing to delve into any particulars of the assault
for fear that it will further upset the victim. The
investigating officer is embarrassed or unsure of the situation.
Yet, regardless of these various problems, a full and complete
description of the assault is crucial to obtain the physical
evidence needed for a successful scientific analysis.
Obvious questions regarding the date, time and location of
the assault, as well as the description or identity of the
subject and the general circumstances of the assault, are rarely
missed. However, the less obvious questions can be just as
important. The interviewer should ascertain whether the victim
bathed, showered, douched, urinated, defecated, vomited, brushed
teeth, or gargled with mouthwash at any time after the assault
and prior to the examination. If any of these activities were
performed, the probability of obtaining useful serological
results could be greatly diminished.
The physician should then discuss the basics of the assault.
It should be determined from the interview whether penile
penetration of the vagina, anus or mouth was attempted and/or
successful, as well as if the perpetrator ejaculated at any time.
If penetration was unsuccessful or not attempted in certain
orifices, the physician may opt not to collect evidence from the
unviolated areas. The physician must realize that it may be
difficult or impossible for a victim to know whether the suspect
ejaculated or there may be a psychologically induced reluctance
by the victim to admit during the early post-assault period that
ejaculation occurred. In these instances, failure to collect
samples from all body cavities may result in lost body fluid
evidence.
The extent of oral/genital sexual contact, whether fellatio
or cunnilingus, should be evaluated. If contact was indicated,
the medical examiner should externally swab the genital area for
later serological analysis. It would also be important to
determine whether the assailant used a lubricant or condom during
the assault or whether the victim was menstruating.
The victim should also be asked about the particulars and
extent of any consensual intercourse within a 72-hour period
prior to the assault. The final serological conclusions reached
from the scientific analysis of the physical evidence of the
assault may be greatly affected by previous consensual sexual
activity and resultant body fluid mixtures. It would then be
important to obtain blood and saliva samples from the previous
partner(s) for comparison and elimination purposes.
The presence and extent of any injuries to the assailant
inflicted by the victim is also extremely important. The suspect
may have bled on the victim from a victim-inflicted injury during
the struggle. A victim during menstruation could also transfer
blood to the suspect during vaginal assault, which could be
serologically identified in the suspect's underwear.
After all background information is obtained from the
victim, the staff nurse and attending physician should conduct a
thorough pelvic examination of the victim. All signs of pelvic
injury should be documented, either in drawing or photographic
form. After the background information has been gathered, the
next step is to collect physical evidence from the victim as
carefully and thoroughly as possible.
COLLECTING PHYSICAL EVIDENCE
Many of the items used for evidence collection are available
in sexual assault kits. However, these commercial kits vary
widely, and basic minimums should be considered.
Vaginal, oral and/or anal swabs should be taken from the
victim using sterile cotton swabs. These swabs should then be
air-dried, appropriately labeled, initialed by the examiner and
packaged separately. In most assaults involving vaginal
penetration, two to four vaginal swabs and two cervical swabs are
adequate for analysis. In cases of oral or anal sodomy, oral or
anal swabs should be obtained from the victim. Two clean swabs
taken from the same package as the unstained control swabs should
be submitted to show that any useful serology results obtained
during analysis were due to body fluids and not any contaminant
initially on the swabs.
Smear slides, unfixed and unstained, are sometimes useful
for demonstrating the presence of sperm cells (spermatozoa).
Vaginal, oral and/or anal smear slides should be obtained from
the victim using the same swabs mentioned above. The smear
slides should be appropriately labeled and should indicate which
individual swab was used to create which microscope smear slide.
Examining physicians in some jurisdictions prepare and
microscopically examine smear slides to determine the presence of
motile sperm cells indicative of recent sexual activity. In such
cases, examining physicians may be required to testify in court
proceedings regarding their observations. In any event, stained
and fixed smear slides are useless for further serological
analysis and should not be submitted to crime laboratories.
Pubic combings should be taken from the victim to identify
any foreign hairs or fibers that may have been transferred during
the assault. The physician should comb the pubic area and submit
the comb and any resultant debris in an appropriately marked,
sealed envelope. Head hair combings should be obtained from the
victim in cases where other evidence is insufficient to show
interpersonal contact. Pubic and head hair combings should also
be obtained from the suspect if appropriate to the investigation.
Any obvious debris (soil, fibers, hair, grass, etc.)
observed during the examination of the victim should be collected
and submitted in a separate envelope describing the location of
the debris. The examining physician should also scrape all
residue from under the fingernails of each hand of the victim and
place the residue in a specimen envelope or clip the fingernails
and place the clippings in separately labeled envelopes.
Using a sterile pad that has been moistened lightly with
distilled water, the physician should swab the vulva and the
inner portion of the victim's thighs adjacent to the vaginal
area. The genital swabbing pad should then be air-dried and
submitted for laboratory analysis in an appropriately labeled
specimen envelope.
The physician should swab any dried secretions observed
during the examination of the victim, i.e., saliva around bite
marks, using a sterile pad that has been moistened lightly with
distilled water. The pad should also be air-dried and submitted
for analysis. In cases where dried blood or encrusted semen is
observed, the material should be scraped from the body into a
specimen envelope and submitted for analysis. Encrusted matter
should never be re-hydrated, since it dilutes the sample. The
location of each sample should be noted on a body diagram. Pubic
or head hair containing encrusted semen should be carefully
clipped and placed in a labeled specimen envelope.
In the event of oral ejaculation, gagging, swallowing or
regurgitation during the assault may force air carrying semen
through the nasal passages. The victim should blow her nose,
very hard, several times into the center of filter paper. The
resultant nasal mucous sample should be allowed to air dry and
then submitted for analysis.
STANDARDS
Head hair and pubic hair standard samples should be obtained
from the victim and any suspects developed from the sexual
assault investigation. The hair samples should be pulled with
the bulb intact, not clipped. Head hair samples should be taken
from four separate areas of the scalp. Twenty-five full-length
hairs are generally considered adequate to represent an
individual's hair characteristics.
Liquid blood samples should also be obtained from the
victim, any consensual sexual partners from at least 72 hours
prior to the assault, and any developed suspects. Known blood
and saliva samples from a suspect in a sexual assault case must
usually be obtained through a court order issued by a judge or
local magistrate. Blood samples from each individual should be
collected in both red-topped and purple-topped blood collection
tubes. Red-topped tubes are used for traditional serological
analysis, such as ABO grouping, secretor status and enzyme
electrophoresis. A red-topped tube indicates that the collected
blood is exposed to no preservatives or blood anticoagulants.
Purple-topped tubes are used for DNA profiling only. These tubes
contain a chemical chelator (EDTA) that inhibits the action of
enzymes that would normally act to break down the DNA molecules
in the blood samples. In the event that toxicology examinations
will be requested, an additional blood sample taken in a
grey-topped tube (containing sodium fluoride) and a 10 cc. urine
sample should also be collected. All of the collected blood and
urine should be refrigerated, not frozen, and submitted for
analysis as soon as possible.
Dried saliva samples should also be obtained from the
victim, from consensual sexual partners from at least 72 hours
prior to the assault, and from any developed suspects. The donor
should expectorate on filter paper to produce a stain
approximately 1 1/2 inches in diameter. Saliva should be clean
and undiluted. Prior to giving the sample, the donor must have
abstained from eating (food, gum, chewing tobacco), drinking and
smoking for about 30 minutes. The stain should be circled in
pencil before the drying is complete. When the samples have
air-dried completely, they should be placed in a specimen
envelope that has been dated and initialed.
MISCELLANEOUS SAMPLES
The officer should make sure that the clothing worn by the
victim during the sexual assault is collected. The victim should
always disrobe over examination paper. The victim's panties,
pantyhose, jeans, shirt, shoes, socks, dress, or nightgown should
be separated and individually packaged as appropriate. Any
physical evidence from the crime scene that may bear suspected
semen stains, such as bed sheets, towels, wash cloths, paper
towels, toilet paper or tissue paper, should also be collected.
The examination paper should also be submitted for analysis in
the event that hair or fiber mixtures from the assault fell from
the victim while disrobing.
The collected items should be clearly described for the
laboratory, including whether the items came in contact with the
victim and/or suspect before, during, and/or after the assault.
Stained areas believed to exhibit evidence of the assault should
be described or highlighted. For example, only a small area on a
bedsheet may be relevant to the investigation. Therefore,
forensic examination of the entire bedsheet for semen may not
only be unnecessary and wasteful of forensic services but may
also dilute the effectiveness of the examination.
PRESERVATION AND PACKAGING
Bacteria begin to degrade biological fluids immediately
after deposition. They especially thrive on the rich nutrients
present in semen. If unchecked, contaminant bacteria can
completely destroy DNA and other genetic markers of value. To
counteract this phenomenon in all of the above instances in
which moist body fluids are collected, it is imperative that the
samples be completely dried. After drying, the specimen(s)
should be placed into breathable paper bags or envelopes and
frozen or refrigerated until submitted to the laboratory for
analysis.
All collected items of evidence should be properly
catalogued with preserved chain-of-custody records for court
presentation purposes. All items should be dated and initialed
by the collector. In cases where samples were taken by health
professionals, they should identify, date and initial the items
and hand the evidence to the investigating officer. Whenever
possible, collection of known blood, urine and saliva samples
should be performed under the supervision of the investigating
police officer.
CONCLUSION
Each sexual assault occurs under circumstances unique to the
victim, the crime scene, and the suspect. If extensive
information is provided to the examiner in the crime laboratory,
the examiner can conduct a more thorough and complete scientific
analysis. Probative value and relevance to the investigation are
the watchwords in collecting and preserving the evidence of a
sexual assault. If collected materials are stored in plastic
bags under room temperature for any extent of time, the
biodegrading action of contaminant bacteria may jeopardize
conclusive test results. Sexual assault evidence kits with all
of the above-mentioned materials are commercially available and
stocked by hospitals and rape crisis centers.
Proper collection, identification, packaging and storing of
evidence in sexual assault investigations will greatly improve
the chances for a successful prosecution of the perpetrator, as
long as the investigating officer follows up with good
communication and contact with the laboratory examiner in all
stages of the case. The evidence sent to the forensic laboratory
should be accompanied by a transmittal letter that completely
describes the facts of the crime, the inventory of the evidence
seized, and the scientific examinations requested. It is
suggested that the report of the initial examining physician be
included along with the evidence.
Criminal investigators and prosecutors must familiarize
themselves with proper procedures concerning the collection,
identification, and packaging of serological evidence, as well as
to establish effective communication with the examiner. By doing
this, and by understanding the practical capabilities and
limitations of modern forensic serology, the interests of justice
can be best served with a successful prosecution of the
perpetrator in a sexual assault.
APPENDIX A:
THE ROLE OF SEROLOGY
Forensic serology is best defined as the science involving
the identification and characterization of blood, semen, and
other body fluids, usually found in dried stain form on items of
physical evidence. Because of its supportive nature to the
prosecution, it is absolutely essential that the investigators
and the prosecutor understand, at least in general terms, the
capabilities, as well as the limitations, of forensic serology.
Under most conventional serology protocols, items of
physical evidence in sexual assault cases are sceintifically
screened for the presence of human semen and blood. Semen is
identified where sperm cells are microscopically identified
and/or a semen-specific protein associated with human semen,
known as p30 or prostatic antigen, is determined to be present
in extracts of dried stains under examination.
Once the presence of semen is established, the stain
extracts may be analyzed for the presence or absence of blood
group substances. Eighty percent of the general population
secrete chemicals (blood group substances) into their body
fluids that are consistent with their red blood cell ABO type.
In these people, known as secretors, analysis of their saliva,
vaginal secretions, or semen will indicate whether they belong
to blood group A, B, AB or O.
If human blood is identified on an item, attempts may be
made to determine whether the blood is of A, B, AB or O. Then,
depending upon the size of the dried stain, it may be analyzed
using electrophoresis to determine as many genetic marker
protein types as possible. In order for any of this to make
sense, known blood and saliva samples from both the victim and
suspect of the sexual assault must be submitted for analysis and
comparison purposes.
The known blood samples may be grouped as to A-B-O blood
type and assorted genetic enzyme markers. The serologist may
also attempt to determine if the individual is a secretor from
analysis of the liquid blood and/or dried saliva standard.
Once all of the scientific information is assembled, the
serology examiner may be able to make expert conclusions from
the findings. Forensic serology is a comparison science. If
all of the information from the analysis of the questioned
samples is identical to that obtained from the known blood and
saliva samples from the suspect, then the serologist can
conclude that the suspect was a possible source of the deposited
semen or blood stain. If, however, one element of the profile
differs, then the serologist may be able to absolutely exclude
the suspect as a source of that body fluid. It is further
emphasized that without known standards from the victim,
suspect(s) and any other involved parties for comparison, the
effective serological analysis of items of evidence from the
assault may be rendered meaningless.
Using this technology, the best that a serologist may be
able to say is that the suspect is a possible depositor of the
body fluid. This is because other potential suspects in the
general population may share the same A-B-O blood type, secretor
status, and enzyme profile, althought the implementation of DNA
testing in forensic samples has dramatically altered this
thinking.
APPENDIX B:
DNA PROFILING
Deoxyribonucleic acid (DNA) is an organic substance found
primarily in the chromosomes within the nuclei of cells. Using
electrophoresis and radioactive probing techniques, a DNA
profile can be developed from dried blood and semen stains.
DNA profiling is the FBI Laboratory's primary method of
choice for the serological analysis of physical evidence from
violent personal crimes. This technology has revolutionized the
processing of serological evidence and has superceded
traditional serology techniques in its associative value.
Currently, the FBI Laboratory screens probative items of
evidence in sexual assault matters, such as vaginal, oral, and
anal swabs and panties from the victim, for the presence of
semen. At the conclusion of this analysis, the remaining stain
may be submitted for DNA profiling. If human semen is
identified, the sample is submitted for DNA analysis.
DNA profiles in the questioned samples are compared side by
side with the DNA profiles in the known blood samples from the
victim and suspect. If the patterns on the suspect's blood
profile match those generated from the questioned samples, the
serologist can testify that the semen or blood was deposited by
the suspect or a member of a group of individuals who share this
profile. As with conventional serology, if DNA profiles do not
match, the serologist can absolutely exclude the suspect as the
contributor of that DNA sample.
Many U.S. crime laboratories do not possess the capability
to conduct DNA testing. Therefore, investigators and
prosecutors should become familiar with the capabilities of
their local laboratories in this regard. The current protocol
used at the FBI Laboratory necessitates an 8- to 10-week time
period from initial DNA extraction to final probing results in
each case sample. Additonal delays may also exist due to high
caseload and the requirement for analysis in other forensic
disciplines. DNA analysis may be more time-consuming and labor
intensive than traditional serology techniques; however, the
results may be far more significant or informative. The law
enforcement community must be aware that DNA analysis of
forensic samples is a lenghty process and trial continuances may
be necessary.
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