Examination of Firearms Review: 2007 to 2010 - Interpol

used in the RT reaction. The expression of miRNAs was subsequently examined
using a SYBR Green real time PCR assay. Unfortunately this procedure cannot be
multiplexed and has to be performed in a separate reaction for every miRNA 16 .
Nonetheless attempts could be made to develop multiplex reactions using multiple
dyes. Zubakov et al 15 in 2010 reported three miRNAs (miR-20a, miR-106a, and
miR-185) that have high expression levels in circulatory blood relative to semen,
saliva and vaginal fluid. They used the TaqMan MicroRNA Reverse Transcription
assay, which is species specific, however there are miRNAs which do appear in other
species (e.g. non-human primates). Of the three miRNAs mentioned above only
miR-106a is human specific.
3.3 Semen
3.3.1 Current confirmatory techniques
The tests that can be used for the identification of semen are the microscope test,
immunological tests, and electrophoretic methods 2 .
The microscope test can be used for the detection of semen. The microscopic
detection of sperm cells gives a direct identification of semen. Another advantage of
the microscope test is the possibility to show the presence of semen in a mixture with
blood and vaginal fluid. Unfortunately not all males have sperm cells in their
ejaculate, meaning that the absence of sperm cells does not mean there is no
semen. The absence of sperm cells can be explained by azoospermicity of the donor
due to natural causes or by vasectomy. To be able to identify semen without sperm
cells other techniques have been developed. An example is the use of prostate
specific antigen (PSA), as mentioned in the introduction, for the identification of
semen. Semen of azoospermic males will still contain this protein which is present in
the seminal plasma 6 . PSA can be detected in semen which is 10 6 times diluted 2 . Due
to its extreme sensitivity other body fluids (e.g. breast milk and vaginal fluid) that also
contain PSA but in much lower concentrations, can produce a false positive result. In
addition PSA can give a false positive reaction on male urine 17 .
Semen can also be detected using a lateral flow immunochromatographic strip test.
This test is a part of the Rapid Stain Identification test (RSID TM ) and is developed to
detect the presence of semenogelin. The detection limit of this test is 2.5nL of human
semen, and takes about 10 minutes to give a result. While it gives no positive result
when tested on blood, saliva, vaginal fluid, and urine.
3.3.2 Emerging and future techniques
Like with blood, the newly developed techniques are aiming at identification and are
species specific as well. Most of these new techniques deal with RNA. As mentioned
before Alvarez et al 8 revealed a method to isolate RNA and DNA simultaneously from
the same bloodstain extract. This method can also be applied to semen samples.
Naturally the RT-PCR method described by Juusola and Ballantyne in 2005 also
applies to semen. Juusola and Ballantyne 9 used the semen specific markers PRM1
and protamine 2 (PRM2). Less than 200 pg of input RNA was needed for a positive
result with both genes. The specificity of these genes for semen has been shown,
because they were not detected in saliva, blood, menstrual blood, and vaginal fluid 9 .
Accordingly Fleming and Harbison 11 mention that PRM1 and PRM2 have been found
in ejaculated spermatozoa and dried semen stains, while being absent in blood,
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