Semen/Sperm Detection

UV-Biochemistry-Microscopy-Immunolog y

One Day

Semen Stained Sample

$50.00

Each Additional Sample

$35.00

Oral Sex Determination

Y-Chromosome STR analysis

$125

Amylase Activity

$50

DNA Profiling

5-8 Days

Differential lysis and two DNA STR profiles

$250.00

Each additional STR profile for comparison

$100.00

Y-Chromosome STR analysis
(Differential lysis not required)

$125.00

Order form (PDF) To view the order form you will need a copy of Adobe®Acrobat® Reader™. If you have access to the Internet, please click here to download the Adobe Acrobat Reader.  The reader  is a free.

During and after a sexual encounter, bodily secretions may be deposited on a number of items, including clothes, bed sheets, undergarments, discarded bathroom tissues, bath towels, carpet, couch cushions, condoms and feminine products, and can be identifiable for years as dried stains. With an average composition of 200–300 million sperm in 2.5 –3.5 mL of seminal fluid, sperm can be detected in semen stains present on a variety of materials. Sexually active women will often have sperm present in the vagina immediately following sexual intercourse, and sperm can last as long as 3–5 days in the vagina. 

Genetic Identity utilizes six different approaches to confidently answer questions or suspicions related to infidelity.  Every test is completed by a skilled Ph.D. scientist. Each approach in infidelity is designed to answer infidelity questions based upon the sample submitted by using a combination of biochemistry, microscopy, immunology and molecular biology.  

All infidelity test results are held in the strictest confidence; information is only released to the submitting party. Every question is answered with discretion and respect and we have scientists on staff to answer your questions; call us toll free at 866-437-1597.  Alternatively, you can e-mail your question to a staff scientist at

Method

Test

Assay

Explanation

Biochemistry

Prostatic Acid Phosphatase

Presumptive
Possible

This assay is a presumptive test because there are other sources of acid phosphates that can lead to false positive results.

Biochemistry

Amylase

Confirmation

Positive amylase tests confirm amylase activity. Amylase activity is consistent with saliva.

Ultraviolet light

UV Light: 254 and 365 nm

Presumptive
Possible

Many biological fluids will fluoresce under UV light.

Immunology

Anti-p30

Confirmation
Proof

Antibody specific for human prostate specific antigen.

Microscopy

Sperm Visualization

Confirmation
Proof

Visual sperm identification based upon unique morphology.

Differential lysis

DNA Profiling

Confirmation
Proof

Unique DNA profiles generated from the sample.

Y-chromosome

DNA Profiling

Confirmation
Proof

Male specific profile generate from the sample.

Sample Submission: If you have your specimen ready to be tested, place in a paper sack or envelope.  DO NOT use a Ziploc® bag or any other plastic bag.  Download and print the information forms, completely fill them out and mail them with your payment to the address indicated on the forms.
Liquid semen:  Absorb suspected liquid semen onto a clean cotton cloth or Q-tip (swab).   Air-dry the cloth or swab for 10-20 minutes and pack in clean paper or an envelope with sealed corners.  Do not use plastic bags.
Dry semen-stained objects: Submit suspected dry semen-stained objects to the Laboratory (i.e. underwear). Pack to prevent stain removal by abrasive action or packaging materials during shipping.
Semen stains from immovable objects: When possible, cut a large sample of suspected semen stains from immovable objects with a clean sharp instrument.  Alternatively, absorb suspected semen stains onto a clean cotton cloth or Q-tip (swab) moistened with distilled water.  Leave a portion of the cloth or swab unstained as a control. Air-dry the swab or clean cotton cloth and place in clean paper or an envelope with sealed corners.

Information/Order form (PDF) To view the order form you will need a copy of Adobe®Acrobat® Reader™. If you have access to the Internet, please click here to download the Adobe Acrobat Reader. The reader is a free.

Biochemistry

One of the unique properties associated with semen is the presence of an enzyme called prostatic acid phosphatase (PAP). PAP is not a single enzyme but an array of related isoenzymes from a variety of sources. The PAP assay is a well-documented presumptive assay for the presence of semen (1-4). Acid phosphatase activity is 50-1000 times greater in human semen than in any other bodily fluid.  Unfortunately, the use of acid phosphatase as a marker for semen is compromised because the vagina is also a source of vaginal acid phosphatase.  Since seminal and vaginal acid phosphatase can not discriminate, the only approach to differentiating semen in vaginal secretion is by quantitative analysis.  Finding a significantly elevated acid phosphatase level is consistent with the presence of semen.  For example, if semen is present the acid phosphatase assay is very robust and solution will immediately turn a deep purple color.  If the solution does not immediately turn purple or takes several minute to hour to turn color then you are more than likely detecting endogenous vaginal acid phosphatase and not semen.

Principle of enzyme-linked detection.

Ultraviolet light Illumination

Short wave 254 nm/Long wave 365 nm

The use of ultraviolet (UV) light can be of great assistance in many forms of forensic investigation. Since body fluids like semen, saliva, perspiration and vaginal fluids are naturally fluorescent, the use of a UV light source offers a unique method for locating these stains.  The dried bodily fluids will naturally fluoresce when illuminated with UV light source.  We can then isolate the exact location of the stain(s) to test, instead of testing the entire, large pieces of evidence such as a mattress, a carpet, a sheet, an article of clothing, etc.

Immunology-Anti-p30

Unlike the presumptive acid phosphatase test, the detection of the p30 antigen requires the presence of the protein without the need of the target to perform an enzymatic function. This aids in the identification of semen in aged evidence samples in which the acid phosphatase enzyme is functionally inactive. Prostate specific antigen (PSA, also known as p30), is a glycoprotein produced by the prostatic gland and secreted into seminal plasma (fluid).  The p30 is secreted in seminal fluid at concentrations of 200,000 to 5.5 million ng per mL. The sensitivity of the p30 test is 4 ng/mL and therefore seminal fluid diluted up to 1 in a million can also be detected.  This means we can detect semen in samples that have been rinsed or washed (without detergent). In addition, p30 protein is produced from a larger protein that is degraded to release the p30 protein.  Since the p30 is a product of protein degradation it is readily detected in very old samples and sample that have been stored in a plastic bags for a long periond. The detection of the p30 antigen in forensic samples is often helpful because it confirms the presence of semen even in samples that involve vasectomized or azospermic individuals. The reported frequency of azoospermia of 1-9% in seminal stains or swabs examined in sexual assault cases (1) can be expected to rise, since the frequency of contraceptive vasectomy has been estimated to be 750 000 to 1,000 000 per year in the United States (2).

Microscopy

Additionally, individual sperm heads can be accurately identified based on their morphological characteristics, using a phase contrast microscope and 20–100x magnification. An ideal, mature spermatozoon has an oval shaped head with a regular contour (4.0-5.0 mm long and 2.5-3.5 mm wide) with a pale anterior part (acrosome; 40-70% of the head area) and a darker posterior region.  The length-to-width ratio of the head should be 1.50 to 1.75. The sperm tail should be attached in a symmetrically situated fossa in the base of the head.  The base of the head should be broad and not arrow-like. Only one tail should be attached (about 45 mm long), not coiled, nicked or bent over itself. Immediately behind the head the first part of the tail, the mid piece, should be somewhat thicker (maximum width = 1 µm) and about 7-8 mm long.

Oral Sex Determination-Saliva Evidence

Y chromosome:  Since the differential lysis procedure is unsuitable for saliva evidence, the ratio of male to female cells becomes the deciding factor in cases of sexual infidelity. For neat saliva stains or body surface swabs, it can be possible to generate a clean or dominant male type using current STR technology.  For vaginal secretions it becomes more difficult to show the presence of male DNA with in a sea of female DNA.  Y-STR testing can help to verify the presence of male DNA were infidelity involving oral sex is considered.

Amylase activity:  The most common tests for the identification of saliva are dependent upon the detection of amylase (1-3).  Confirmation of amylase means that results are consistent with saliva and then consistent with oral sex. A high level of amylase activity is usually indicative of the presence of saliva.  With the exception of feces, no other body fluid approaches the level of amylase activity in dried stains.  Amylase is a very stable enzyme and can be detected even on dried articles.

Salivary amylase:   As with most enzymes, the function of salivary amylase can be deduced from the name. "Salivary" refers to the fact that it is found in saliva.  The suffix "ase" is usually added to the end of enzymes as an indication that the name refers to an enzyme.  Amylose is a kind of starch in which glucose monomers are joined by α(1—4) linkages. α-Amylase (α-1,4-glucan 4-glucanohydrolase, EC 3.2.1.1) is an enzyme that degrades starch to oligosaccharides and in turn to maltose and glucose by hydrolyzing α-1,4-glucan bonds.  In digestion, the role of α-amylase is primarily the first reaction of this process, generating oligosaccharides that are then hydrolyzed by other enzymes. Salivary amylase is very specific in the way it breaks down starch to glucose.  It inserts a water molecule at the α(1—4) linkage closest to the end of the amylose molecule thus releasing a glucose molecule.  We can measure this break down to determine if salivary amylase is present (Fig 1).  Positive amylase tests confirm amylase activity. Amylase activity is consistent with saliva.

The amylase assay is very sensitive and can detect amylase activity down to a final concentration of 1 x 10-4 U/mL (0.001 units).  Amylase concentration in human saliva is 42–59 U/mL (4).  One unit (U) is defined as the amount of enzyme required to liberate 1 mg of maltose from starch in 3 minutes at 20° C, at pH 6.9.

Figure 1.  Amylase reaction

The mechanism of α-amylase.  The enzyme protonates (hydrolizes) the acetal linkage between the n-sugar residue (sugar on the left) and the rest of the saccharide chain to yield a free glucose residue to convert to energy.

References:
1. Gaensslen, R.E., "Blood, Sweat and Tears and saliva and semen," Law Enforcement Communications, February 1980, pp.23-30.
2. Gaensslen, R.E. et al. Physical Evidence In Criminal Investigation, Narcotic Enforcement Officers Association, Westbrook, CT, 1991, pp.140-143.
3. Keating, S.M. "Oral Sex—a review of it's prevalence and proof, "Journal of the Forensic Science Society, 1988; 28; 341-355.
4. Hoek et al.  �?Toothbrushing affects the protein composition of whole saliva�?  Eur J Oral Sci 2002; 110: 480–481.

                             Molecular Biology-DNA Profiles

Differential Lysis-Extraction

Once a suspected sample has been identified to contain sperm it is often contaminated with other cell types.  The most common contaminating cells are the epithelial cells lining the vaginal wall, but can include epithelial cells from the mouth (buccal cells) and skin, as well as those found in urine.

DNA from contaminating epithelial cells can be removed using a procedure called a differential extraction (5), which takes advantage of unique properties associated with each cell type.  In this procedure, the cells are removed from the suspected material by soaking them in a gentle solution. Epithelial cell DNA is isolated under mild conditions that break open the epithelial cells but leave the sperm cells intact DNA.  The DNA in sperm can then be extracted using a more harsh extraction procedure.

Differential Lysis-DNA Profiles

Differential Lysis and Two Genetic Profiles (Male and Female Genetic Profiles)

$250.00

Differential Lysis and Three Genetic Profiles (Male and Female Genetic Profiles plus a control profile for comparison)

$350.00

Each Additional Profile

$100.00

Sample Submission
If you have your specimen ready to be tested, place in a paper sack or envelope.  DO NOT use a Ziploc® bag or any other plastic bag.  Download and print the information forms, completely fill them out and mail them with your payment to the address indicated on the forms.

Control profile: For the control profile use two clean Q-tips (swab).  Rub one swab on the inside of your left cheek.  Rub the other swab on inside of your right cheek.  Let the sample air-dry for 10-20 minutes. Place the samples in a paper envelope and label it control profile.

Information/Order form (PDF)
To view the order form you will need a copy of Adobe®Acrobat® Reader™. If you have access to the Internet, please
click here to download the file. Adobe Acrobat Reader is a free

References:
1. Brauner, P., "The Evidence Notwithstanding--A case Report on a Rape," Journal of Forensic Sciences, JFSCA, Vol. 37, No.1, Jan. 1992, pp.345-348
2. Brauner, P.& Gallili, N., "A Condom--the Critical Link in a Rape," Journal of Forensic Sciences, JFSCA, Vol. 38, No.5, September 1993, pp.1233-1236.
3. "Forensic Science Symposium On The Analysis of Sexual Assault Evidence", Proceedings, Forensic Science Research and Training Center, Laboratory Division, FBI Academy, Quantico, Virginia, 1983, July 6-8
4. McCloskey, K.L. & Muscillo, G.C. & Noordewier, B., "Prostatic Acid Phosphatase Activity in the Postcoital Vagina," Journal of Forensic Sciences, 1975, vol.20, p.630-636
5. Gill, P., Jeffreys, A.J., Werret, D.J., Nature 1985, 318, 577-579

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