SPERM HY-LITER™ – FAQs
Q:Can DNA profiles be obtained from SH stained slides?
A: Easily. SPERM HY-LITER is an immunofluorescent detection system for human sperm and the reagents do not react, inhibit or interact with DNA in any way that interferes or inhibits standard DNA extraction or PCR amplification protocols. Numerous laboratories have successfully obtained full DNA profiles, both somatic and sex chromosome based, from SH-stained slides. In addition protocols are available to perform in situ digestion of stained slides in order to obtain DNA profiles.
Q: How does SPERM HY-LITER stain ‘hidden’ sperm?
A: The protocol has been specifically designed to allow the Alexa488 tagged monoclonal antibody to reach sperm at all layers of the stained slide. As fluorescent detection is not inhibited by cell layers, sperm under cells, over cells, and at all layers of the preparation are easily identified.
Q: Does the SPERM HY-LITER staining protocol damage sperm or epithelial cells?
A: The staining procedure is simple, and extremely reproducible and works particularly well on highly dense cell preparations. Cells are not damaged in any way, and no morphology, if it is originally present, is lost during the staining method. If tails, midsection and acrosomes are present at the beginning of the staining, they will be present at the end – however the antibody targets the sperm head, the sole remaining sperm substructure in sexual assault evidence, uniquely.
Q: Does SPERM HY-LITER have species specificity?
A: SPERM HY-LITER uniquely and specifically stains human sperm heads – neither domestic, farm , or higher primate sperm are labeled by the kit.
Q: Does SPERM HY-LITER have body fluid specificity?
A: Only human sperm heads react – no other cell type and no other body fluid is stained by the kit.
Q: What reagents does the laboratory have to provide?
A: The forensic laboratory must have 1M DTT (ph 8.0, recipe of course supplied), and mol bio quality water. All other components are supplied in the kit. The final concentration of DTT used in the Sample Prep solution is ~12.5 mM.
Q: Can SPERM HY-LITER be used on smear slides often included in sexual assault evidence collection kits?
A: SPERM HY-LITER is a very effective and efficient staining method for smear slides and extracts. In fact the more dense the cell preparation, the greater the need for SPERM HY-LITER – which performs particularly well in dense cell preparations.
Q: What equipment does the laboratory have to have in order to visualize SPERM HY-LITER stained slides?
A: The laboratory must have a fluorescent microscope fitted with the correct light source (there are many suitable models and manufacturers) and the correct filters (called ‘cubes’). We strongly recommend the addition of phase contrast as an optical staining method to the fluorescent microscope, but it is not strictly required.
Q: How long does the fluorescence last?
A: There are two fluorescent dyes in the SPERM HY-LITER staining method – a non-specific nuclear dye, DAPI, that is somewhat analogous to KPIC in that all cells are non-specifically stained (epi, sperm, fungi, spores, pollen, bacteria, etc) and the Alexa488. The dyes themselves are quite stable – at least 8-10 years when stored in the dark. The slide preparation will reduce this lifetime – by making the material on the slide accessible for DNA analysis, should this be required, the slides are mounted with a water soluble mounting media that contains anti-fade reagents. Current experiments demonstrate that slides are stable for at least two years (as this is ongoing research, we anticipate longer shelf life).
Q: What is contained in a SPERM HY-LITER microscope package?
A: The SPERM HY-LITER microscope package has been extensively optimized specifically for crime laboratories and includes a modern research grade fluorescent microscope with the most appropriate light sources, objectives, filters, and optical components. In addition, the package includes a state-of-the-art image capture, archiving and analysis software package with a color CCD camera. This allows permanent, digital signed storage of all sperm searches, customized reports, and full documentation of results as required by ISO-17025 audit requirements. The package also includes installation and training for forensic laboratory staff on all aspects of the microscope and staining.
Q: What are the benefits of SPERM HY-LITER screening for sperm from sexual assault evidence?
A: There are several answers to this question: tremendous time savings in identifying sperm (up to 80%), greater DNA-STR profile success from SAE as DNA profile success will now correlate with observed sperm cell counts, court defensible results on the identification of sperm and the identification of sperm can now be scientifically and procedurally justified.
Q: Are other laboratories using SPERM HY-LITER in case work?
A: Yes. Please contact Ms Dina Mattes for a current list of laboratories that have purchased systems, validated systems and released the method for case work. This list is constantly expanding.
Q: Are there publications describing SPERM HY-LITER?
A: Yes, several and more in press. Please contact Ms Dina Mattes for a current list of abstracts and publications describing various applications of the technique.
Q: Can SPERM HY-LITER slides be stained in ‘batch’ mode?
A: Absolutely – numerous slides can be stained all at once – this is in fact a very efficient method of staining slides with SPERM HY-LITER.
Q: Is SPERM HY-LITER compatible with automation, or automated slide scanning methods?
A: Yes, in fact, SPERM HY-LITER is the only method currently available that provides sufficient signal to noise for software-driven sperm identification – methods that rely on KPIC stained preparations cannot distinguish sperm from background signal in case work samples.
There is also an effective and efficient liquid handling robot that can stain up to 100 SPERM HY-LITER slides a day with minimal analyst intervention.
Q: What is the shelf life of a SPERM HY-LITER kit?
A: Currently SPERM HY-LITER kits have a six (6) month expiration date.
Q: Are SPERM HY-LITER slides available in different configurations?
A: Yes, the standard SPERM HY-LITER slide has two eleven millimeter wells, but a variety of other sizes and configurations are available as well. Please consult the current SPERM HY-LITER catalog.