Letters to the editor |
Reply: Development of a novel home sperm test – What are the limitations?
1 Andrology Centre, Department of Medicine, Karolinska University Hospital and Institute, Huddinge, Stockholm, Sweden 2 Assisted Conception Unit, Birmingham Women’s Hospital and 3 Reproductive Biology and Genetics Research Group, Birmingham
4 To whom correspondence should be addressed at: Division of Reproductive and Child Health, The Medical School, University of Birmingham, Birmingham, B15 2TT, UK. E-mail: c.barratt{at}bham.ac.uk
Sir,
We appreciate the comments from Dr Alvarez and Dr Agarwal and welcome the opportunity to address them.
Regarding point 1
- We clearly stated in the Material and Methods the selection criteria for the study (see Björndahl et al., 2006
). We stated that we intended that a target minimum of 25% of subjects should have progressively motile sperm concentrations below the test threshold (10 million progressively motile cells/ml semen). The selected research donors had a variety of semen characteristics—some were recruited specifically because they had poor semen quality—thus the authors statement ‘88% of semen samples were not obtained from normal donors’ is incorrect.
- High viscosity and incomplete liquefaction are different issues (see Mortimer, 1994; WHO, 1999), and it is inappropriate to combine them. In the article, we stated that all samples were allowed to liquefy for 30 min at 20°C. This was set to try and mimic the home environment. Increased viscosity is not necessarily associated with impaired fertility potential, but even if it did reduce the ability of sperm to migrate from the semen into the hyaluronate, then this biological effect would be detected by the device.
- Incomplete liquefaction and non-liquefaction are different issues, but both are considered diagnostic findings that require medical assessment by a properly trained clinical andrologist. Should the liquefaction be so incomplete as to impair sperm penetration into hyaluronate, this is diagnostic (no red line would appear) and the patient with a problem will seek medical advice. Non-liquefaction is considered a pathological finding, and hence would be detected by the device (no red line—sufficient number of sperm unable to penetrate into the hyaluronate).
Regarding point 2
There is no ‘sample port’ in the device. We clearly stated in the article how the test works (see Figure 1 in the article and associated text). In our experiments, the semen samples were ejaculated directly into containers and after 30 min loaded into the device. When the man is using this at home, he ejaculates directly into the sample collection container, which is integral to the device—no transfer of semen/sperm is necessary (see http://www.fertell.co.uk for patient instructions and photographs). The man is instructed that after 30 min when semen has filled the sample well, he needs to put the top on the collection container and push a button (blue one in Figure 1 in the article). When this occurs, a portion of the semen sample (600 µl) comes in direct contact with the hyaluronate (via a mesh). Only if the volume is less than the critical limit (600 µl) will the test fail, i.e. no red line will appear. Men are advised to abstain for at least 2 days (not greater than 7 days) before doing the test. If they have a semen volume of 600 µl or less, this is a diagnostic finding considered to have clinical importance.
Regarding point 3
Once again the authors are confusing increased viscosity with incomplete liquefaction (see above). The interface between the gel and semen is deliberately large to allow more sperm to successfully penetrate the hyaluronate (see Tang et al., 1999). This may compensate for potential heterogeneity in the semen sample. In our trials, we obtained a high degree of accuracy (>95% compared with the Hamilton Thorn and modified Kremer testing). Thus, we do not believe that sample heterogeneity is a significant issue when using this device.
Regarding point 4
CD59 was used because it is well known to be present on sperm (and within the cell), as stated in our article (see review Harris et al., 2006). Studies performed by other authors have shown that CD59 is present on the vast majority of sperm (in semen or in a swim-up, see D’Cruz and Hass, 1992; Fenichel et al., 1994; Simpson and Holmes, 1994; approximately >95% in our experiments). We have not specifically examined the relationship between individual sperm morphology and sperm CD59 as, in the samples we examined, no obvious relationship exists.
Our device is based on sperm penetrating hyaluronate. Many authors have studied the relationship between sperm penetration into cervical mucus (and its substitutes) and semen characteristics (e.g. Mortimer et al., 1990; Aitken et al., 1992). Penetration is highly dependent on sperm motility. As there is a strong relationship between motility and morphology, men with severe teratozoospermia will have poor penetration. Cases with isolated teratozoospermia are rare (e.g. Mortimer et al., 1982).
We do not believe that it is appropriate in an internationally peer-reviewed journal to discuss the merits of different assays. For the authors’ information, Fertell is cleared by the FDA for OTC use (K041039).
Finally, we do not understand the need to use the device for washed sperm samples. The authors have misunderstood the fundamental principle of the device and its purpose (as the title of the article stated—it is a home test). From a biological perspective, sperm would not migrate in the same way (and with very much reduced numbers) from washed preparations in culture medium (even if in non-capacitating medium) into the HA gel (Katz et al., 1990).
Commercial interest
As stated on the original article, CLRB is a member of the Scientific Advisory Board of Genosis. Genosis provided a grant to the University of Birmingham to fund the development of the home sperm test.
References
Aitken RJ, Bowie H, Buckingham D, Harkiss D, Richardson DW, West KM. (1992) Sperm penetration into a hyaluronic acid polymer as a means of monitoring functional competence. J Androl 13:44–54.
Björndahl L, Rattle S, Hart G, Kirkman-Brown JC, Barratt CLR. (2006) Development of a home sperm test. Hum Reprod 21:145–149.
D’Cruz OJ and Hass GG Jr. (1992) Flow cytometric quantitation of the expression of membrane cofactor protein as a marker for the human sperm acrosome reaction. Fertil Steril 58:633–636.[Web of Science][Medline]
Fenichel P, Cervoni F, Hofmann P, Deckert M, Emiliozzi C, His BL, Rossi B. (1994) Expression of complement regulatory protein CD59 on human spermatozoa: Characterisation and role in gametic interaction. Mol Reprod Dev 38:338–346.[CrossRef][Web of Science][Medline]
Harris CL, Mizuno M, Morgan PB. (2006) Complement and complement regulators in the male reproductive system. Mol Immunol 43:57–67.[CrossRef][Web of Science][Medline]
Katz DF, Morales P, Samuels SJ, Overstreet JW. (1990) Mechanisms of filtration of morphologically abnormal sperm by cervical mucus. Fertil Steril 54:513–516.[Web of Science][Medline]
Mortimer D. (1994) Practical Laboratory Andrology(Oxford University Press, Oxford).
Mortimer D, Templeton AA, Lenton EA, Coleman RA. (1982) Semen analysis parameters and their interrelationships in suspected infertile men. Arch Androl 8:165–171.[Web of Science][Medline]
Mortimer D, Mortimer ST, Shu MA, Swart R. (1990) A simplified approach to sperm–cervical mucus interaction testing using a hyaluronate migration test. Hum Reprod 5:835–841.
Simpson KL and Holmes CH. (1994) Differential expression of complement regulatory proteins decay-accelerating factor (CD55), membrane cofactor protein (CD46) and CD59 during human spermatogenesis. Immunology 81:452–461.[Web of Science][Medline]
Tang S, Garrett C, Baker HWG. (1999) Comparison of human cervical mucus and artificial sperm penetration media. Hum Reprod 14:2812–2817.
WHO. (1999) WHO laboratory manual for the examination of human semen and sperm–cervical mucus interaction 4th edn (Cambridge University Press, Cambridge) World Health Organization.
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