PARENT SESSION

(T209) ULTRA-SENSITIVE TIME-RESOLVED FLUORESCENT IMMUNOASSAY OF SEX HORMONE-BINDING GLOBULIN IN HUMAN SPERM.

Selva, David¹, Bassas, Lluis², Munell, Francina³, Tekpetey, Francis⁴, Hammond, Geoffrey¹, ¹ University of British Columbia, Vancouver, BC, Canada² Fundacio Puigverd, Barcelona, Spain³ Hospital Vall d'Hebron, Barcelona, Spain⁴ University of Western Ontario, London, ON, Canada

ABSTRACT- A sex hormone-binding globulin (SHBG) isoform produced by male germ cells has been found in human sperm. Although the majority of SHBG transcripts in the human testis lack exon 7 sequences which result in premature truncation of the SHBG coding sequence, the SHBG isoform in human sperm contains oligosaccharides linked to Asn residues that can only be present within sequences encoded by exons 7 and 8. The SHBG isoform in human sperm is, however, 4-5 kDa smaller than SHBG in human plasma. These data indicate that the sperm SHBG isoform is a result of translation initiation at an ATG codon that corresponds to Met-30 in the mature SHBG polypeptide sequence. To study the physiological and clinical relevance of this SHBG isoform, we have developed a time-resolved immunoassay to measure its concentrations in human sperm, in which washed sperm samples were diluted at 100 million/ml, and 100 ul aliquots were used for measurements. Different methods for SHBG extraction from the sperm were tested, and one freeze and thaw cycle was found to release the SHBG from the acrosome into the assay buffer for analysis. The amount of SHBG detected was directly proportional to the number of the sperm analyzed. The mean intra-assay and inter-assay variability (SEM) was 5.8% and 8.5%, respectively. The SHBG levels in sperm samples taken from donors on two different occasions were within the range of assay variability in 7 out of 9 individuals studied. In samples from potential sperm donors, sperm SHBG levels ranged from 6 - 50 pM/10 million sperm. In these subjects, sperm SHBG levels did not correlate with the serum levels of SHBG. In patient samples, the lowest sperm SHBG levels were found in 3 men requesting a vasectomy (mean ± SD, 9.7 ± 0.9 pM/10 million sperm). By contrast, sperm SHBG levels were highest 3 varicoceole patients (46.24 ± 8.9 pM/10 million sperm). Values in patients with suspected infertility were more variable. Thus, the ultra-sensitive method we have developed for measuring SHBG in human sperm samples is accurate and precise, and should help define the possible function of SHBG in the acrosome of human sperm in relation to male fertility.

KEY WORDS: sperm, Sex Hormone Binding Globulin, acrosome, male infertility