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R. G. Lea and A. E. Bolton

Summary. Two-dimensional crossed immunoelectrophoresis of sera from pregnant and non-pregnant horses, using antisera developed against early pregnant mare serum, revealed the presence of two immunologically related proteins one of which appeared to be specific to the pregnant state. This pregnancy-specific protein had β2-electrophoretic mobility and was first detectable at Day 6 after successful mating with a stallion. The second protein had γ2-electrophoretic mobility and was present in sera from pregnant and non-pregnant horses. The proteins were termed β2-horse pregnancy protein and γ2-horse protein respectively. The latter appeared to be immunologically related to the former in that the precipitin lines of the 2 proteins showed continuity. Samples from 16 mares mated with a stallion were investigated for the β2-protein during the first 3 weeks after mating. Of the 11 successful matings, confirmed by ultrasonic scanning at 90 days or by a successful outcome, 10 mares showed the presence of the protein. In all of 14 non-pregnant sera taken from mares not recently mated, the protein was not detectable. The validity of detection of β2-protein as an indication of pregnancy was clinically significant at the 10% level. The presence of the protein in 2 out of the 5 recently mated mares that did not become pregnant may be indicative of a biochemical pregnancy that failed at a later stage of gestation.

Keywords: horse; pregnancy protein; immunochemical techniques

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E. Kessopoulou, M. J. Tomlinson, C. L. R. Barratt, A. E. Bolton, and I. D. Cooke

Summary. Peroxidative damage induced by reactive oxygen species (ROS) has been proposed as one of the major causes of defective sperm function. In previous studies of the production of ROS in semen, the contribution of contaminating leucocytes was not assessed. We determined the levels of ROS in 60 semen samples from men attending our infertility clinic and demonstrated by performing extraction experiments with antibody-coated magnetic beads that, within this unselected population of patients, leucocytes were the major source of ROS in the low-density Percoll fraction. Of the sperm motion parameters examined using computerized semen analysis, beat-cross frequency was the only one significantly affected by the ROS in semen.

Keywords: antibody-coated magnetic beads; computerized sperm motility analysis; leucocytes; reactive oxygen species; spermatozoa; human

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R. G. Sutcliffe, A. E. Bolton, F. Sharp, L. V. B. Nicholson, and R. MacKinnon

Summary. Human alpha uterine protein (AUP) has been prepared from extracts of decidua by antibody affinity chromatography, DEAE Sepharose chromatography and by filtration through Sephadex G-150. This procedure yielded a protein fraction containing AUP, which was labelled with125I by chloramine T. When analysed by SDS gel electrophoresis this radioiodinated protein fraction was found to contain predominantly a single species of protein which was precipitated by antibodies against AUP in antibody–antigen crossed electrophoresis. Rabbit anti-AUP precipitated 55–65% of the tracer in a double-antibody system. Sephadex G150 gel filtration of AUP obtained before and after affinity chromatography provided a molecular weight estimate of 50 000. Since SDS gel electrophoresis revealed a polypeptide molecular weight of 23 000–25 000, it is suggested that AUP is a dimer.

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O. M. R. Westwood, M. G. Chapman, N. Totty, R. Philp, A. E. Bolton, and N. R. Lazarus

Summary. Human placental protein 14 (PP14) has been purified in high yield from first trimester decidual cytosol. High-performance liquid chromatography on anion exchange, gel filtration and reverse-phase chromatography were used. The protein obtained is approximately 97% pure with an overall recovery of about 50% from the original tissue extract. The first 24 amino acids of the N-terminal were found to be Met-Asp-Ile-Pro-Gln-Thr-Lys-Gln-Asp-Leu-Glu-Leu-Pro-Lys-Leu-Ala-Gly-Thr-Glu-His-Glu-Met-Ala-Met. PP14 has been characterized in this study to be a dimeric glycoprotein of M r 60 000, with homologous subunits having an M r of 28 000.

Keywords: human; PP14; decidual cytosol; h.p.l.c. techniques