Application of enzymeimmunoassay to measure oestrone sulphate concentrations in cows' milk during pregnancy

• Published in: The Journal of steroid biochemistry and molecular biology Vol. 50; no. 3; pp. 189 - 196
• Main Authors: Henderson, K.M., Camberis, M., Simmons, M.H., Starrs, W.J., Hardie, A.H.M.
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 01.08.1994; Elsevier Science
• Subjects: Animals; Biological and medical sciences; Cattle; Cross Reactions; Estrone - analogs & derivatives; Estrone - analysis; Female; Fundamental and applied biological sciences. Psychology; Gestational Age; Hormone metabolism and regulation; Immunoenzyme Techniques; Milk - chemistry; Pregnancy; Pregnancy, Animal; Pregnancy. Parturition. Lactation; Vertebrates: reproduction; Pregnancy; Vertebrata; Mammalia; Steroid hormone; Estrogen; Ox; Artiodactyla; Measurement method; Enzyme immunoassay; Ungulata; Breast milk
• ISSN: 0960-0760; 1879-1220
• DOI: 10.1016/0960-0760(94)90028-0

The characteristics of antigen- and antibody-coated enzymeimmunoassay (EIA) formats to measure oestrone sulphate (OS) were studied using a murine monoclonal antibody as the primary antibody. In an antigen-coated format the most sensitive EIA (9 fmol/well) was achieved using 6-ketoestrone-6- O-carboxymethyloxime (OCMO) coupled to bovine serum albumin (BSA), as the coating antigen, and horseradish peroxidase (HRP), as the enzyme label. In an antibody-coated format, comparable sensitivity could be achieved using HRP conjugated to either OCMO, oestrone-3-glucuronide (OG) or oestrone-3-hemisuccinate (OHS) as the steroid ‘tracer’. In both the antigen- and antibody-coated formats replacing HRP with alkaline phosphatase (AP) markedly aggravated the assay sensitivity. The antigen-coated EIA format was used to measure OS concentrations in cows' milk directly without an initial defatting step, and a progressive increase in OS concentrations in milk as pregnancy progressed was observed. Median OS concentrations rose from 1.1 nmol/1 at days 70–99 of pregnancy ( n = 44) to 3.2 nmol/1 at days 140–160 ( n = 92). Oestrone sulphate concentrations in milk from non-pregnant cows ( n = 51) ranged from non-detectable to 1.3 nmol/1 with a median value of 0.4 nmol/1. Only 5% of cows 120 or more days pregnant had milk OS concentrations within the range of values found in milk from non-pregnant cows. Accurate discrimination of non-pregnant and pregnant cows can thus be achieved on the basis of OS concentrations in milk samples taken at least 120 days after mating/insemination. This EIA for OS may have a role in the dairy industry as an alternative non-invasive means of determining pregnancy status in cows.