A simple direct-addition microtitre plate enzymeimmunoassay (EIA) for progesterone in whole milk is described. The assay used antiserum raised against 11 alpha-hydroxyprogesterone 11-hemisuccinate (progesterone 11-hemisuccinate) and a heterologous label prepared by conjugation of 11 alpha-hydroxyprogesterone 11-glucuronide (progesterone 11-glucuronide) with alkaline phosphatase using an active ester procedure. The sensitivity, analytical recovery, linearity of response and precision of the assay compared favourably with radioimmunoassay (RIA). Results from EIA of milk samples were compared with determinations made after isolation of progesterone by HPLC (r = 0.910). Milk samples (200) were assayed by RIA at both the Milk Marketing Board and the Cattle Breeding Centre and the results were correlated with EIA performed at the Cattle Breeding Centre (r = 0.890 and r = 0.833 respectively). Calving data were obtained from a further 110 cows for which the milk progesterone EIA had provided a pregnancy test 24 days after AI; 46 cows were correctly identified as non-pregnant and 58 as pregnant and there were 4 false positive and 2 inconclusive results.
Peripheral blood mononuclear cells were collected from a sheep immunized against progesterone-11 alpha-hemisuccinate-ovalbumin. Following fusion with NS1 mouse myeloma or heteromyeloma cells, a large number of hybrid colonies was established. These were screened for the production of sheep antibodies to progesterone. Twenty-four cell lines were cloned and one was stabilized. This cell line, O/MP.1A9.D7B2, produced a high-affinity ovine immunoglobulin G1 (dissociation constant 4.8 pmol/l) with a high degree of specificity for progesterone. The antibody was substituted into a competitive enzyme-linked immunosorbent assay for the measurement of progesterone in bovine milk, originally established using an ovine polyclonal antibody, and the results were compared. The monoclonal antibody produced an assay with a lower limit of detection and a greater degree of discrimination than the polyclonal antiserum.
Abstract— In piglets affected with congenital tremor type AII the CNS was not morphologically underdeveloped; spinal cord weight, total DNA content and fat‐free dry matter differed little from control values. However the total lipid extractable from affected spinal cords was only about 63% of values established for normal newborn piglets. In particular, the cerebroside content (a myelin‐specific lipid) was reduced to about 30% of the ‘normal’ value. This was parallelled by the results of an in vitro assay of cerebroside synthesis from [3H]galactose which indicated a metabolic impairment. The altered fatty acid profile of isolatcd cerebrosides further suggested a derangement of fatty acid synthesis. Unlike the spinal cords of normal newborn piglets, tissues from affected piglets contained significant amounts of cholesterol esters carrying the characteristic fatty acids associated with demyelination. This implied that the reduced quantities of possibly abnormal myelin were unstable. Abnormal swollen oligodendrocytes were commonly present in the spinal cords of affected piglets and this was consistcnt with the observed impairment of myelin formation.
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