Non-surgical recovery of eggs from single-ovulating bovines

Egg recovery was attempted by non-surgical technique 128 times from 38 single-ovulating cows and heifers 7 or 8 days after insemination. Ninety-one (71%) eggs were recovered of which 81 (89%) were fertilized.     

Non-surgical transfer of deep-frozen bovine embryos

Preservation of cattle embryos by methods of deep-freezing has recently been established (1, 11, 12) and provides a valuable addition to the possibilities of controlled breeding by embryo transfer in cattle.Already long distance transport of frozen embryos has been demonstrated (2, 6) and adopted by some commercial interests. However, in all publications to date, embryos have been transferred to recipients by surgical methods, even though non-surgical methods of embryo recovery and transfer would be preferred for commercial embryo transfer.The purpose of the present experiments was to utilize non-surgical methods of embryo recovery and transfer of deep-frozen cattle embryos to demonstrate the feasibility of the procedure for a farm service to interested breeders. The particular advantage of non-surgical embryo transfer methods is that neither the donor nor recipient need to leave the farm. Embryo preservation by freezing obviates the necessity for synchronization of recipients for immediate transfer from the donor and allows considerable freedom in the choice of the recipients and the timing of embryo transfers.     

A new technique for the recovery of Toxocara eggs from soil.

A new flotation technique for the extraction of Toxocara eggs from soil has been devised. It has a fairly high recovery efficiency for Toxocara eggs, can be used for replicated quantitative experiments, and may also be used for recovery of other ascarid eggs from soil.     

Patterns of intracellular Ca2+ concentrations in fertilized bovine eggs.

Sperm-induced calcium (Ca2+) changes were examined in zona pellucida-intact, mature bovine eggs injected with the fluorescent Ca2+ indicator fura-2 dextran (fura-2 D). Fifty four percent (37/68) of the dye-injected, inseminated bovine eggs were fertilized and 43% (16/37) of the fertilized eggs exhibited Ca2+ elevations during the time of measurement. All (16/16) of the eggs with Ca2+ elevations were fertilized but none of the unfertilized eggs (0/31) showed intracellular Ca2+ elevations. Six of 13 eggs that were later examined and found to be fertilized at the time of the Ca2+ recordings did not show sperm-induced Ca2+ elevations. Fifty percent (8/16) of the eggs with Ca2+ elevations exhibited a single Ca2+ rise as a response to sperm penetration during the 60-min period in which these eggs were monitored. Twelve percent (2/16) of the eggs responded with two Ca2+ elevations spaced by 50- and 51-min intervals and 38% (6/16) of the eggs exhibited multiple elevations with intervals of 15-29 min. In the latter group, one egg was polyspermic. The mean amplitude of the sperm-induced Ca2+ elevations was 564 +/- 58 nM. Eggs with single elevations reached higher peak concentrations than eggs with multiple elevations (p < 0.05). The mean duration of the Ca2+ elevations was 166 +/- 13 sec and was similar among eggs with different Ca2+ patterns. The first elevations detected occurred at a mean of 6.6 +/- 0.5 h after insemination. Fertilization in this study was confirmed by looking at pronuclear formation 16 h post-insemination or by DNA staining immediately after the fluorescence readings. Eggs exhibiting Ca2+ elevations ranged in stage of fertilization from just penetrated to pronuclear. Injection of inositol 1,4,5 trisphosphate (5 microM in the injection pipette) into 6 bovine eggs induced an immediate Ca2+ elevation with a mean peak Ca2+ value of 700 +/- 60 nM and a mean duration of 103 +/- 21 sec. Incubation of bovine eggs with 200 microM thimerosal induced periodic Ca2+ rises. The mean number of Ca2+ elevations observed in 35 min of recordings was 3.0 +/- 0.5 (n = 9, range 1-5). The mean peak Ca2+ value of the first thimerosal-induced Ca2+ elevation was 990 +/- 210 nM. The results of this study indicate that fertilization can evoke intracellular Ca2+ elevations in bovine eggs and that the periodicity of these Ca2+ elevations is different among eggs. Furthermore, both inositol 1,4,5-trisphosphate and thimerosal were able to induce intracellular Ca2+ release in bovine eggs.     

Transvaginal recovery of uterine eggs in the cow

The uterus was flushed with a transvaginal technique 5 to 6 days after isemination in 6 naturally ovulated and 6 superovulated cows. When 50–100 of fluid were injected, an average of 82% of the volume was recovered, jgs were recovered in 3 of the 6 cows having a single ovulation and 5 of ie superovulated animals. The recovery rate was 43% of the number of corpora Jtea (20/46).     

Comparative recovery of uninjured and heat-injured Listeria monocytogenes cells from bovine milk.

The standard selective enrichment protocols of the Food and Drug Administration (FDA) and U.S. Department of Agriculture (USDA) were compared with an experimental nonselective broth enrichment (NSB) protocol and variations of the standard cold-enrichment (CE) protocol for the recovery of heat-injured Listeria monocytogenes. Bacterial cells (10(7)/ml) were suspended in sterile milk and heated at 71.7 degrees C in a slug-flow heat exchanger for holding times ranging from 1 to 30 s. Surviving cells were determined (50% endpoint) by the given protocols, and the following D values were obtained: NSB, D = 2.0 +/- 0.5 s; FDA, D = 1.4 +/- 0.3 s; USDA, D = 0.6 +/- 0.2 s; CE, D less than or equal to 1.2 s. The respective direct-plating media used in these enrichments were also analyzed for recovery, and the following D values were calculated from the enumeration of surviving cells; NSB, D = 2.7 +/- 0.8 s; FDA, D = 1.3 +/- 0.4 s; USDA, D = 0.7 +/- 0.2 s. The low levels of heat-injured L. monocytogenes cells which were detected at inactivation endpoints on the optimal nonselective media (25 degrees C for 7 days) failed to recover and multiply during experimental CEs (4 degrees C for 28 days). Initial inactivation experiments in which raw whole milk was used as the heating menstruum gave much lower recoveries with all protocols. The detectable limits for uninjured cells that were suspended in raw milk were similar (0.35 to 3.2 cells per ml) for the standard CE, FDA, and USDA protocols.(ABSTRACT TRUNCATED AT 250 WORDS)     

Transient recovery from trypsin-caused reduction in fertilizability of sea urchin eggs

Summary The effect of trypsin on the fertilizability of sea urchin eggs was studied withParacentrotus lividus andPseudocentrotus depressus. The main effects were two reductions of fertilizability, with a transient increase intervening. The first decrease was probably caused by degradation of sperm-binding sites at the vitelline sheet and the second by degradation of binding sites on the plasma membrane.     

Non-surgical uterine flushing for the recovery of preimplantation embryos in rhesus monkeys: Lack of seasonal infertility

A non-surgical uterine flushing technique was employed to recover rhesus monkey preimplantation embryos during April–-September, a period thought to be associated with reduced fertility. A total of 22 females of proven fertility, maintained indoors under strict light and temperature control, were employed for the study in which 72 menstrual cycles were monitored. The average length of their menstrual cycle was 27.9 ± 3.8 days. The percentages of cycles that showed normal cyclic patterns of estrogen, LH, and sex-skin color were 84.7%, 91.7%, and 90.3%, respectively. Following natural mating, uterine flushing was performed on days 4–7 of pregnancy. Of 58 attempts, 27 (46.6%) resulted in the recovery of embryonic materials. Two recoveries produced unfertilized oocytes; 20 yielded embryos which were morphologically normal, and 7 yielded damaged and/or degenerate embryos. Luteal function in cycles involving uterine flushings was evaluated by examining ovaries by laparoscopy, ovulation being confirmed in 34 of 35 examinations. Serum progesterone was > 2 ng/ml in 39 of 45 cycles. The conception rate of 46.6% noted in this study was similar (P > 0.4) to those observed from other natural matings in our colony, either during the same period (49.3%; n = 67) or during October–-March (46.9%; n = 113). These results show that rhesus monkeys, maintained under appropriate environmental conditions, can experience normal fertile menstrual cycles throughout the summer months, and extend previous observations to demonstrate, for the first time, that preimplantation embryos can be recovered from this species year-round. © 1993 Wiley-Liss, Inc.     

Characterization of the zona pellucida glycoproteins from bovine ovarian and fertilized eggs

Bovine zone pellucida (ZP) glycorproteins from ovarian egg emerged as three bands with molecular mass of 78 kDa, 64 kDa and 21 kDa in SDS-PAGE under reducing conditions. Endo-β-galactosidase (EβG) digestion of the glycoproteins yielded five products with molecular mass of 76 kDa (EβG-76), 68 kDa (EβG-68), 63 kDa(EβG-63), 47 kDa (EβG-47) and 21 kDa (EβG-21) under the same conditions. The N-terminal amino acid sequences of EβG-76 and EβG-21 were identical. This fact together with the results of diagonal SDS-PAGE indicated that EβG-21 (N-terminal region) is linked to EβG-63 (C-terminal region) through disulfide bond to form EβG-76. Immunoblot analysis using anti-pig ZP protein antibodies revealed that bovine EβG-76, EβG-68 and EβG-47 correspond to pig PZP2, PZP3α and PZPEβ glycoproteins, respectively. The EβG-76 and EβG-68 components were shown to be specifically cleaved during fertilization.     

Non-surgical management of peripheral vascular disease: a review.

Although the outlook for a pronounced improvement in the ischaemic limb that cannot be surgically treated remains bleak, cessation of smoking, encouragement of exercise, and the withdrawal of vasoconstricting agents may give some symptomatic relief. There is little evidence that vasodilators or antiplatelet agents have much to offer.     

Optimization of bovine serum albumin sorption and recovery by hydrogels

Aqueous two-phase systems are composed of aqueous solutions of either two water-soluble polymers, usually polyethylene glycol (PEG) and dextran (Dx), or a polymer and a salt, usually PEG and phosphate or sulfate. Partitioning of proteins in such systems provides a powerful method for separating and purifying mixtures of biomolecules by extraction. If one of the phase forming polymers is a crosslinked gel, then the solution-controlled gel sorption may be considered as a modification of aqueous two-phase extraction. Since PEG/dextran systems are widely used in aqueous two-phase extraction and dextran gels (Sephadex) are common chromatographic media, we choose a PEG/dextran gel system as a model system in this study. The partitioning behavior of pure bovine serum albumin (BSA) in PEG/dextran gel systems is investigated to see the effects of variations in PEG and NaCl concentrations on the partition coefficient K. By making use of the Box-Wilson experimental design, K is shown to be maximized at 9.8 (%, w/w) PEG and 0.2 M NaCl concentrations, respectively, as 182.