Anti-rat myoglobin antisera in the immunocytochemical diagnosis of rhabdomyosarcomas of rats

Anti-rat myoglobin (Mb) was prepared and used in the avidin-biotin-peroxidase complex (ABC) method on paraffin-embedded sections of nine soft tissue tumors (including two rhabdomyosarcomas) of rats. Distribution and nature of the reactive substance to Mb antiserum were compared to those of desmin antiserum. Rat Mb was isolated from the skeletal muscle; monospecificity of the rat antiserum was confirmed by the immunoblotting procedures. The Mb antiserum reacted specifically to normal and neoplastic striated muscle cells. Mb-staining reactions were present diffusely in the cytoplasm, while desmin-staining substances were localized at Z-bands or were diffuse in the cytoplasm as separated aggregates. Reaction to the Mb serum was also detected in cells of thick portions of Henle's loop and distal convoluted tubules.     

Physico-chemical properties of calf-diarrhea coronavirus

Replication of calf diarrhea coronavirus was not inhibited by 5-iodo-2′-deoxyuridine, indicating that the virus is an RNA virus. Sensitivity to ether and chloroform indicated that the virus is enveloped, and this was confirmed by electron microscopic observation of the virion. The virus was readily inactivated by trypsin and sodium deoxycholate. The virus was labile at 50°C in diluted medium, but readily stabilized in the presence of MgCl₂. It was stable at pH 5 and 7, while a slight loss of infectivity was observed at pH 3. The virus was readily filtered through membrane filters of 200 and 100-nm pore sizes, but not through 50-nm filters. The buoyant density of the virion in CsCl was estimated to be 1.25 g/ml.     

Establishment of a large-scale purification procedure for purified recombinant bovine interferon-tau produced by a silkworm-baculovirus gene expression system

We developed a procedure for the large-scale purification of bovine interferon-tau (boIFN-tau) by means of a silkworm-baculovirus gene expression system. Recombinant boIFN-tau (rboIFN-tau) was efficiently produced in the silkworm infected with boIFN-tau cDNA recombinant baculovirus and accumulated in the haemolymph. To establish a purification method suitable for mass production, we tried three crude purification methods, namely, an acidification and neutralization treatment (ANT), silica gel column chromatography (SGCC), and Blue sepharose column chromatography (BSCC) with a combination of Q-sepharose (QSC) and chelating sepharose column chromatographies (CSCC). As a result, the acidification and neutralization treatment was found to be the most efficient and cost effective. With this combination, we obtained 91% pure products. To confirm the applicability of the procedure for mass production, we inoculated 100 silkworms with the recombinant virus, and recovered about 4.55 mg (1.26 x 10(8)U/mg) of 91% pure rboIFN-tau by means of a combination of the ANT, followed by QSC and CSCC.     

Catalase in manipulation buffer enhances the developmental competence of DNA-injected embryos

To improve the efficiency of transgenesis, we investigated the effects of a radical scavenger during microinjection on the development to blastocysts or pups of mouse pronuclear embryos, microinjected with the enhanced green fluorescent protein (EGFP) transgene. When embryos were microinjected in medium containing 0-1,000 units/ml catalase, the developmental rate to blastocysts was significantly higher (P<0.01) in 100-units/ml catalase (81%) than those in 0 and 1,000 units/ml (56 and 65%). To investigate the ontogenetic ability of DNA-injected embryos, EGFP-injected embryos manipulated under 0 or 100 units/ml catalase were transferred separately to recipient mice. The proportion of fetuses derived from EGFP-injected embryos manipulated under 100 units/ml catalase (29%) was significantly higher (P<0.05) than that manipulated under 0 units/ml catalase (19%). Furthermore, the numbers of transgenic pups were 17 in 100 units/ml catalase and 14 in 0 units/ml catalase. The results of the present study indicate that scavenging reactive oxygen species during in vitro micromanipulation is beneficial for the development of DNA-injected embryos.     

Fate of cystic ovarian follicles and the subsequent fertility of early postpartum dairy cows

The fate of cystic ovarian follicles that developed spontaneously during the early postpartum period of 50 lactating dairy cows was traced by ultrasonography to characterise the follicular dynamics in relation to the fertility of the cows. The absence of postpartum ovulations caused by repeated waves of anovulatory large follicles was also characterised and evaluated. Fifteen of the 50 cows developed cystic follicles, and these follicles became follicular cysts in five of the 15 cows. Most of the cystic follicles emerged before the first postpartum ovulation of the cows. The transition from cystic follicles to follicular cysts delayed the cows' first ovulation, oestrus and insemination, but had less influence on their fertility after they had recovered spontaneously. In addition to the 15 cows that developed cystic follicles or follicular cysts, six of the cows had five to 13 waves of follicles before their first ovulation. These repeated waves of follicles caused a more severe delay in the early postpartum reproductive events but did not affect the cows' fertility.