Primer-induced labeling of pea and field bean chromosomes in situ and in suspension

A protocol for primed in situ DNA labeling (PRINS) was optimized for pea (Pisum sativum L.) and field bean (Vicia faba L.) chromosomes attached to coverslips. Cloned DNA or synthetic oligonucleotides were used as probes for repetitive DNA sequences (rDNA, Fok-element) and different reaction conditions were tested to achieve the highest specific signal-to-background ratio. A procedure based on direct labeling by fluorescein-dUTP was compared with an indirect one using digoxigenin detected by fluorescently labeled antibody. Under optimal conditions, strong and specific signals were obtained exclusively on chromosome regions known to contain respective DNA sequences. Compared to the direct labeling, significantly stronger signals were obtained when the indirect procedure was used. Both types of labeling were successfully applied to chromosomes in suspension and were shown to produce signals comparable to that obtained with chromosomes attached to coverslips. It is expected that primed in situ DNA labeling en suspension (PRINSES) will provide a basis for flow-cytometric discrimination and sorting of otherwise indistinguishable chromosomes according to their specific fluorescent labeling.     

Epitope specificity of CD44 for monoclonal antibody-dependent facilitation of marrow engraftment in a canine model

Primary graft rejection after marrow transplantation occurs more frequently in patients receiving HLA-haploidentical compared with HLA-identical sibling transplants. Both human and experimental animal data suggest that the cells responsible for this phenomenon are either host natural killer (NK) cells, T cells, or both. To investigate the mechanisms of graft rejection, we have developed a canine model of marrow transplantation, which uses DLA-nonidentical unrelated donors in the absence of postgrafting immunosuppression. In this model most animals rejected their marrow grafts after a preparative regimen of 9.2 Gy total body irradiation (TBI). However, engraftment of DLA-nonidentical marrow can be facilitated when the recipients are pretreated with monoclonal antibody (MoAb) S5, which recognizes CD44. In this report, we extended these observations by first cloning the canine CD44 and, next, mapping the epitope recognized by S5, which was located in a region conserved among human and canine CD44 and was distinct from the hyaluronan binding domain. However, in vitro binding of S5 caused a conformational change in CD44, which allowed increased hyaluronan binding. Then, we reexamined the in vivo model of marrow transplantation and compared results with MoAb S5 to those with two other anti-CD44 MoAbs, IM7 and S3. Only MoAb S5 significantly increased the engraftment rate of DLA-nonidentical unrelated marrow, whereas the two other anti-CD44 MoAbs were ineffective. The enhanced in vivo effect was not related to differences in the MoAbs' avidities, since both S5 and IM7 had equivalent binding to CD44, but most likely related to the specific epitope that S5 recognizes. Thus, this study shows that the effect of the anti-CD44 MoAb S5 in facilitating engraftment is epitope specific and if one is to use an anti-CD44 to facilitate engraftment of marrow in humans, one cannot assume that any anti-CD44 would work.     

Temperature and modulation characteristics of resonant-cavitylight-emitting diodes

Resonant-cavity light-emitting diodes (RCLED) are novel, high-efficiency light-emitting diodes which employ optical microcavities. These diodes have higher intensities and higher spectral purity as compared to conventional LEDs. Analytical formulas are derived for the enhancement of the spontaneous emission along the optical axis of the cavity. The design rules for high-efficiency operation of RCLEDs are established. The temperature dependence of the emission intensity is analyzed in the range 20-80° and it is described by an exponential dependence with a characteristic temperature of 112 K. The modulation characteristics of RCLEDs exhibit 3 dB frequencies of 580 MHz. Eye diagrams at transmission rates of 622 Mb/s are wide open indicating the suitability of RCLEDs for high-speed data transmission     

Laboratory evaluation of the XR-Bond Dentin/Enamel Bonding System

The shear bond strengths of the XR-Bonding System used in conjunction with Herculite composite, to the dentine of forty extracted human permanent first and second molars were determined after the test specimens were stored in physiological saline at 37 degrees C for 48 hours, one week, two weeks and four weeks, respectively. A shear load was applied to the base of the bonded composite cylinders with a knife-edged rod at a crosshead speed of 0.5 mm/minute. The shear bond strengths were expressed in megapascals (MPa). The quantitative microleakage of Class V preparations in dentine (cementum) in forty-eight extracted human maxillary permanent canines restored with the same dentinal bonding system and after storage in physiological saline at 37 degrees C for the same time intervals as for the shear bond strength tests, was determined. On the final day of each time interval the teeth were thermocycled X 500 in a 2 per cent methylene blue solution between 8 degrees C and 50 degrees C with a dwell time of 15 seconds. Microleakage was determined by a spectrophotometric dye-recovery method and expressed in microgram dye/restoration. There was a significant trend for the shear bond strengths to increase with duration of storage (p = 0.01) but the quantitative microleakage was not significantly different (p = 0.75).