Miller Technique Research Articles

Interface control mechanisms in horizontal zone-melting with slow rotation

Using slow rotation to control the interfaces in horizontal zone-melting (HZM) is a widely adopted method for organic materials, i.e., the Pfann–Miller technique. A short and nearly axisymmetric zone can be achieved easily even with a rotation rate of only 1 or 2 rpm. However, the melt convection might not be the crucial mechanism for such a control. Through flow and interface visualization during HZM of succinonitrile, the role of the slow rotation was investigated. Three-dimensional numerical simulation was further performed to illustrate the key mechanisms for determining the zone shape. Besides the good agreement between the simulation and the experiments, we found that the heat of fusion released or absorbed during the rotational regrowth and remelting played a crucial role, and it could not be ignored in the simulation.

EXPERIMENTAL PARTIAL UNILATERAL URETER OBSTRUCTION. I. PRESSURE FLOW RELATIONSHIP IN A RAT MODEL WITH MILD AND SEVERE ACUTE URETER OBSTRUCTION

Purpose To create an animal model with mild and severe partial unilateral ureter obstruction in young rats using a modified Ulm and Miller technique and to characterize the model by acute renal pelvic pressure measurements. Material and Methods During anesthesia the upper fourth (n = 15) or the upper two-thirds of the left ureter (n = 15) was embedded into the psoas muscle causing either a mild or severe partial obstruction. Sham-operated control rats were prepared in paralllel (n = 20). The baseline pelvic pressure, the perfusion pelvic pressure (perfusion rates: 0.2 to 1.0 ml. per minute) and peristaltic waves were recorded after a resting period. Results Mean baseline pelvic pressure and perfusion pelvic pressure were significantly higher in obstructed kidneys than in non-obstructed kidneys, and significantly higher in severely obstructed kidneys than in mildly obstructed kidneys (p <0.05). A fair linear relationship existed between the increase in pelvic pressure and the increase in perfusion rates in all groups. In the severely obstructed kidneys, baseline pelvic pressure was 16.9 ± 2.3 cm. H 2 O and the perfusion pelvic pressure increased significantly from 41.7 ± 3.3 cm. H 2 O to 68.6 ± 6.3 cm. H 2 O. The pelvic peristaltic amplitude increased significantly following the increase in perfusion rate and there was a significant difference in the amplitude between severely obstructed and non-obstructed kidneys when the perfusion rate exceeded 0.4 ml. per minute (p <0.05). Conclusion Embedding either the upper fourth or the upper two-thirds of the ureter into the psoas muscle produced a mild or a severe partial obstruction. In the latter model a significant increase in pelvic pressure and amplitude was observed, indicating the existence of severe obstruction.

Structural organization of kinetoplast DNA and its compactization in vitro

Studies on compactizatlon and decompactizatlon of the genome are of great importance for elucidation of structural mechanisms taking part in the regulation of gene activity. Kine to plast DNA (kpDNA) is a convenient model for studies of compactizatlon processes. Leishmania gymnodactyli kpDNA represents a gigantic association of catenated circular molecules of two types: 0.3μm minicircles and 10μm maxicircles. Fig. 1 demonstrates that kpDNA prepared by protein film method is a network of linked circular DNA molecules shaped as a shallow cupor a basket. Part of minicircles exists in relaxed form, part-in a supertwisted. Electron microscopic studies of Leishmania gymnodactyli cells by Miller's technique showed that kpDNA looks like irregularly out line dagglomerations of crossed or sometimes branched rod-like structures 100 to 200nm long and 20nm thick (Fig.2). They seemtore present compacted minicircles. On the electron microscopic preparation these accumulations are lying separately from nuclear chromatin agglomerations and are often torn out together with the flagellum at the basis of which the kinetoplast is located.

Ultrastructural localization of the herpes simplex virus major DNA-binding protein in the nucleus of infected cells.

Using immunocytochemical procedures in conjunction with electron microscopy we have examined the distribution of the major DNA-binding protein (DBP) of herpes simplex virus (HSV) in infected nuclei. In embedded specimens, DBP was preferentially associated with fibrillar material of electron-translucent viral inclusions, and to a lesser extent with peripheral (marginated) host chromatin. The latter association was sensitive to a non-ionic detergent ('Photo flo'). In chromatin spread by the Miller technique, DBP was found to be a component of the 10 nm 'thick filaments' previously described in HSV infection.

Isolation and characterisation of a transcribing polynucleosomal chromatin fraction.

A method is described for preparation of a fraction of chromatin enriched in transcribing regions from nuclei of mouse GR cells. This fraction, released by mild staphylococcal nuclease digestion of isolated nuclei, contains 2 to 10% of the DNA as polynucleosomal chromatin together with 50-70% of pulse-labelled RNA and about 90% of all template-engaged RNA polymerase B molecules, titrated with (3H)-alpha-amanitin. Hybridisation of DNA from this chromatin fraction to total nuclear RNA in excess shows that it is enriched in frequently-transcribed DNA sequences. A modification of the Miller technique, allowing the spreading of the active chromatin fraction for electron microscopy, has been developed. Examination of the spreads reveals that this chromatin fraction contains 20-100 nucleosome-long polynucleosomal chains bearing lateral RNP fibrils interpreted as nascent transcripts. The average length of the DNA fragments in the fraction is greater than that of average transcribed regions, suggesting that the transcribed regions are linked to flanking segments whose chromatin conformation probably contributes to the selective release of transcribing chromatin.

Comparison between proliferative changes and nuclear events during ageing of human fibrobalsts in vitro

Age-related alterations in nucleoproteins were examined in resting human embryonic fibroblasts at various population doubling levels (PDL) with three techniques for electron microscopy. Conventional glutaraldehyde fixation showed nuclear modifications consistent with previously published studies. Miller's technique permitted the detection of transcription complexes in young as well as in old cells. The technique involving the ultrathin sectioning of partially loosened nucleoproteins revealed at high PDL a marked rarefaction of the chromatin threads and the appearance of large parts of the lamina densa devoid of attached threads. In addition, the nucleolar filamentous masses were considerably less tufty than at earlier PDL, suggesting a decrease in the ribosomal transcriptional activity. These changes in nucleoprotein organization were rare before the 40th PDL, then increased to about 5% of the nuclei when the percentage of cells initiating DNA during the first 24 h remained at low levels. They were the rule for the last 6 to 7 PDL and coincided with the time when the maximal densities started to fall rapidly. They appeared after treatment with a loosening medium at neutral and at alkaline pH.

Non-nucleolar transcription complexes of rat liver as revealed by spreading isolated nuclei.

Miller's technique was applied to isolated nuclei of rat liver. Both the usual nucleolar and non-nucleolar transcription complexes were visualized. In addition, an unusual type of putative non-ribosomal transcription unit was revealed. It was charcaterized by a high density of the lateral ribonucleoprotein (RNP) fibrils. Although these particular units exhibited a regular increase of fibril lengths, the length of the transcript-covered deoxyribonucleoprotein (DNP) fibres and the morphological aspect of the RNP fibrils distinguished them from the nucleolar 'Christmas-tree'-like figures. The linear and granular configuration of the transcripts and the absence of terminal knobs made them similar to non-nucleolar nascent RNP fibrils.

Ribosomal transcriptional complexes in subnuclear fractions of chinese hamster ovary cells after short-term actinomycin D treatment

The early effects of a low dose of actinomycin D (AMD) on the ultrastructure of ribosomal transcriptional complexes (RNP) have been studied in Chinese hamster ovary (CHO) cells. After various periods of treatment with a concentration of AMD which inhibits preferentially ribosomal RNA biosynthesis, either whole nuclei or nucleoli were isolated and spread for electron microscopic observation according to a procedure adapted from Miller's technique. Even after the shortest period of AMD action, striking modifications could be detected in the actively transcribed ribosomal matrices. After a 1-min treatment, most of the elongating RNP fibrils were still attached to the DNP strand but abnormally large “gaps” appeared in most of the ribosomal matrix units. After 2 min of action the spacing of the RNP fibrils on the matrix unit became irregular, occasionally simultaneous with a shortening of the length of the RNP-covered segment. Some bound RNA plymerase molecules were devoid of an associated RNP fibril. The visualization of tandem repeat units became exceptional. However, when they were detected, adjacent matrices displayed different patterns of alteration. On and after 5 min of treatment, the frequency of detection of ribosomal matrix units decreased considerably. The persistent ribosomal complexes corresponded generally to the presence of a few residual RNP fibrils.

Electron microscopic visualization of transcribed genes in the nucleus of Amoeba proteus

An electron microscopic examination by Miller's technique of nuclei of Amoeba proteus has revealed two types of transcriptionally active genes. The putative nucleolar genes have an average length of 6.07 μm and are transcribed throughout their entire length. On occasion a second type of active gene is observed as discrete regions on long chromatin fibers. These regions measure 3.21 μm and are not tandemly repeated. We have used Miller's technique [1–3, 13] to observe the transcriptionally active genes in the nucleus of the unicellular protozoan, Amoeba proteus. We report here preliminary observations on the morphology of two types of structures that are interpreted as transcriptionally active genes.

Calculation of the degree of blood mixing by the Davis and Miller transfusion technique

Formulas are derived for determining the amount of blood mixing that occurs when blood transfusions using the Davis and Miller technique are carried out. These formulas indicate that the degree of mixing depends on the blood volumes of the donor and recipient animals, the amount of blood that is withdrawn and injected with each crossing and the total number of crosses that are carried out. It is also shown that the maximum amount of foreign blood that can be transferred from one animal to another depends only on the blood volumes of the two animals.

The Relative Frequency of Sparse Cell Elements -- An Application to Reticulocyte Blood Counts

THE PROBLEM of determining the relative frequency of sparse elements frequently arises in making blood counts. For example, reticulocytes in normal humans comprise about 1% of the total red cells in the blood. With certain types of anemia this percentage rises considerably. The current, standard method for making reticulocyte counts involves counting 1000 red cells, and noting the number of reticulocytes among the 1000. This is a relatively time-consuming technique, and to reduce the work involved another technique has been suggested. The new technique, which we call the Miller technique, involves using an ocular disc which is divided into two areas, as in figure 1, below.

STUDIES ON MENINGOCOCCUS INFECTION

The investigation of this isolated epidemic of meningococcus meningitis at a C.C.C. camp gave an opportunity to examine the carrier state in contacts carrying what were presumably virulent epidemic strains of organisms. With the aid of Miller's technique for the enhancement of the demonstrable virulence of meningococci for mice, it proved possible to test the virulence of the carrier strains from Camp Rusk. These results were consistent despite the interval of from 3 to 4 weeks which intervened between the isolation of the strains and the virulence titrations. Type I strains were found to have a high virulence, while the virulence of Type II strains was moderately high but definitely less than that of the Type I, and atypical strains and strains of N. catarrhalis isolated from carriers showed a very low virulence. The question of the precise nature of the carrier state was investigated. No evidence has been obtained yet as to the existence of a relationship between pharyngitis, coryza or upper respiratory disease and the presence and degree of the carrier state. This is unlike the situation with regard to pneumococcus carriers. On the other hand, it has proved possible to demonstrate reactions within the body to the meningococci in the nasopharynx, consisting of the formation of agglutinins and protective antibodies in the blood serum. 32.3 per cent of Type I and 60 per cent of Type II carrier sera showed moderate or good agglutinins for homologous organisms and 80 per cent of Type I and 40 per cent of Type II sera showed moderate or good protective antibodies against virulent homologous strains. No idea could be obtained as to the relationship of the presence or absence and the degree of serological reaction and the duration of the carrier state.