Optimization of a method for simultaneous analysis of cell surface phenotype and cell cycle in human bone marrow and peripheral blood leukocytes by flow cytometry: the value of both internal and external standards

Three different methods for the simultaneous analysis of surface phenotype and DNA quantification were compared. One method, involving the fixation of cells in 70% ethanol, was convincingly superior, both with regard to the CV of the G0G1 peak and the intensity of the DNA labelling. Furthermore, the correlation between the surface antigen densities before and after fixation were high. Experiments evaluating the intraday and the interday variation of the DNA ratio (the mean channel of the G0G1 peak of the sample divided by the mean channel of the G0G1 peak of chicken erythrocytes), documented the former to be small, with S.D. values varying from 0.0 to 0.016, while the latter were considerably higher with S.D. values varying from 0.077 to 0.123. Since the intraday variation of the DNA ratio was consistently low and the interday variation strongly correlated to the position of the red fluorescence test beads, it was possible to minimize the interday variation of the DNA ratio, by calculating the DNA index as the ratio between the DNA ratio of the sample and that of an external control (buffy coat leukocytes). Analyzing normal bone marrow and calculating the DNA index (DI) on the basis of these ratios, the confidence limits of the DI were decreased by more than half the values obtained when DI calculation was based solely on an internal standard, thereby making subsequent ploidy determinations of patient samples more precise. We conclude that this setup of internal and external standards allows accurate determinations of DNA aneuploidy even in an assay where whole cells labelled for surface antigen and DNA content are analyzed.     

Analysis of linkage between beta-lactoglobulin and J blood group system loci in cattle

Linkage between loci controlling variants of beta-lactoglobulin and blood groups of the J system in cattle was studied by means of stochastic genetic methods reported earlier. The studies were conducted on a herd of Black Pied cattle improved with Holstein sires; population genetic data were analyzed. A plot for lod score was constructed, and point (r - 0.28) and interval estimations of the coefficient of recombination were obtained. The results are in good agreement with earlier reported data on other subjects.     

Endoplasmic reticulum Ca(2+)-ATPase in microsomal vesicles isolated from bovine retinae

Active Ca2+ transport was measured in microsomal vesicles prepared from bovine retinae and was compared with that in disk membranes of the photoreceptor cells of the same retina. The 45Ca uptake was dependent on the presence of Mg(2+)-ATP and was inhibited by vanadate or when GTP substituted for ATP. The dependence of calcium uptake on the external free Ca2+ concentration gave a KM = 13 microM or a KM = 0.1 microM for disks and microsomal vesicles, respectively. A phosphorylated intermediate (E-P) of Ca(2+)-ATPase of about 100 kDa was isolated in microsomal vesicles. The E-P formation was strongly inhibited by thapsigargin and partially by 2,5-di-(-butyl)benzohydroquinone. Digestion of disks or microsomes with calpain had no effect on the phosphorylated intermediate, while digestion with trypsin produced two fragments of approximately 55 kDa and 35 kDa. These results suggest that bovine retinal microsomes contain a calcium pump belonging to the SERCA family.     

Prediction of fire classification for wood based products. A multivariate statistical approach based on the cone calorimeter

Wood has long traditions as a building material, and is often used in construction elements, and as interior and exterior surfaces in the Nordic countries. In most applications, there are reaction to fire requirements to products used as surfaces, e.g. in escape routes and larger public spaces. Most wood products will therefore have to be treated with fire retardant (FR) agents to fulfil the strict requirements to properties connected to heat release and flame spread. Unfortunately, FR agents usually also increase the smoke production, as they cause a more incomplete combustion of the wood. The wood product manufacturers seek to find the optimal amount of FR additives where both heat release and smoke production in the classifying test are within the requirements given in the building regulations. This paper describes models for prediction of the European reaction to fire classes of wood products. The models are based on multivariate statistical analysis, and use test results from the cone calorimeter test as input. The presented models are, with very good precision, able to predict which Euroclass and additional smoke class a wood based product would obtain if it were to be tested in the single burning item test. Copyright © 2006 John Wiley & Sons, Ltd.     

Correlated response of udder dimensions to selection for milk yield in Holsteins

Response to selection for milk yield in Holsteins was examined by a controlled selection experiment initiated in 1964. Foundation cows were paired by sire and divided randomly into two breeding groups, selection and control. Selection group was mated to four sires each year highest for Predicted Difference milk. Twenty bulls selected in 1964 as near breed average for milk sired all control cows. Milk yield was recorded for each lactation, and height of udder from ground, distances between teats, and perimeter and area bound by the four teats were measured at 30 to 75 days postpartum on 153 selection and 202 control cows over 14 yr. Most estimates of repeatability and heritability of udder dimensions before and after milking were .45 or larger. Correlations with milk yield were negative for udder height and positive for other udder measurements. Selection cows exceeded controls for lactational milk yield. Daughters of high milk bulls had greater distances between teats, greater perimeters, and larger areas of udder floor. Selection cows did not differ from controls in udder height for first parity but were below controls in udder height and had udders that collapsed more for all parities.     

Vascular malformations of the jaw bones. Report on nine patients

Partial complementary DNA (cDNA) for thymidine phosphorylase (dThdPase) was cloned by means of a polymerase chain reaction. There was complete sequence identity between the amino acid sequence deduced from the nucleotide sequence of a clone (288 nucleotides) and the residues of platelet-derived endothelial cell growth factor (PD-ECGF). The amino acid sequence of all four peptide fragments from purified human dThdPase could be aligned with that of PD-ECGF. Our data indicate that residues 125-244 of PD-ECGF are identical to the sequence of human dThdPase. The molecular weights of human dThdPase and recombinant PD-ECGF (rPD-ECGF) that lacks 10 amino acids at the amino terminal were 55 and 52 kDa, respectively. Anti-PD-ECGF antibody recognized dThdPase, and anti-dThdPase antibody recognized rPD-ECGF. rPD-ECGF had dThdPase activity and its specific activity was similar to that of purified human dThdPase. dThdPase activity and molecules were detected in COS cells transfected with human PD-ECGF cDNA, but not in nontransfected cells. The sizes of PD-ECGF and dThdPase in the transfected COS cells were identical. These data suggest that human dThdPase is identical to PD-ECGF.     

Long-term prognosis for women with chronic pelvic pain without laparoscopically demonstrable cause

With the object of assessing the long-term prognosis and the frequencies of recurrence and remission in women chronic low abdominal pain without laparoscopically visible cause, questionnaire were sent in 1985 and 1991 to 55 women who had been submitted to laparoscopy in 1982-1984 for this reason. These women had been told that there was no demonstrable explanation of the pain experienced and were then discharged. 65% and 55% respectively had experienced and unfavourable course with considerable and continued symptoms. Only 22% stated in both investigations that they had experienced a favourable course and that they were, by and large, free from pain. 36% changed from an unfavourable to a favourable course or the reverse. The assessment made by the women was confirmed by a series of subordinate questions and this demonstrated a marked difference between the favourable and unfavourable courses of the condition. It is concluded that laparoscopy with exclusion of significant pathology is not, in itself, satisfactory as treatment of this patient group and that no improvement occurs in the course of time. The condition varies greatly with many recurrences and remissions and, for this reason, uncontrolled reports of the therapeutic effects are of no significance. When compared with the literature, it is suggested that this patient group should be referred early in the course of the condition to a therapist with specialist psychological/sexological insight and/or to a physiotherapist with interest in this patient group.