Evaluation of the diagnostic application of an enzyme immunoassay for Clostridium perfringens type A enterotoxin

The diagnostic application of an enzyme immunoassay for Clostridium perfringens type A enterotoxin was evaluated. Test results from 100 individuals associated with C. perfringens gastroenteritis outbreaks and 111 control individuals were included. The assay sensitivity was 93.7%, and the assay specificity was 98.7%.     

pHG165: A pBR322 copy number derivative of pUC8 for cloning and expression

During the construction of the Messing pUC plasmid series, the rop(rom) gene of pBR322 which mediates the activity of RNAI was deleted. This has resulted in an elevated copy number for the pUC plasmids which makes the expression of beta-galactosidase activity constitutive in a host containing the Iqtss lac repressor. We describe the construction of a new series of vectors which retain the pUC multiple cloning site (MCS) but in which copy number control has been recovered. In addition, the lac alpha/lac promoter expression region has been inserted into a HpaI cassette. This facilitates the movement of recombinant DNA clones within the MCS. It also increases the complementation activity of the lac alpha peptide by an order of magnitude, allowing selection of recombinants by their Lac- phenotype on MacConkey agar.     

Photosynthetic pathways in a midwestern rock outcrop succulent,Sedum nuttallianum Raf. (Crassulaceae)

Shoots of Sedum nuttallianum exhibited CAM^* acid fluctuations in the field. These nocturnal acid accumulations persisted in the laboratory under well-watered and water-stressed conditions. Simultaneous measurements of transpiration, however, indicated daytime stomatal opening and nocturnal stomatal closure. Measurements of CO₂ and H₂O vapor exchange continuously for six days after watering substantiated these results in part: the majority of CO₂ uptake occurred during the day early in the experiment; however, after several days without water, nighttime CO₂ uptake was stimulated and eventually was greater than the drastically reduced daytime CO₂ uptake. This nighttime uptake was never quite sufficient to account for all estimated increases in tissue acidity. Thus, a combination of CAM and CAM-cycling occurred early in the desiccation experiment. Evidence for CAM and a form of CAM-idling was found later in the experiment. Though nighttime CO₂ uptake occurred and persisted after only one day without water, rates were too low to alter the tissue ¹³C/¹²C value from a C₃-like number (–30). Thus, although CAM and CAM-idling may have survival value during extended droughts, shoots of S. nuttallianum apparently utilize the C₃ pathway to obtain most of their carbon.Abbreviations C₃ pathway - CO₂ fixation pathway in which an intermediate containing 3 carbon atoms is formed - CAM Crassulacean acid metabolism - Chl Chlorophyll - c_i internal CO₂ concentration - DW Dry weight - g_c mean conductance to CO₂ - FW Fresh weight - PAR Photosynthetically active radiation - SD Standard deviation - vpd Vapor pressure deficit - WUE Water use efficiency     

The murine Hox-2 cluster of homeo box containing genes maps distal on chromosome 11 near the tail-short (Ts) locus

Two probes derived from a mouse recombinant lambda-clone (H24.1), that contains a sequence closely homologous to the Drosophila antennapedia homeo box, were mapped to mouse chromosome (MMU) 11 by filter hybridization of somatic cell hybrid DNA. This sequence is highly homologous to a human homeo box gene (HOX2) and appears to represent one of the two genes in the Hox-2 cluster previously assigned to MMU 11. To regionally map the Hox-2 cluster, we have carried out in situ hybridization of the two H24.1 probes and of an independently isolated Hox-2 probe. The autoradiographic silver grain distributions were similar in all three experiments with a peak over band 11D. This region contains the locus for the tail-short (Ts) mutation which causes skeletal abnormalities in heterozygotes and early embryonic death in homozygotes.     

Mechanism of intramolecular recyclization and deletion formation following transformation of Escherichia coli with linearized plasmid DNA.

The deletion end-points of a number of type I (less than monomeric) plasmid deletants obtained by transforming recA+ or recA- E. coli with linear pBR322 DNA were determined by DNA sequencing. In both monodirectional and bidirectional deletions the recyclization point was normally characterized by recombination between directly repeated sequences of between 4 and 10 bp present on each arm of the linearized pBR322 molecule. Frequently, short tracts of uninterrupted homology involved in recombinational recircularization were embedded in regions of relative non-homology. A model predicting the probability of matching sequences in either end of a linear plasmid molecule is presented. It is proposed that exonucleolytic processing of the exposed termini of linear plasmid molecules generates substrates for subsequent recombinational recyclization and deletion. The activity of host recombination and repair functions in recircularizing linear DNA molecules explains the generation of many of the aberrant recombinant DNA constructs obtained during gene cloning procedures.     

The full length coding sequence of rat liver androsterone UDP-glucuronyltransferase cDNA and comparison with other members of this gene family.

Cloned cDNAs coding for hepatic UDP-glucuronyltransferase (UDPGT) have been isolated from a rat liver cDNA library in the expression vector bacteriophage lambda gt11 using anti-UDPGT antibodies. Four different mRNAs have been identified by sequencing of 15 UDPGT cDNA clones. The sequences of the four classes of cDNA were determined to be 85-95% homologous. Restriction fragments were isolated from the cDNA in each class and used as class specific probes. Hybridisation of these probes to northern blots of total RNA prepared from the livers of normal and genetically deficient Wistar rats identified the cDNA in class 4 with androsterone UDPGT. Translation of the cDNA sequence of clone rlug 23, the longest member of class 4, allowed determination of the complete amino acid sequence of androsterone UDPGT.     

Monoclonal antibodies against human ovarian tumor associated antigen NB/70K: Preparation and use in a radioimmunoassay for measuring NB/70K in serum

Summary Murine monoclonal antibodies (MCAs) against human ovarian tumor associated antigen NB/70K have been prepared. One of these MCAs, NB12123, was chosen for the development of a radioimmunoassay for measuring serum NB/70K levels. In this assay, the average NB/70K level in 75 normal, healthy controls was 11.9 activity units (AU) with an SD of 14.9 AU. The normal cut off value for this assay was set at 45 AU (mean +2 SD). 24 of 46 (52%) ovarian cancer patients, 7 of 18 (39%) patients with benign ovarian cysts or tumors and 3 of 85 (4%) control samples had elevated serum NB/70K levels. Comparison of NB/70K levels measured in the NB12123 assay with levels measured in an assay using a polyclonal antiNB/70K previously developed in our laboratory [13] indicated that although both assays had approximately the same percentage of positive ovarian cancer patient samples, there appeared to be no correlation between the absolute NB/70K levels measured by the two assays. The rank of ovarian cancer patient samples was also different for the two assays. Also, almost 40% of patients with benign ovarian cysts and tumors had elevated serum NB/70K levels as measured by the NB12123 assay as compared to 0% for the polyclonal assay. Reciprocal cross-blocking experiments, absorption studies, and immune precipitate analysis indicated that both the monoclonal NB12123 assay and the polyclonal antiNB/70K assay measured the same population of NB/70K molecules. However, the polyclonal antibody recognizes epitopes in addition to that recognized by NB12123. Taken together, these results suggest that the epitope recognized by NB12123 is not as specific for malignant ovarian tumors as the epitope(s) recognized by polyclonal antiNB/70K and/or that more than the one epitope detected by the MCA is responsible for the specificity for ovarian cancer of the polyclonal NB/70K assay. In spite of this, the greater sensitivity and range of the monoclonal NB12123 assay make it possible to monitor serum NB/70K levels in ovarian cancer patients. In four patients examined, the fluctuating serum NB/70K levels appeared to correlate well with clinical statusSupported in part by ACS # PDT 231 and a grant from the Elsa U. Pardee Foundation     

Characterization of RNA synthesis in an Escherichia coli mutant with a temperature-sensitive lesion in stable RNA synthesis

Previous experiments with Escherichia coli strain 2S142 have shown that the synthesis of stable RNA is preferentially blocked at the restrictive temperature. In this paper, we have examined the capacity of this mutant strain to synthesize RNA in vitro. Growth of the strain for as short a period as 10 min at 42 degrees C resulted in a 40 to 60% loss of RNA synthetic capacity and a fourfold decrease in percent rRNA synthesized in toluenized cell preparations. The time course for the loss and recovery of this RNA synthetic capacity correlated very well with the changes in RNA synthesis observed in vivo. We found no difference in temperature sensitivity of the purified RNA polymerase from the mutant and the parental strains. Moreover, there was no detectable alteration in the amount of enzyme, specific activity of the enzyme, or electrophoretic mobility of the subunits when the mutant strain was grown at 42 degrees C. The capacity for rRNA synthesis was also measured with the Zubay in vitro system (Reiness et al., Proc. Natl. Acad. Sci. 72:2881-2885, 1975). Supernatant fractions (S-30) prepared from cells grown at 30 degrees C were capable of up to 31.2% rRNA synthesis, using phi 80d3 DNA as template. S-30 fractions from cells grown at 42 degrees C synthesized 8.6% rRNA. The bottom one-third of the S-100 fraction and the ribosomal salt wash from 30 degrees C cells contained one or more factors which partially restored preferential rRNA synthesis in S-30 fractions from cells grown at 42 degrees C. Preliminary evidence suggests that the factor(s) is protein in nature.     

Peripheral glucose uptake in young men with myocardial infarction

Forearm glucose uptake (FGU) was studied during 100 g oral glucose tolerance tests (GTT) in nonobese, nondiabetic men who had suffered a myocardial infarction (MI) at or before the age of 40, and the results compared with the response in age-matched normal men. In the MI group the rise in both glucose and insulin concentrations after glucose loading was similar to that in normal subjects, although in the former, peak levels tended to be slightly higher. Concomitant FGU, however, was significantly greater in the MI group than in control subjects in the period 0-90 min and in the test as a whole (0-180 min). The results show that at least in some nondiabetics suffering MI at an early age hyperinsulinism is not a feature and peripheral tissue sensitivity is increased.     

Action of hydroxyethyl starch on the flow properties of human erythrocyte suspensions

Hydroxyethyl starch (HES) has often been used as a plasma expander, but questions still remain concerning the mechanisms by which it produces changes in the rheological properties of blood and erythrocyte (RBC) suspensions under various flow conditions. The present investigation has shown that the dynamic viscosity of HES (232,000 and 565,000 daltons) solutions rises in a nonlinear fashion with increasing HES concentration, and for a given concentration of HES exhibits Newtonian behavior at shear rates between 0.15 to 124 sec-1. At low (less than 0.9 sec-1) shear rates the apparent viscosity of a 40% RBC suspension increases with lower concentrations of HES because of RBC aggregation. At higher concentrations of HES, increases in suspension viscosity are due to an increase in the viscosity of the continuous phase since the RBC are largely disaggregated. At high (greater than 36 sec-1) shear rates the relative viscosity (eta/eta O) of RBC suspensions slowly decreases with increasing HES concentration. At low shear rates eta/eta O increases and then decreases with increasing HES concentration. Evidence of the concentration-dependent effects of HES on RBC aggregation is provided not only by the viscometric analysis but also from measurements of erythrocyte sedimentation rate (ESR) and the zeta sedimentation ratio (ZSR). HES is a more potent aggregating agent in phosphate buffered saline (PBS) than it is in plasma. Polymer size has only a slight effect on the extent of RBC aggregation produced, but does have a significant effect on the concentration of polymer at which maximum aggregation occurs. The viscosity-corrected electrophoretic mobility of RBC in HES rises monotonically with the concentration of HES in the suspending medium. Decreases in the extent of RBC aggregation with increasing polymer concentrations probably result from an increase in the electrostatic repulsive forces between the cells.