Massive air embolus to the left ventricle: diagnosis and monitoring by serial echocardiography.

Air embolization is an unusual but potentially dangerous complication in left heart catheterization. Microbubbles can be detected with two-dimensional echocardiography, which is often used for this purpose during open heart and bypass surgeries. Permanent neurologic sequelae and hemodynamic collapse can result from embolization of air bubbles to the cerebral and coronary circulations, respectively. Hyperbaric oxygen is the treatment of choice for cerebral air embolization. We present a 39-year-old man who had air embolization during left ventriculography in the form of a large pocket of "pooled" air. The patient was treated with conservative therapy successfully. Two-dimensional transthoracic echocardiography was used to document the presence of the air and follow its dissolution.

Nucleic acid hybridization assays employing dA-tailed capture probes. I. Multiple capture methods.

A quantitative hybridization assay termed "reversible target capture" is described. The technique is designed to extensively purify the target nucleic acid from crude cell lysates in about 1 h without phenol extraction. Simple, rapid methods are described that explain how each process in the assay is optimized. The procedure involves hybridizing the target nucleic acid in solution with a dA-tailed capture probe and a labeled probe. The capture probe-target-labeled probe "ternary complex" is then captured on magnetic beads containing oligo(dT). After the excess unhybridized labeled probe, cell debris, and other sample impurities are washed away, the intact ternary complex is further purified by chemical elution from the beads and recapture on fresh beads. The ternary complex is then eluted thermally and recaptured on a third set of beads or on poly(dT) filters. This triple capture method results in a detection limit of approximately 0.2 amol (100 fg) of target with 32P-labeled riboprobes. This is approximately 1000 times more sensitive than sandwich assays employing only a single capture step. The method is illustrated by detecting Listeria cells in the presence of heterologous bacteria. With three rounds of target capture, as few as six Listeria cells have been detected in the presence of 1.25 x 10(7) control cells.

Translational regulation of the spc operon in Escherichia coli. Identification and structural analysis of the target site for S8 repressor protein.

The spc ribosomal protein operon of Escherichia coli is feedback-regulated by ribosomal protein S8, a translational repressor. We have analyzed the region of the spc mRNA that is responsible for this regulation. First, we have established that the S8 target site on the mRNA is near the translation start site of the third gene encoding ribosomal protein L5 in the operon. This was done by constructing hybrid plasmids carrying spc operon ribosomal protein genes under lac transcriptional control, as well as their deletion derivatives, and carrying out both in vivo and in vitro protein synthesis experiments. Next, the secondary structure of this region was studied by analyzing 5' end-labeled RNA synthesized from the phage SP6 promoter using structure-specific nucleases. A secondary structure model consistent with the results was deduced with the aid of a computer prediction of RNA folding. In addition, we cloned and sequenced the corresponding region from Salmonella typhimurium, Proteus vulgaris and Serratia marcescens and found five "compensating" substitutions that support some of the deduced helical structures of mRNA. None of the base changes was inconsistent with the deduced secondary structure model. Finally, site-directed mutagenesis experiments have identified bases important for regulation, including two base-paired sites representing each of two helical regions. This has led to the conclusion that some specific nucleotide residues located between these two helical regions are directly involved in S8 recognition, and that the function of the two helical regions is to maintain the proper orientation of these nucleotide residues. Comparison of the structure of the S8 target site on the spc mRNA with the known S8 binding site on rRNA has revealed a striking similarity in both primary and secondary structures. In particular, primary sequences of rRNA conserved among distantly related bacterial species in this region is found to be identical with the sequences at the corresponding positions in mRNA. These results suggest that the same structural features of the S8 repressor protein are involved in the interaction with both 16 S rRNA and the mRNA target site.

Secondary structure of the autoregulatory mRNA binding site of ribosomal protein L1.

The secondary structure of the autoregulatory mRNA binding site of Escherichia coli ribosomal protein L1 has been studied using enzymatic methods. The control region of the E. coli L11 operon was cloned into a vector under control of the Salmonella phage SP6 promoter, and RNA transcribed using SP6 RNA polymerase. The secondary structure of this RNA was probed using structure-specific nucleases, and by comparison of the data with computer predictions of RNA folding, secondary structural features were deduced. The proposed model is consistent with elements of some previously proposed models, but differs in other features. Finally, secondary structure information was obtained from two mutant mRNAs and the structural features correlated with observed phenotypes of the mutants.

X-ray solution-scattering studies of active and inactive Escherichia coli ribosomal subunits.

That ribosomal subunits can exist in active and inactive functional states, and that subunits in the two states are interconvertible, has been known for some time (see Zamir et al., 1974). The magnitude of the conformational perturbation accompanying this functional transformation, however, was not known. In the present study 30 S and 50 S subunits in the two functional states have been compared by small-angle X-ray scattering. The results indicate that the structural differences between active and inactive subunits are small, at the limit of resolution of this technique. Model studies show that the data imply conformational differences at or below the limit of resolution of other physical methods now in use to examine the detailed structure of these particles.