Structural and functional characterization of proteolytic fragments derived from the C-terminal regions of bovine fibrinogen

A number of new as well as previously described fragments derived from the D region of bovine fibrinogen by limited proteolysis have been characterized by sequence analysis, differential scanning calorimetry and circular dichroism. Determination of the extremities of the polypeptide chains forming individual fragments allowed the scheme of proteolysis and the borders between domains in the D region of fibrinogen to be established. It was also found that the most thermostable region of the D fragment (TSD) can be substantially reduced in size without loss of its compact structure. The alpha-helical content of the newly prepared 21-kDa TSD2 and 16-kDa TSD3 fragments were 82% and 75%, respectively, strongly supporting a coiled-coil structure for this region of the fibrinogen molecule. The DX and DZ fragments, prepared from a chymotryptic digest of the DLA fragment, were found to be similar to the DL and DY fragments, respectively, except for an internal cleavage at K393-T394 in their beta chains. This cleavage leads to destabilization of all thermolabile domains, indicating interaction between them. The DL and DY fragments, containing only one polymerization site in their beta chains, were able to inhibit fibrin polymerization at high concentration. However, these same fragments failed to bind to fibrin-Sepharose under conditions where their structural analogues, DX and DZ, were tightly bound, indicating that cleavage after K393 substantially increases the affinity of this site.     

Long-range correlation properties of coding and noncoding DNA sequences: GenBank analysis

An open question in computational molecular biology is whether long-range correlations are present in both coding and noncoding DNA or only in the latter. To answer this question, we consider all 33301 coding and all 29453 noncoding eukaryotic sequences--each of length larger than 512 base pairs (bp)--in the present release of the GenBank to dtermine whether there is any statistically significant distinction in their long-range correlation properties. Standard fast Fourier transform (FFT) analysis indicates that coding sequences have practically no correlations in the range from 10 bp to 100 bp (spectral exponent beta=0.00 +/- 0.04, where the uncertainty is two standard deviations). In contrast, for noncoding sequences, the average value of the spectral exponent beta is positive (0.16 +/- 0.05) which unambiguously shows the presence of long-range correlations. We also separately analyze the 874 coding and the 1157 noncoding sequences that have more than 4096 bp and find a larger region of power-law behavior. We calculate the probability that these two data sets (coding and noncoding) were drawn from the same distribution and we find that it is less than 10(-10). We obtain independent confirmation of these findings using the method of detrended fluctuation analysis (DFA), which is designed to treat sequences with statistical heterogeneity, such as DNA's known mosaic structure ("patchiness") arising from the nonstationarity of nucleotide concentration. The near-perfect agreement between the two independent analysis methods, FFT and DFA, increases the confidence in the reliability of our conclusion.     

The OKT3 Antibody Response Study: a multicentre study of human anti-mouse antibody (HAMA) production following OKT3 use in solid organ transplantation

Human anti-murine antibody titres following patient exposure to the monoclonal antibody Orthoclone OKT3 (muromonab-CD3) are determined by laboratories using diverse analytical methods which are not standardized and whose concordance is not established. A multicentre study group therefore compared testing for IgG anti-OKT3 antibody among seven laboratories. A set of 270 sera was obtained from 30 heart, 30 kidney and 30 liver transplant recipients with no previous exposure to OKT3 who were receiving OKT3 for induction immunosuppression. Sera were collected from each patient prior to and at 24 +/- 2 days and 31 +/- 2 days following initial OKT3 exposure. Identical aliquots of all 270 sera were tested for IgG anti-OKT3 antibody by each laboratory. In addition, the limit of detection of each laboratory's method was estimated by titration of an affinity-purified IgG anti-OKT3 reference material of known concentration. Anti-OKT3 antibody formation differed greatly among the three organ groups. Cardiac patients demonstrated the least sensitization and almost exclusively lower titres, while kidney recipients had more frequent and higher titre antibody formation. Liver recipients yielded the highest sensitization rate and the most frequent high titre sera. Importantly, the seven laboratories differed widely in the number of pretreatment sera reported as positive (ranging from 0% to 41% among laboratories), the number of post-OKT3 sera reported as positive (17-63%), the number of post-OKT3 samples with titre > or = 1000 (2-31%), and the number of patients sensitized 19-69%). Concordance among laboratories was highly variable, with interlaboratory agreement ranging from 38% to 83% on the sample titres assigned to 180 post-OKT3 sera. Many of the discordant results were consistent with differences in the limit of detection of the analytical methods, which ranged from 0.19 microgram/ml to > or = 15 micrograms/ml, a nearly 100-fold difference among laboratories. This study demonstrated the presence of both good concordance and significant discordance among laboratories in determining human anti-mouse antibody titres, and demonstrated that common titre categories (100, 1000, 10,000) were not equivalent among laboratories. The level of concordance among methods should be considered when comparing anti-OKT3 antibody results from different centres and their correlation with clinical events. Universal comparative testing, patterned after proficiency testing programmes, is needed to assess differences among laboratories and to bring uniformity and a sound interpretative basis to this field of testing.     

Patient compliance in mobile screening mammography

RATIONALE AND OBJECTIVES: We assessed the follow-up behavior of women who had abnormal results of screening mammograms taken on a mobile van. METHODS: A retrospective cohort study was conducted between 1988 and 1991 of all women served by a mobile mammography van in rural North Carolina. Compliance with radiologist recommendations for follow-up was assessed through review of patient records and mail surveys of patients with incomplete records. RESULTS: Compliance was 44% for negative or benign mammograms, 57% for indeterminate mammograms, and 62% for probably malignant or malignant mammograms. Women who had a previous mammogram or had a malignant finding were more likely to comply with follow-up recommendations (p < .0001) than women with normal or benign results and no history of mammography. Compliers and noncompliers did not differ with respect to family history of breast cancer or personal history of breast discomfort. CONCLUSION: Compliance with recommendations in this setting was lower than expected. This may be because rural women using mobile van mammography have limited access to the resources needed for appropriate follow-up. Further research is needed to examine explanations for poor compliance in this setting.     

Performance of children with ADHD on the Rey-Osterrieth complex figure: a pilot neuropsychological study

This study evaluates the performance of boys with Attention Deficit Hyperactivity Disorder (ADHD) on the Rey-Osterrieth Complex Figure (ROCF) taking into consideration familiality and comorbid psychiatric and learning disorders (LD). Sixty-five children with ADHD performed at developmentally lower levels of Copy Organization and Recall Style than did 45 controls. ADHD children with LD scored significantly lower on Copy Organization than did ADHD children without LD, whereas psychiatric comorbidity and familiality had no effect. These results suggest that a developmental analysis of the ROCF identifies organizational difficulties associated with ADHD and that these impairments cannot simply be attributed to comorbidities associated with ADHD.     

'Tunable' positive and negative surface charges on a capillary wall: exploiting the Immobiline chemistry

The Immobiline (weak acrylamido acids and bases) chemistry has been applied to the covalent attachment of a positively (or, if needed, negatively) charged layer onto the inner surface of the silica wall. In particular, the following basic Immobilines have been used: pK 6.2, pK 7.0, pK 8.5 and pK 9.3. In order to avoid pK changes, the charged Immobilines are mixed with neutral acrylamido derivatives (in particular the highly resistant and hydrophilic N-acryloyl aminoethoxyethanol) so as to form a co-polymer having a 1:5 molar ratio (charged to neutral). The mu(eo) vs. pH curves have a slope opposite to that of a naked capillary and fan out on the pH scale following the titration curves of the different weak bases. Such chemistry allows the covalent attachment of charged species having known pK values and offering controlled charged densities on the wall. However, with the atomic force microscope, it is found that such soft coatings (whether charged or neutral) do not seem to provide complete coverage of the surface: naked patches of fused silica are found interdispersed among the polymer-coated ones. A good solution is a hybrid bonded and dynamic coating, obtained by adding short chain linear polyacrylamides to the background electrolyte. Good separations of polycations [poly(L-histidine)] and of histones are reported up to pH 5.7.