Biological activities of phthalocyanines--III. Photoinactivation of V-79 Chinese hamster cells by tetrasulfophthalocyanines.
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Biomedical subjects
Publications and source records attributed to H Ali.
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In addition to the highly conserved AATAAA sequence, there is a requirement for specific sequences downstream of polyadenylic acid [poly(A)] cleavage sites to generate correct mRNA 3' termini. Previous experiments demonstrated that 35 nucleotides downstream of the E2A poly(A) site were sufficient but 20 nucleotides were not. The construction and assay of bidirectional deletion mutants in the adenovirus E2A poly(A) site indicates that there may be redundant multiple sequence elements that affect poly(A) site usage. Sequences between the poly(A) site and 31 nucleotides downstream were not essential for efficient cleavage. Further deletion downstream (3' to +31) abolished efficient cleavage in certain constructions but not all. Between +20 and +38 the sequence T(A/G)TTTTT was duplicated. Function was retained when one copy of the sequence was present, suggesting that this sequence represents an essential element. There may also be additional sequences distal to +43 that can function. To establish common features of poly(A) sites, we also analyzed the early simian virus 40 (SV40) poly(A) site for essential sequences. An SV40 poly(A) site deletion that retained 18 nucleotides downstream of the cleavage site was fully functional while one that retained 5 nucleotides downstream was not, thus defining sequences required for cleavage. Comparison of the SV40 sequences with those from E2A did not reveal significant homologies. Nevertheless, normal cleavage and polyadenylation could be restored at the early SV40 poly(A) site by the addition of downstream sequences from the adenovirus E2A poly(A) site to the SV40 +5 mutant. The same sequences that were required in the E2A site for efficient cleavage also restored activity to the SV40 poly(A) site.
Mast cells from different species and even from given tissues within a particular animal are shown to differ in their responsivity to histamine liberators and anti-allergic drugs. The significance of these findings is discussed.
The 3' terminus of most, if not all, eucaryotic polyadenylated mRNAs is formed as a result of endonucleolytic cleavage of a larger precursor RNA. That is, transcription does not terminate at the mRNA 3' sequence but rather proceeds through this site, terminating at some distance downstream. Using a plasmid containing the adenovirus E2A transcriptional unit, we have investigated the sequence requirement for the formation of a mature mRNA 3' terminus, focusing on the role of sequences immediately distal to the poly(A) addition site. Deletion mutants were constructed in the region distal to the poly(A) addition site and assayed by transfection into human 293 cells. The results demonstrate that 35 nucleotides distal to the site of poly(A) addition are sufficient for the formation of a mature E2 mRNA. However, removal of an additional 15 nucleotides, leaving 20 nucleotides distal to the poly(A) site, abolished the ability to produce functional E2A mRNA. The defect in the production of functional mRNA from such a mutant appears to be in the proper cleavage of the primary transcript at the poly(A) addition site. It would thus appear that sequences immediately distal to the site of poly(A) addition do not contribute to the mature mRNA but are essential for the formation of mature mRNA.
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Hospital treatment staff are constantly faced with the problem of psychiatric patients who discharge themselves against medical advice before their treatment is completed. A total of 50 patients who signed themselves out of the 9th floor service of the Clarke Institute of Psychiatry, over a two year period, were analyzed. When compared with a random sample of patients who remained for the duration of their treatment, the authors found the AMA patients to have the following characteristics: The patient is likely to be a young male, admitted as an emergency, remain in hospital for three days or less and have a diagnosis of personality disorder or schizophrenia. The authors conclude with a number of recommendations to assist in dealing with the problem.
Misdiagnosis of maxillary sinus hypophasia usually as sinus infection, sometimes as neoplasm, can lead to unnecessary and difficult surgical explorations. Associated anatomical abnormalities, e.g., caudal displacement of the ipsilateral frontal lobe of the brain or central position in the maxilla of the infraorbital nerve may create unexpected surgical hazards. Associated orbital enlargement can lead to diagnostic confusion in the investigation of headache, especially if the superior orbital fissures show marked asymmetry suggesting erosion. Projection of the fissure into the antrum in Waters view can simulate trabeculation of the sinus or fracture of the inferior orbital rim. Radiologic examination of 500 patients without intracranial or intraorbital lesions revealed maxillary sinus hypoplasia in 36 cases (7.2%) and in half the hypoplasia was unilateral (sinus asymmetry); aplasia was not encountered. Fissure asymmetry was present in 30 cases (6%), being present in 3.66% of patients with normally developed sinuses and in 36.1% of the hypoplasia patients. The appearances and measurements of the fissure are presented and examples of marked normal fissure asymmetry are demonstrated. Maxillary sinus hypoplasia is classified in this series as grade I-mild hypoplasia with limited inferolateral expansion (4 cases) and grade II in which there is also a curved orbital floor and lateral displacement of the adjacent nasal wall (32 cases).
Two pathways of ammonium assimilation are known in bacteria, one mediated by glutamate dehydrogenase, the other by glutamine synthetase and glutamate synthase. The activities of these three enzymes were measured in crude extracts from four Rhizobium meliloti wild-type strains, 2011, M15S, 444 and 12. All the strains had active glutamine synthetase and NADP-linked glutamate synthase. Assimilatory glutamate dehydrogenase activity was present in strains 2011, M15S, 444, but not in strain 12. Three glutamate synthase deficient mutants were isolated from strain 2011. They were unable to use 1 mM ammonium as a sole nitrogen source. However, increased ammonium concentration allowed these mutants to assimilate ammonium via glutamate dehydrogenase. It was found that the sole mode of ammonium assimilation in strain 12 is the glutamine synthetase-glutamate synthase route; whereas the two pathways are functional in strain 2011.
We synthesized the m-maleimidobenzoyl derivative of digoxigenin-3-0-succinate (through a p-phenylenediamine bridge) as a hapten derivative directed towards coupling to sulfhydryl grouos of beta-galactosidase. Prepared enzyme conjugate had about 97% of the enzyme labeled with the hapten derivative while retaining full enzyme activity. The enzyme immunoassay for digoxin we prepared showed a maximum sensitivity of 30 pg per assay (c.v. = 3%) with minimal cross-reaction with digotoxin (3.8%). Our method for hapten conjugation to beta-galactosidase is highly efficient and is simple and easily replicated.
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Conjugates between bovine serum albumin and (R,S)-2-aminonicotine were produced, and these conjugates were employed in rabbits and goats for the production of nicotine antibodies. In the assay of nicotine, an 125I-tyrosine methyl ester derivative of (R,S)-6-aminonicotine was employed as radioligand. The antibody-bound derivative was separated from the free derivative by charcoal adsorption (0.5% charcoal, 0.1% dextran T-70, 0.1% bovine serum albumin pH 7.3). Among the twenty five nicotine derivatives and metabolites examined, (R,S)-6-aminonicotine gave the highest cross-reaction. Cross-reaction with cotinine, a major mammalian metabolite of nicotine, was less than 0.1% for both the rabbit-derived and goat-derived antisera. Cross-reaction by other metabolites, such as (S)-nicotine-N'-oxide, (S)-nornicotine, and N-methylpyrrolidine was less than 1%. The antibodies produced were thus highly specific to nicotine. The radioimmunoassay for nicotine showed a maximum sensitivity of 10 ng/ml in 50-microliter plasma samples for both antisera. After the smoking of a single cigarette (1.2 mg nicotine content in mainstream) the peak of blood plasma level of nicotine in the subjects varied from 20--104 ng/ml, and high levels of nicotine were not necessarily found in heavy smokers.
Enzyme immunoassay for cortisol was developed using beta-galactosidase as an enzyme label and m-maleimidobenzoyl derivatives of cortisol, i.e. cortisol-21-m-maleimidobenzoate (CT-MB) and cortisol-21-hemisuccinate conjugated with m-maleimidobenzoic acid through p-phenylenediamine linkage (CHS-MB), as the haptens coupled to sulfhydryl groups of the enzyme. This enzyme-coupling procedure was highly efficient; over 90% of the enzyme was labeled while full enzyme activity was retained. CHS-MB-beta-galactosidase conjugate showed high immunoreactivity to antibody produced against cortisol-21-hemisuccinate-bovine serum albumin but showed poor displacement with the added cortisol. CT-MB-beta-galactosidase conjugate, however, showed not only a high immunoreactivity to the antibody but also displaced well with cortisol, showing maximum sensitivity of 1 microgram/dl with a 20-microliter sample size. Modification around the linkage of cortisol derivative resulted in high sensitivity to cortisol. Cross-reactions to cortisol-21-acetate, cortisone, and corticosterone were 150%, 10%, and 7%, respectively. The accuracy, precision and correlation with RIA of this method were satisfactory for clinical application.
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