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At least 37 records · Page 2Linked to original sources

PubMed1978

Changes in fluorescence of 8-anilino-1-naphthalene sulfonate after bacteriophage T5 infection of Escherichia coli. Initial fluorescence rise coincides with onset of rubidium efflux.

Escherichia coli cells pre-loaded with 86Rb+ begin to lose 86Rb+ immediately after phage T5 addition. The loss proceeds with negative-exponential (first-order) kinetics for up to approximately 15 min after phage addition. The constant which characterizes the rate of loss increases with increasing numbers of infecting phage per cell. It is known that anaerobic, fermenting cells of E. coli show a two-step increase in 8-anilino-1-naphthalene sulfonate (ANS) fluorescence upon infection with bacteriophage T5; the first rise begins immediately upon phage addition, the second 6 min later. The onset of 86Rb+ release, therefore, is correlated with the first fluorescence rise with respect to timing and response to the multiplicity of infection.

Aerobiosis↗

PubMed1976

Fluorescence and the structure of proteins. XXII. Fluorescence of aminotyrosyl residues formed in ribonuclease A.

1. Tyrosyl residues on ribonuclease A were nitrated with tetranitromethane and then reduced to aminotyrosyl residues. By variation of reaction conditions and degree of exposure of tyrosyl residues it was possible to convert from 1 to all 6 tyrosyl to aminotyrosyl residues. 2. At the lower levels of 1-3 aminated tyrosyl residues/molecule the change in conformation seemed minor and 70% of the enzymatic activity was retained. When the three buried tyrosyl residues or all six residues were aminated only 5% of the enzymatic activity was retained. 3. Titration data, susceptibility to urea denaturation, and fluorescence characteristics indicated that some of the aminotyrosyl residues were buried in the interior and others were exposed on the surface of the protein. On the basis of the activation/emission wavelengths it was possible to distinguish buried (288/320 nm) and exposed (288/365-395 nm) aminotyrosyl residues as well as exposed tyrosyl residues (275-305 nm). 4. The modification of specific tyrosyl residues on a protein to aminotyrosyl residues appears to have some promise for observation of changes in environment of the residues that accompany various conformation changes by monitoring the fluorescence.

Amines↗

PubMed1979

Application of a high-performance liquid chromatographic fluorescence method for the rapid determination of alpha-tocopherol in the plasma of cattle and pigs and its comparison with direct fluorescence and high-performance liquid chromatography-ultraviolet detection methods.

High-performance liquid chromatography is used to develop a sensitive method for the determination of tocopherol levels in the plasma of cattle and pigs. This method is compared with a similar method using UV detection and one using direct fluorescence determination of tocopherol. Finally a double injection technique used in conjunction with fluorescence detection is shown to enhance the rate of analysis of the tocopherol levels in bovine plasma extracts.

Animals↗

PubMed1977

Comparison of rabies humoral antibody titers in rabbits and humans by indirect radioimmunoassay, rapid-fluorescent-focus-inhibition technique, and indirect fluorescent-antibody assay.

Rabies humoral antibodies were induced in eight New Zealand rabbits by a single intramuscular injection of inactivated suckling mouse brain rabies vaccine. The primary response to immunization was measured in blood samples taken at selected intervals for 6 months. The anamnestic response was measured in blood samples obtained 2 weeks after the rabbits received a booster immunization. The humoral antibody concentrations were measured by the rapid-fluorescent-focus-inhibition technique (RFFIT), indirect fluorescent-antibody assay (IFA), and indirect radioimmunoassay (RIA). The maximal neutralizing antibody titers as measured by RFFIT were attained by the 4th week and persisted into the 24th week. After booster immunization the antibody response was almost 10-fold higher than the highest level attained in the primary response. The antibody levels as measured by IFA and RIA were similar, but the titers as measured by either procedure were almost 10-fold lower than those determined by RFFIT. After booster immunizations the antibody levels, as measured by IFA and RIA, were three- and sixfold higher, respectively, than the maximal levels attained in the primary response. Twenty-two human serum specimens were tested by the same serological procedures, with disparate results. Both RIA and RFFIT effectively differentiated antirabies-positive sera from antirabies-negative sera.

Animals↗

PubMed1976

Influence of various fluorescent and non-fluorescent labels on the kinetics of the complex formation of alpha-chymotrypsin with basic pancreatic trypsin inhibitor (Kunitz).

The association of alpha-chymotrypsin with basic pancreatic trypsin inhibitor was studied using extrinsic signals produced by fluorescent and nonfluorescent labels. The reactive dyes were covalently bound to the proteins in the complexed state, in which the binding region was protected. The signals were sufficiently large to measure the complex formation at protein concentrations of 10(-9)M by fluorescence and down to 10(-6)M by absorption. Therefore, the association and dissociation could be followed over a broad range of concentration. Good correspondence was observed between data which were obtained with different labels and with published values for the unlabeled proteins. Existing differences could be explained by different buffer conditions used by the different authors. Also the pH dependence of the dissociation rate constants was essentially unaltered by the introduction of the labels. The large signals allowed a direct measurement of the equilibrium constants of dissociation, even at high pH, at which they are in the range of 10(-8)M. The experimentally determined binding constants were in agreement with those calculated from the rate constants. The temperature dependence of the binding constants revealed a small positive and pH-dependent enthalpy change [deltaHo = 4.0 kcal/mol (16.7 kJ) at H 7.0[. The results prove that the labeling can be performed in such a way that the equilibrium and kinetic parameters of the system studied are not significantly influenced.

Affinity Labels↗

PubMed1978

[The differentiation of clostridial microcultures on nuclepore-filters by fluorescent antibodies as a rapid screening test for spores. II. Application of fluorescent antibodies (author's transl)].

The study deals with the question of whether a combination of microcolony-cultivation on polycarbonate-membrane-filters with the fluorescent-antibody-method will lead to a rapid-diagnosis method for clostridial-spores. Formaldehyd-inactivated vegetative cells of eight different clostridial-strains (Table 1) were used for the immunisation of rabbits according to the method published by Böhm and Strauch (5). The results of the immunisation procedure, developed for aerobic-sporeformers, showed, that satisfactory results could not be obtained in all cases (Table 2). The eight conjugates were tested against microcolonies of 32 strains of different clostridial-species on membrane-filters, produced by the technique described in the first communication. The results showed (Table 4) that differentiation was always possible when homologous antisera were used. In most cases, in order to differentiate between the various species, conjugates of several serotypes had to be employed together. However due to their uniform antigenic nature, Cl. feseri and Cl. tetani could both be identified by using one strain for each to immunize. With clostridial-spores the species could be identified within a period of 6 to 12 hours by using the membrane-filter-fluorescent-antibody technique.

Antibodies, Bacterial↗

PubMed1977

Fluorescent probes for antibody active sites. II. Further studies on two groups of anti-MANS antibodies with significantly different effects on MANSamide fluorescence produced by a single rabbit.

As described previously, a single rabbit (No. 13) produced two groups of anti-MANS antibodies with significantly different effects on MANSamide fluorescence. These groups, anti-MANS antibody (415 nm) and anti-MANS antibody (465 nm), had emission maxima at 415 nm and 465 nm, respectively. This paper deals with the immunochemical properties of the two groups of antibodies (1) The results of fluorometric titration of these groups with MANSamide indicated that anti-MANS antibody (465 nm) had a higher affinity for MANSamide than did anti-MANS antibody (415 nm). (2) More than 70% of anti-MANS antibody (415 nm) was adsorbed on Sepharose covalently coupled with bovine serum albumin, which was used as a carrier during immunization, while a hlaf of anti-MANS antibody (465 nm) was adsorbed. (3) Only one of the groups, anti-MANS antibody (465 nm), had precipitating activity toward MANS-rabbit serum albumin used as a test antigen.

Anilino Naphthalenesulfonates↗

PubMed1976

Effects of ATP and magnesium ions on the fluorescence of harmala alkaloids. Restrictions for the use of harmala alkaloids as fluorescent probes for (Na+ + K+)-ATPase.

1. Harmine and harmaline were investigated as potentially useful fluorescent inhibitors of (Na+ + K+) activated ATPase. 29 From spectroscopic measurements both compounds were shown to form 1 : 1 complexes with ATP, the dissociation constants being 0.65 mM and 1.83 mM for harmine and harmaline respectively. Addition of Mg2+ and enzyme further affected these equilibria. 3. Although it was possible to demonstrate a competitive effect of harmine at the sodium-loading site of the enzyme, other inhibitory effects, including inhibitions of ouabain binding and the ouabain-insensitive ATPase were found. 4. It was concluded that the harmala alkaloids can inhibit (Na+ + K+)-activated ATPase in a complex way involving both Na- and ATP-binding sites. This severely limits their usefulness as spectroscopic probes.

Adenosine Triphosphatases↗

PubMed1979

Separation of mouse T cell subsets by a fluorescent activated cell sorter using fluorescence-labeled peanut agglutinin.

Nylon non-adherent spleen T cells obtained from concanavalin A-injected mice were labeled with fluorescein-labeled peanut agglutinin and separated into peanut agglutinin-positive (PNA+) cells and peanut agglutinin-negative (PNA-) cells by a fluorescent activated cell sorter. PNA+ cells were found to exert marked suppressive effect on primary anti-sheep red blood cells antibody response, but PNA- cells did not affect the antibody response. From these results and from the sugar-binding specificity of PNA, suppressor T cells are supposed to possess abundant galactosyl residues exposed on the cell surface, which are not masked by sialyl residues.

Animals↗

PubMed1978