Search

The phenomenon of low initial growth rates in atomic layer deposition (ALD) of various oxides on HF-treated (thus H-terminated) Si(001) substrates, which is termed the incubation effect or incubation period, can be effectively avoided by use of OH-terminated Si(001) substrates. Two ways of preparing OH-terminated Si(001) were devised in this work: one from atomically clean Si(001) and the other from H-terminated Si(001). The effect of reducing the incubation period was confirmed in the ALD process of aluminum oxide (Al2O3) thin films in which trimethylaluminum and water were used as sources of aluminum and oxygen, respectively. The use of OH-terminated Si(001) substrates has another beneficial effect of producing thin films with very smooth surface morphology. We propose the use of OH-terminated Si(001) substrates for growing thin films of many other metal oxides that have shown the incubation period on H-terminated Si(001) substrates.

Antibodies against blood group substance T in normal human sera consist of two types, depending upon whether or not the molecules remain immunochemically active at 37 degrees C. The two agglutinins were isolated by means of T antigen-coupled Sepharose 48 affinity chromatography. Temperature-sensitive agglutinin was eluted from the affinity column at 37 degrees C, while temperature-independent agglutinin remained bound to the antigen. Subsequently, the latter was dissociated from the column in the presence of 2.5 M MgCl2. Examination of six normal sera revealed that the levels of temperature-independent agglutinin was about twice that of temperature-sensitive agglutinin. More than 90% of anti-T agglutinins in normal sera were of the IgM class, in 8 of 11 samples studied. No direct relationship appeared to exist between anti-T titers and blood type. Thermal effects on hemagglutination, at least for anti-T agglutinin interacting with T antigen bearing red cells, is primarily due to the molecular characteristics of the agglutinin and not to the conformational change of the red cell membranes.

A specific and sensitive radioimmunoassay was developed to investigate the immunochemical properties of serum anti-T antibodies. Evidence was presented that the radioimmunoassay can measure non-agglutinating as well as agglutinating anti-T antibodies, while the hemagglutination method can detect only the latter type. Although the hemagglutination method failed to detect anti-T agglutinins in cord sera, the radioimmunoassay results demonstrated the presence of non-agglutinating antibodies. Presence of 1% polyethylene glycol potentiated some of the originally nonagglutinating antibodies to produce a hemagglutination reaction. Immunochemical characteristics associated with the human anti-T antibodies are discussed.

Anti-lactose antibodies induced in a horse with a bacterial vaccine were specifically purified and separated into IgM adn 7 S populations. These were further fractionated by liquid preparative isoelectric focusing. The fractions thus obtained were characterized by their affinity for a monovalent lactose-containing hapten and the deviation from homogeneous binding. In the IgM population there was a major component which exhibited the maximum affinity observed in the IgM fractions and functional homogeneity with a K 0 value of 4 × 10 5 M −1 . It was presumably a monoclonal product which persisted as the major IgM antibody during, at least, an 8-week period, from the 6th week to the 14th week after initial immunization. The emergence of this component is regarded as the result of an early maturation process. Another, non-homogeneous IgM fraction appeared to undergo maturation during the 8-week period but did not attain the affinity of the major component. The 7 S population gave rise to several major isoelectric components one of which was isolated in a functionally homogeneous form. All of the 7 S fractions exhibited a striking increase in affinity during the 8-week period and reached a common value of K 0 of 2 × 10 6 M −1 . This result provides further evidence for the selective limitation of the affinity of IgM antibody. The binding results with functionally homogeneous IgM component also demonstrate the functional equivalence and independent reactivity of the 10 combining sites of the IgM molecule. Finally, the disparity between the maximum affinities for the IgM and 7 S antibodies raises a question about the notion of an IgM -7 S switch. However, the occurrence of such a transition can be made compatible with our results by introducing special features for this process.

The nature of the pigment-pigment, pigment-protein, and protein-protein interactions in the bacteriochlorophyll-protein complex (BChl-P) isolated from a green photosynthetic bacterium Chloropseudomonas ethylicum has been investigated by means of absorption spectrum, optical rotatory dispersion, and circular dichroism. Pure bacteriochlorophyll in appropriate compositions of methanol-water mixture has been used as the model system to understand the state of the pigment molecules in the complex. There is a strong interaction between the pigment molecules in the BChl-P complex at neutral pH, but the interaction is very weak in both the Bph-P and the alkaline form of the complex. Despite significant structural modifications at acid and alkaline pH values, the pigment molecules are still attached to the protein moieties. Urea (8 m ) or 0.8% Triton X-100 do not produce any change in the conformational state of the BChl-P complex at, neutral pH, but they drastically alter the structures of the Bph-P and the alkaline forms of the complex. Hence, the BChl-P complex possesses the most compact and stable conformation at neutral pH, but becomes less stable at either of the extreme pH values. The results are discussed in view of the factors that are essential for stabilizing the pigment-protein relationship in the photosynthetic apparatus.

The copolymer poly Glu42Lys28Ala30 was crosslinked by S–S bridges by reaction with butyrothiolactone followed by air oxidation of the mercaptan covalent adduct. The modified copolypeptide was more compact hydrodynamically, more resistant to the structurally disruptive effects of urea, and able to bind dye cations with much increased avidity.

The interactions of anionic, cationic, and nonionic detergents with bacteriochlorophyll-protein (Bchl-P)3 complex isolated from a green photosynthetic bacterium Chloropseudomonas ethylicum have been examined. The effects of the detergents on the Bchl-P complex have shown a considerable variation depending on the type of the polar head-group and the nonpolar group of the detergent molecules. The anionic detergents, sodium dodecyl sulfate and sodium dodecyl benzene sulfonate, acting on Bchl-P, separate bacteriochlorophyll from the protein moiety and pheophytinize the Bchl by ligand bonding interaction. Pure Bchl is also converted into bacteriopheophytin by SDS. The interaction of a cationic or nonionic detergent with Bchl does not produce Bph. The Bchl-P complex is dissociated into Bchl and protein component by the cationic detergent, HTAB, or by a mixed solution of non-ionic detergent, Triton X-100, and 8 m urea. The ORD spectrum of Bchl-P in the uv range suggests that a significant change in protein conformation has taken place as a result of the detergent actions. Probable mechanisms for the combined action of Triton and urea on the complex are discussed.

Pleural and peritoneal fluids from humans with pathological diseases were examined for the presence of carcinoembryonic antigen-like substances (CEA-LS). Among eight samples tested by a solid phase radioimmunoassay, two pleural fluids and one peritoneal fluid showed significantly elevated CEA-activity. The substances responsible for the CEA-activity were isolated by perchloric acid-extraction followed by two successive Sephadex G-200 chromatography into two pools, Pool I (PI) and Pool II (PII). According to their sedimentation properties, PII was slightly smaller than CEA from tumor tissue-extract (CEA-TTE), while PI was larger than CEA-TTE and approximately twice the size of PII. Micro-double diffusion and antibody binding studies showed that CEA-LS possessed identical antigenic determinants as CEA-TTE, which did not cross-react with normal colon antigen (NCA).

The ultracentrifugation technique utilizing Airfuge was employed to determine the intrinsic association constant of mouse monoclonal IgM antibodies to human hepatitis B surface antigen (HBsAg). The IgM antibody was reduced to Fab fragments by tryptic digestion in the presence of mercaptoethanol, followed by two passages over a Sephacryl S-200 column. The centrifugation conditions were optimized to sediment Fab-HBsAg complexes, leaving the unbound Fab in the supernatant. The analyses of the binding data by Scatchard and Sips methods yielded good agreement, and the intrinsic association constants ranged from 0.13 to 5.3 X 10(7) (l/mol) for three IgM antibodies. The heterogeneity indices deduced from the Sips analysis were 1.0. The present protocol would allow determination of the intrinsic association constant for the binding of a multivalent antibody to a comparatively large antigen bearing multiple combining sites.

An attempt has been made to evaluate the maximum affinity of anti-lactoside antibody by the use of a large haptenic group (SLAC-lysyl), formula weight 797, containing the p -azophenyl- β -lactoside group as its terminal portion. Its synthesis and that of a number of intermediate compounds are described and their identity established. The preparation and characterization of the corresponding ligand, formula weight 853, used for the measurement of affinity are given, including its synthesis in tritiated form. The immunization of a horse with an immunogen consisting of multiple SLAC groups coupled to poly- L -lysine SLAC-PLL) produced IgG antibodies of functional homogeneity with an association constant of the order of 10 8 M −1 . Bleedings taken over a period of 6 weeks to 25 months following the primary immunization yielded antibody populations which showed no systematic and only minor variations of affinity. In the case of the rabbit, SLAC-PLL proved unsuitable as an immunogen for the indication of anti-SLAC antibody. However SLAC coupled to bovine-γ-globulin was an effective antigen. The rabbit anti-SLAC antibody, obtained after 5 weeks of immunization, also showed functional homogeneity with association constants of the order of 10 7 M −1 .

Immunization of a horse with a bacterial vaccine of Streptococcus faecalis, strain N led to the sustained and concomitant production of IgM and 7 S antibody with anti-lactoside specificity. Restriction of heterogeneity was apparent in both mol. wt groups with the IgM population exhibiting three major components when analyzed by isoelectric focusing. Equilibrium dialysis studies of the specific binding of a lactoside hapten, p(p-dimenthylaminophenylazo)-phenyl-β-lactoside (Lac dye), were carried out with three preparations of purified IgM antibody. The intrinsic association constants (K0) ranged between 1 × 105M−1 and 2 × 105M−1 for bleedings taken from 6 weeks to 14 weeks after the initiation of immunization. The availability of 10 binding sites per IgM molecule was clearly evident with no indication of intramolecular heterogeneity although substantial deviation from homogeneous binding was observed. Conversion of the pentameric IgM molecule to the monomeric form resulted in identical binding behavior. The 7 S antibody fraction gave a maximum value of K0 of 8 × 105M−1. The binding of lactose by IgM yielded a value of K0 which was 24-fold smaller than that for the Lac dye. Thus, the nonspecific contribution of the aglycoside, arising presumably from its hydrophobic character, to the free energy of complex formation was - 1·8 kcal/mole of hapten. The temperature dependence of the IgM binding of the Lac dye demonstrated that the enthalpy contribution was primarily responsible for the affinity. From this result it was inferred that the specific binding of lactose involves multiple hydrogen bonds, perhaps as many as 10.