Determination of proteins in unconcentrated normal human urine

The protein content of repeatedly dialyzed urine is determined by measuring its absorbancy at 210 nm.     

The localization of proteoglycans and glycoproteins in the hyaline cartilage

Summary The innervation of the cornea of newborn (two day old) and adult rats was investigated using glyoxylic-acid-induced fluorescence (GIF) for catecholamines and subsequent acetylcholinesterase reaction.Fluorescent nerves were observed around the limbal vessels and in the pericorneal nerve plexus, from which they branched towards the central parts of the cornea. The fluorescent corneal nerves were either nonvaricose or had varicosities at intervals of 10 micra. When the animals had been pretreated with nialamide, noradrenaline and propranolol, some fluorescent branching nerve terminals with numerous varicosities also appeared. All fluorescent nerves disappeared two days after ipsilateral superior cervical sympathectomy.When the acetylcholinesterase (AChE) reaction was performed subsequently to the GIF reaction the following nerve types could be identified: 1. nerves containing both catecholamine (CA) fluorescence and AChE, 2. nerves containing only AChE.     

Attempt to isolate antierythrocyte antibodies by the method of affinity chromatography. I. IgM--complete antibodies

It follows from the results of the present study that erythrocyte stromata can serve as a suitable insoluble immunosorbent of complete antibodies. When using optimum conditions, such as laboratory incubation timperature, suitable buffers for both washing the column prior to absorption and for elution, it is possible to obtain, by means of the chromatographic column with 0.5 ml of erythrocyte stromata suspended in 40 g of silica glass beads (ballotini), fractions devoid of complete antibodies and subsequently to obtain the latter in purified form by elution. In complete anti antibodies, the elution yield amounted to an average of 35% of the initial amount of antibodies. Although the elution yield is not high and non-specific absorption has been demonstrated, the method can be recommended for the isolation of complete antibodies and thus for the production of monospecific sera because it is easy, rapid and inexpensive due to the regeneration ability of the mentioned columns and the relatively long duration of the immunoabsorption capacity of the incorporated erythrocyte stromata.     

Stereotyped behaviour after cholinergic, but not dopaminergic, stimulation of the substantia nigra in rats

Stereotyped behaviour of the rat was measured after intracerebral drug application in an objective and quantitative way by means of a new method developed in this laboratory. Bilateral intranigral injection of apomorphine /APO/, a specific dopaminergic agonist, did not evoke any signs of stereotyped behaviour. Also ineffective was the application of APO in the amygdaloid nucleus. Dopaminergic blockade of the substantia nigra by topical application of triperidol, a potent dopaminergic antagonist, failed to influence the stereotypy elicited by systematic APO administrationDirect cholinergic stimulation of the substantia nigra with carbachol resulted in a dose-related stereotyped behaviour not distinguishable by sight from that evoked by systematic APO administration. The effect of intranigral carbachol was antagonized by a previous intraperitoneal injection of 10 mg/kg of atropine. Stereotypy could easily be produced also by intracaudate application of APO. Topical triperidol blockade of the caudate nucleaus prevented the stereotypy caused by intraperitoneal application of APO.It is concluded that at least a part of nigral neurons cannot be directly excited by apomorphine. However, they can be excited by carbacol and seem thus, to contain muscarinic receptors. The stimulation of these receptors results in an excitation of the neurons involved and produces marked stereotyped behaviour.     

Teucrins H1-H4, novel clerodane-type diterpenes from Teucrium hyrcanicum

The structure and stereochemistry of teucrins H1, H2, H3 and H4, four novel diterpenes from Teucrium hyrcanicum have been established by detailed studies of the PMR and ¹³C NMR spectra, CD results and chemical transformations. Teucrin H4, a minor constituent is shown to possess a unique stereochemistry hitherto not reported for related diterpenoids.     

Isolation and characterization of wheat peroxidase isoenzyme B1

Two pure peroxidase isoenzymes B1 and D4 were isolated from the upper parts of 10-day-old wheat seedlings by means of gel and ion-exchange chromatography. Their MWs were 85000 and 24000 respectively. B1 was unstable and under various conditions it was converted to another isoenzyme, electrophoretically identical with D4. B1 contains about 40% of neutral sugars: 17.2% arabinose, 15.3% galactose, 5% glucose and traces of mannose. D4 is free of neutral sugars. None of the isoenzymes contained amino sugars. B1 oxidizes ferulic and p-coumaric acids. This oxidation has two pH optima of 4.4 and 5.4–5.6 and is inhibited by high concentrations of substrates, cyanide and azide. B1 oxidizes IAA in the presence of phenolic cofactor and Mn²⁺ ions. IAA oxidation has two pH optima of 4.5 and 5.6 and is inhibited by high substrate concentration, cyanide and azide, and by a number of indole derivatives. The main products of IAA oxidation are 3-methyleneoxindole and indole-3-methanol. o- and p- diphenols induce a lag period prior to IAA oxidation. Ferulic acid is oxidized during this lag period, probably to a dimer. B1 is able to produce H₂O₂ from oxygen. Mn²⁺ ions, a phenolic cofactor and an electron donor (IAA or NADH) are needed. B1 oxidizes α-keto-γ- methylmercaptobutyric acid to ethylene. D4 has a low peroxidatic activity and is inactive as an IAA oxidase. Thus B1 is probably an active cell wall-bound peroxidase isoenzyme, whereas D4 is its decomposition product.     

Isolation of somatic hybrids by cloning Nicotiana heterokaryons in nurse cultures

Fusion of Nicotiana knightiana Goodsp. and kanamycin resistant Nicotiana sylvestris Speg. et Com. protoplasts was induced by polyethylene glycol treatment. Heterokaryons were isolated by micropipette and transferred to nurse cultures of albino cells. Colonies originating from the heterokaryons could subsequently be distinguished by their green colour. The somatic hybrid nature of four such colonies was confirmed by isoenzyme pattern, kanamycin resistance and restored morphogenic potential. An additional kanamycin resistant line with characteristic Nicotiana knightiana isoenzymes was also found indicating that the drug resistance in the kanamycin resistant parent is under cytoplasmic control.Abbreviations ADH alcohol dehydrogenase - Nk Nicotiana knightiana     

Rhesus monkey micro mixed lymphocyte and mitogen reactivity: Optimal conditions and normal variability

Optimal conditions for the rhesus monkey micro mixed lymphocyte system with multiple automated harvesting of samples were evaluated. Parameters studied were cell concentration, length of culture period, methods of inactivation of cell populations, supplementation of media, type of culture plates, and changes in the reactivity of cells from individual animals over an extended time period. This work was supported in part by Portland Veterans Administration Hospital, Portland, Oregon, and the General Research Support Branch of the U.S. Public Health Service Grant RR00163, the Bureau of Medicine and Surgery, Navy Department, Work Unit No. M4318. 01.007ABG2. The opinions or assertions contained herein are the private ones of the authors and are not to be construed as official or reflecting the views of the U.S. Navy Department or the Naval service at large. The animals used in this study were handled in accordance with the provisions of Public Law 89–54 as amended by Public Law 91–579, “Animal Welfare Act of 1970,” and the principles outlined in the “Guide for the Care of Laboratory Animals,” U.S. Department of Health, Education, and Welfare Publication No. (NIH) 73-23.     

Effect of “osmotic stress” on protein and nucleic acid synthesis in isolated tobacco protoplasts

The incorporation of labeled precursors into RNAs and proteins of isolated tobacco (Nicotiana tabacum L.) leaf protoplasts decreases with increasing osmotic pressure in the incubation medium. The incorporation of precursors into RNA and proteins is linear for 15–18 h after the isolation of the protoplasts, irrespective of the osmolarity of the culture media. The uptake of precursors is also affected by the osmolarity of the medium. However, the osmotic stress-induced inhibition of incorporation of precursors into RNA and proteins is also apparent if the differences in uptake are taken into consideration in the calculation. Incorporation of ³²P into TMV-RNA is also inhibited by osmotic stress. As assayed by the double labeling ratio technique, osmotic stress has less unequivocal effect on TMV protein synthesis.Abbreviations PP protoplast - RNase ribonuclease - rRNA ribosomal ribonucleic acid - SDS sodium dodecyl sulfate - SSC 0.1 M Na-acetate in 0.15 M NaCl - TCA trichloroacetic acid - TMV tobacco mosaic virus     

Interaction of intracellular proteases and immune mechanisms

Intracellular proteases play an important role both in nonspecific and specific immune reactions. Results concerning the interaction of these enzymes with phagocytic factors responsible for killing microorganisms are presented. In inflammatory foci, proteases released from destroyed leukocytes have been found to modify the function of antibodies present. They particularly reduce their complement fixation activity. Macrophages with differing proteolytic activity demonstrated diverse effects on the formation of antibodies tested by Jerne's plaque technique in young rabbits immunized with sheep erythrocytes. Whereas alveolar macrophages diminished antigen immunogenicity, peritoneal macrophages enhanced it.