scholarly journals STUDIES ON OXIDATION AND REDUCTION BY PNEUMOCOCCUS

1924 ◽  
Vol 39 (3) ◽  
pp. 357-366 ◽  
Author(s):  
Oswald T. Avery ◽  
James M. Neill
Keyword(s):  
Yeast Extract ◽  
Room Temperature ◽  
Prolonged Exposure ◽  
Freezing And Thawing ◽  
Peroxide Formation ◽  
Cell Extracts ◽  
Repeated Freezing ◽  
Phosphate Solutions ◽  
Animal Tests

In the present paper methods have been described for the preparation of sterile extracts of pneumococci. These extracts may be obtained by dissolving the bacteria in broth cultures by means of bile, or by extraction of the cellular substances by repeated freezing and thawing of broth or saline suspensions of unwashed cells. Under special precautions these extracts may be passed through Berkefeld filters without loss of potency. In this procedure, as in all other manipulations incident to their preparation, the extracts should be protected as far as possible from contact with air. All extracts were proved sterile by cultural and animal tests. Sterile extracts of unwashed pneumococcus cells promptly form peroxide on exposure to air. Peroxide formation is almost as active in extracts aerated at 2°C. as in those exposed to the air at room temperature. Detectable amounts of peroxide may be produced by these cell extracts within the reaction range of pH 5 to 9, the optimal zone lying at reactions less acid than pH 6. The peroxide-forming activity of the extracts is gradually diminished by prolonged exposure to 55°C., and is completely destroyed by heating at 65°C. for 5 minutes. Cell extracts of pneumococci which have been thoroughly washed prior to extraction in salt or phosphate solutions exhibit no peroxide-forming activity. These extracts of washed cells may be activated by the addition of the cell washings, yeast extract, or muscle infusion.

scholarly journals Cultured bovine endothelial cells produce both urokinase and tissue-type plasminogen activators.

1982 ◽  
Vol 94 (3) ◽  
pp. 631-636 ◽  
Author(s):  
E G Levin ◽  
D J Loskutoff
Keyword(s):  
Molecular Weight ◽  
Endothelial Cells ◽  
Degradation Products ◽  
Tissue Type ◽  
Freezing And Thawing ◽  
Prolonged Incubation ◽  
Aortic Endothelial Cells ◽  
Cell Extracts ◽  
Bovine Endothelial Cells ◽  
Repeated Freezing

Cell extracts and conditioned media (CM) from cultured bovine aortic endothelial cells (BAEs) were fractionated by PAGE in the presence SDS, and plasminogen activator (PA) activity was localized by fibrin autography. Multiple molecular weight forms of PA were detected in both preparations. Cell-associated PAs had Mr of 48,000, 74,000, and 100,000 while secreted PAs showed Mr of 52,000, 74,000, and 100,000. A broad zone of activity (Mr 80,000-100,000) also was present in both cellular fractions. In addition, PAs of Mr 41,000 and 30,000 appeared upon prolonged incubation or repeated freezing and thawing of the samples, and probably represent degradation products of higher molecular weight forms. This complex lysis pattern was not observed when CM was subjected to isoelectric focusing. Instead, only two classes of activator were resolved, one at pH 8.5, the other at 7.6. Analysis of focused samples by SDS PAGE revealed that the activity at pH 8.5 resulted exclusively from the Mr 52,000 form; all other forms were recovered at pH 7.6. The activity of the Mr 52,000 form was neutralized by anti-urokinase IgG but was not affected by antitissue activator IgG indicating that it is a urokinaselike PA. The activities of the Mr 74,000-100,000 forms were not affected by anti-urokinase. They were blocked by antitissue activator suggesting that all the forms in this group were tissue-type PAs. The multiple forms of PA were differentially sensitive to inactivation by diisopropylfluorophosphate (DFP). Treatment of CM with 10 mM DFP for 2 h at 37 degrees C only partially inhibited the 52,000-dalton form. However, it completely inactivated the 74,000-dalton partially inhibited the 52,000-dalton form. However, it completely inactivated the 74,000-dalton PA. The activity of the Mr 100,000 form was not affected by this treatment, or by treatment with 40 mM DFP. Thus, cultured BAEs produce multiple, immunologically distinct forms of PA which differ in size, charge, and sensitivity to DFP. These forms include both urokinaselike and tissue-activator-like PAs. The possibility that one of these forms is a zymogen is discussed.

scholarly journals STUDIES ON OXIDATION AND REDUCTION BY PNEUMOCOCCUS

1924 ◽  
Vol 39 (4) ◽  
pp. 543-552 ◽  
Author(s):  
Oswald T. Avery ◽  
James M. Neill
Keyword(s):  
Methylene Blue ◽  
Molecular Oxygen ◽  
Yeast Extract ◽  
Phosphate Solution ◽  
Peroxide Formation ◽  
Hydrogen Acceptor ◽  
Intact Cells ◽  
Washed Cell ◽  
Cell Extracts ◽  
Methylene Blue Reduction

1. Sterile broth extracts of unwashed pneumococci, entirely free from living or intact cells, actively reduce methylene blue. 2. Sterile extracts prepared by extracting washed pneumococci in phosphate solution are unable by themselves to reduce methylene blue. Upon the addition of meat infusion or yeast extract, these washed cell extracts actively reduce methylene blue. 3. The system or systems in pneumococcus extracts responsible for methylene blue reduction are destroyed by exposure to temperatures practically identical with those which have been previously found to destroy the peroxide-forming activity of the same extracts. 4. It is suggested that peroxide formation and methylene blue reduction by pneumococcus extracts are functions of the same or closely related systems, the particular reaction induced depending upon whether molecular oxygen or methylene blue serves as hydrogen acceptor or oxygen donator.

Stability of varenicline concentration in saliva over twenty-one days at three storage temperatures

2021 ◽  
Author(s):  
Maria Novalen ◽  
Meghan J Chenoweth ◽  
Bin Zhao ◽  
Larry W Hawk, Jr. ◽  
Rachel F Tyndale
Keyword(s):  
Quality Control ◽  
Smoking Cessation ◽  
Room Temperature ◽  
Freezing And Thawing ◽  
Storage Duration ◽  
Non Invasive ◽  
Specimen Collection ◽  
Repeated Freezing ◽  
The Stability ◽  
Storage Temperatures

Abstract Introduction Varenicline is the most efficacious drug for smoking cessation; saliva varenicline concentrations can be useful for the evaluation of adherence in smoking cessation trials. Saliva is a useful non-invasive matrix for mail-in specimen collection, if stable. We investigated the stability of varenicline in saliva at different storage temperatures simulating the time it takes to mail in a sample. Methods We evaluated the concentrations of varenicline, nicotine, cotinine, 3’-hydroxycotinine and 3’-hydroxycotinine/cotinine (3HC/COT) ratio in quality control saliva samples (and after repeated freezing and thawing), and in smokers’ saliva samples, stored for up to 21 days at room temperature (~25°C), 4°C and -80°C. Results In saliva quality control samples, concentrations of varenicline, nicotine, cotinine, 3’-hydroxycotinine and 3HC/COT remained unchanged and showed little within-sample variation (CV≤5.5%) for up to 21 days at the three storage temperatures; they were also not altered after three thaw-freeze cycles. In smokers’ saliva, a significant main effect of storage duration, but not temperature, was observed for varenicline, cotinine and 3’-hydroxycotinine, but not for nicotine or the 3HC/COT ratio. However, these changes were within analytical (i.e. equipment) variation resulting in little within-sample variation (CV≤5.8%) for all analytes in smokers saliva. Conclusions Varenicline, the other analytes and the 3HC/COT ratio remained stable in saliva during storage for 21 days at all temperatures tested and after repeated freezing and thawing with only minor changes in concentration over time. These findings support the potential use of mail-in approach for saliva samples in varenicline smoking cessation clinical trials. Implications Assessing saliva varenicline concentrations can be useful for the evaluation of adherence in smoking cessation trials. Saliva is a non-invasive matrix suitable for mail-in specimen collection. This is the first investigation of stability of varenicline in saliva. Varenicline, nicotine, cotinine, 3’-hydroxycotinine and 3HC/COT were stable in saliva for up to 21 days at room temperature (~25°C), 4°C and -80°C, supporting the use of a mail-in approach for saliva specimen in smoking cessation trials.

scholarly journals ISOZYMES OF ACID PHOSPHATASE IN NORMAL AND CALMETTE-GUÉRIN BACILLUS-INDUCED RABBIT ALVEOLAR MACROPHAGES

1968 ◽  
Vol 128 (5) ◽  
pp. 1031-1048 ◽  
Author(s):  
S. G. Axline
Keyword(s):  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Alveolar Macrophages ◽  
Acid Phosphatase Activity ◽  
Enzyme Inhibitors ◽  
Freezing And Thawing ◽  
Ph Optimum ◽  
Lysosomal Fraction ◽  
Repeated Freezing ◽  
Major Acid

The acid phosphatase activity of normal alveolar and BCG-induced alveolar macrophages has been examined. Five electrophoretically distinct forms of acid phosphatase have been identified in both normal and BCG-induced macrophages. The acid phosphatases can be divided into two major categories. One category, containing four distinct forms, is readily solubilized after repeated freezing and thawing or mechanical disruption The second category, containing one form, is firmly bound to the lysosomal membrane and can be solubilized by treatment of the lysosomal fraction with Triton X-100. The Triton-extractable acid phosphatase and the predominant aqueous soluble acid phosphatase have been shown to differ in the degree of membrane binding, in solubility, in net charge, and in molecular weight. The two pre-dominant phosphatases possess identical pH optimum and do not differ in response to enzyme inhibitors. BCG stimulation has been shown to result in a nearly twofold increase in acid phosphatase activity. A nearly proportionate increase in the major acid phosphatase forms has been observed.

Effects of perturbations and stimulants on red raspberry (Rubus idaeus L.) seed germination

1997 ◽  
Vol 73 (4) ◽  
pp. 453-457 ◽  
Author(s):  
R. A. Lautenschlager
Keyword(s):  
Seed Coat ◽  
Temperature Fluctuations ◽  
Rubus Idaeus ◽  
Red Raspberry ◽  
Freezing And Thawing ◽  
Short Term ◽  
Repeated Freezing ◽  
Key Words Animal ◽  
Concentrated Sulfuric Acid ◽  
Seed Coats

Red raspberry (Rubus idaeus L.) seeds germinate only after seed coats are degraded. In nature this happens slowly. Seeds from recently collected fruit (fresh to four years old) germinated only after scarification of the seed coat by 20-minute soaking in concentrated sulfuric acid. Germination was not enhanced by: (1) short-term intermittent soaking, up to 81 hours, in dilute (0.01 normal) hydrochloric acid; (2) passage through the digestive tracts of bears, coyotes, or birds; (3) physical perturbations such as nicking, mechanical scarification, repeated freezing and thawing and/or four years of exposure in the field; (4) exposure to light; (5) increased temperatures or temperature fluctuations; or (6) addition of nitrogen (ammonium nitrate, urea). Key words: animal passage, germination, nitrogen, red raspberry, Rubus idaeus L., seed coat, seed weight, scarification, stratification

scholarly journals Safety evaluation of resveratrol as an active compound for drug-eluting cardiovascular implants

2019 ◽  
Vol 5 (1) ◽  
pp. 331-333
Author(s):  
Valeria Khaimov ◽  
Thomas Reske ◽  
Claudia Matschegewski ◽  
Niels Grabow ◽  
Thomas Eickner
Keyword(s):  
Prolonged Exposure ◽  
Animal Experiments ◽  
Safety Evaluation ◽  
Local Drug Delivery ◽  
Surrounding Tissue ◽  
Interesting Candidate ◽  
Drug Eluting ◽  
Cardiovascular Implants ◽  
Animal Tests

AbstractResveratrol is a member of stilbenoids with promising anti-atherosclerotic properties. This hallmark makes it an extremely interesting candidate for local drug delivery to damaged tissue adjacent to the implant in order to reduce implant-related complications. For the regulatory approval drug-eluting medical devices have to be thoroughly tested for safety, efficacy and interactions with the surrounding tissue, including tests for sensitization among others. Studies for sensitization help to estimate the risk for an allergic reaction upon prolonged exposure to a chemical compound. Due to increased social and regulatory demand for replacement of animal experiments by in vitro approaches a number of reliable predictive non-animal tests have been developed. Here, we assessed the skin sensitization potential of resveratrol by the direct peptide reactivity assay (DPRA), one of the first non-animal tests adopted by the OECD.

97 IMPROVED CRYOPRESERVATION OF DOMESTIC CAT SPERMATOZOA IN A SOY LECITHIN-BASED EXTENDER

2011 ◽  
Vol 23 (1) ◽  
pp. 153 ◽  
Author(s):  
M. M. Vick ◽  
H. L. Bateman ◽  
W. F. Swanson
Keyword(s):  
Sperm Motility ◽  
Room Temperature ◽  
Egg Yolk ◽  
Tris Buffer ◽  
Domestic Cat ◽  
Dna Integrity ◽  
Freezing And Thawing ◽  
Soy Lecithin ◽  
Acridine Orange Staining ◽  
Acrosome Integrity

Development of a chemically defined, plant-based cryopreservation media would reduce extender variability and the potential for transmission of zoonotic pathogens compared with traditional egg-yolk-based extenders. The objective of this study was to compare effects of egg yolk- and soy lecithin-based cryopreservation media and the temperature of glycerol addition on sperm parameters following freezing and thawing of domestic cat spermatozoa. Fresh semen was collected by manual stimulation on 3 separate occasions from 4 adult male cats. Each ejaculate was washed to remove seminal plasma, divided into 4 equal aliquots, and extended at room temperature in one of the following treatments: 1) TEST-egg yolk (Irvine Scientific Inc., Santa Ana, CA, USA) medium with 4% glycerol (EYG); 2) TEST-egg yolk, with 4% glycerol added after cooling to 5°C (EY); 3) TES-Tris buffer with soy-lecithin (1%) and 4% glycerol (SLG); and 4) TES-Tris buffer with 1% soy-lecithin, and 4% glycerol added after cooling to 5°C (SL). Sperm progressive motility (%) and rate of progressive movement (scale of 0–5) were evaluated at 0, 1, 3, 6, and 24 h post-thaw. Sperm capacitation (chlortetracycline staining), acrosome integrity (FITC-PNA staining), and DNA integrity (acridine orange staining) were assessed at 15 min post-thaw. Data were exponentially transformed to achieve normal distribution and then subjected to GLM analysis to determine effects of media and temperature of glycerol addition on sperm traits. At 0 and 1 h post-thaw, acrosome integrity, DNA integrity and % sperm motility did not differ (P > 0.05) among treatments. However, % sperm motility was greater in the soy-based media compared to egg yolk-based media at 3, 6, and 24 h post-thaw (Table 1; P < 0.05). A higher percentage of uncapacitated spermatozoa were present in soy-based compared to egg-yolk based cryopreservation media (63.9 ± 9.3 v. 51.2 ± 11.5, respectively; P < 0.05), regardless of temperature of glycerol addition. Finally, addition of glycerol at 5°C resulted in higher % sperm motility compared to room temperature at 6 and 24 h post-thaw in both medium types (Table 1; P < 0.05). Our results suggest that use of a chemically defined, soy-based medium improves long-term motility and capacitation status of frozen–thawed domestic cat spermatozoa compared with cryopreservation in a traditional egg yolk-based extender. Table 1.Motile spermatazoa and motility score at 3, 6, and 24 h

Simple method for preparing a concentration gradient of serum components by freezing and thawing

1991 ◽  
Vol 37 (7) ◽  
pp. 1225-1229 ◽  
Author(s):  
Tetsuo Hirano ◽  
Toshiaki Yoneyama ◽  
Hiroko Matsuzaki ◽  
Takainitsu Sekine
Keyword(s):  
Concentration Gradient ◽  
Clinical Laboratory ◽  
Freezing Temperature ◽  
Test Results ◽  
Freezing And Thawing ◽  
Serum Samples ◽  
Simple Method ◽  
Gradient Formation ◽  
Repeated Freezing ◽  
Serum Components

Abstract We created a simple method for obtaining a series of successively more-concentrated samples from a serum without changing the ratio of its components. We froze a pooled serum and then allowed it to thaw undisturbed. The serum components formed a gradient of increasing concentration from the top of the sample to the bottom. We found that (a) in test results, each fraction of serum in the gradient showed almost the same relative concentrations of components (i.e., inorganic and organic compounds, proteins, metals, and hormones), irrespective of atomic or molecular mass; (b) the concentration gradient depended on the thawing temperature but not on the freezing temperature; (c) when we thawed the frozen sample with centrifugation, the slope of the concentration gradient increased with increasing centrifugal force; (d) when the thawed sample was fractionated into 10 fractions from the top to the bottom, the original serum concentration was always maintained between the sixth and seventh fractions from the top; and (e) the concentration gradient became steeper with repeated freezing and thawing. By using this method, one can easily prepare serum samples at gradients of concentration useful in the clinical laboratory, although the mechanism of gradient formation is still unclear.

FREEZE-THAW EFFECTS ON GRANULAR STRUCTURE REORGANIZATION FOR SOIL MATERIALS OF VARYING TEXTURE AND MOISTURE CONTENT

1988 ◽  
Vol 68 (3) ◽  
pp. 485-494 ◽  
Author(s):  
S. PAWLUK
Keyword(s):  
Moisture Content ◽  
Lacustrine Sediments ◽  
Field Capacity ◽  
Granular Structure ◽  
Glacial Till ◽  
Microstructural Development ◽  
Clay Loam ◽  
Freezing And Thawing ◽  
Freeze Thaw ◽  
Repeated Freezing

Repeated freezing and thawing of glacial till cores of clay loam texture results in the formation of granic and metafragmic microfabrics. These units of fabric are best developed near the surface of cores kept at moisture levels between field capacity and saturation. Well-sorted lacustrine sediments with fewer voids tend to form banded fabrics. Many of the morphological features such as vesicles, metavughs and desiccation cracks commonly attributed to freeze-thaw processes are evident in all materials tested. Discrete units of fabric observed in this study are very similar to units of fabric observed in the Ah horizons of Black Chernozemic and Cryosolic soils. Results of this investigation strongly support earlier research which suggests that frost processes are major contributors to their microstructural development. Key words: Granic, freeze-thaw, microfabrics

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