scholarly journals Isolation of bacteria from artificial bronchoalveolar lavage fluid using density gradient centrifugation and their accessibility by Raman spectroscopy

2021 ◽  
Author(s):  
Christina Wichmann ◽  
Petra Rösch ◽  
Jürgen Popp
Keyword(s):  
Raman Spectroscopy ◽  
Bronchoalveolar Lavage ◽  
Density Gradient ◽  
Bronchoalveolar Lavage Fluid ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Lavage Fluid ◽  
Biochemical Properties ◽  
Low Concentrations ◽  
A Cell

AbstractRaman spectroscopy is an analytical method to identify medical samples of bacteria. Because Raman spectroscopy detects the biochemical properties of a cell, there are many factors that can influence and modify the Raman spectra of bacteria. One possible influence is a proper method for isolation of the bacteria. Medical samples in particular never occur in purified form, so a Raman-compatible isolation method is needed which does not affect the bacteria and thus the resulting spectra. In this study, we present a Raman-compatible method for isolation of bacteria from bronchoalveolar lavage (BAL) fluid using density gradient centrifugation. In addition to measuring the bacteria from a patient sample, the yield and the spectral influence of the isolation on the bacteria were investigated. Bacteria isolated from BAL fluid show additional peaks in comparison to pure culture bacteria, which can be attributed to components in the BAL sample. The isolation gradient itself has no effect on the spectra, and with a yield of 63% and 78%, the method is suitable for isolation of low concentrations of bacteria from a complex matrix.

Responses to Subchronic Inhalation of Low Concentrations of Diesel Exhaust and Hardwood Smoke Measured in Rat Bronchoalveolar Lavage Fluid

2005 ◽  
Vol 17 (12) ◽  
pp. 657-670 ◽  
Author(s):  
JeanClare Seagrave ◽  
Jacob D. McDonald ◽  
Matthew D. Reed ◽  
Steven K. Seilkop ◽  
Joe L. Mauderly
Keyword(s):  
Bronchoalveolar Lavage ◽  
Diesel Exhaust ◽  
Bronchoalveolar Lavage Fluid ◽  
Lavage Fluid ◽  
Low Concentrations

The subcellular distribution and properties of Crithidia sp. hydrolases with particular reference to pyrophosphate and orthophosphate monoester phosphohydrolases

1976 ◽  
Vol 54 (4) ◽  
pp. 365-381 ◽  
Author(s):  
J. McLaughlin ◽  
H. S. Injeyan ◽  
E. Meerovitch
Keyword(s):  
Density Gradient ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Hydrolytic Enzymes ◽  
Hydrolytic Activity ◽  
Cell Fractionation ◽  
Acid Hydrolases ◽  
Mitochondrial Marker ◽  
Fractionation Scheme ◽  
A Cell

A cell fractionation scheme was developed for studying the distribution of certain hydrolases, especially phosphohydrolases, in a Crithidia sp. (Trypanosomatidae). Whilst between 26–56% of the total cellular hydrolase activities were soluble (probably of flagellar pocket origin), a certain percentage, 5–40%, was sedimentable. A particulate fraction obtained after isopycnic density gradient centrifugation (ρ = 1.187–1.241), designated fraction FA/FB, was enriched in various acid hydrolases (relative specific activities 1.33–6.24) and displayed latent phosphohydrolase activities. The density gradient distributions of these hydrolytic enzymes were compared with reference to one another and malate dehydrogenase (mitochondrial marker). From the results obtained it appears that the sedimentable acid hydrolases of Crithidia are associated with a heterogeneous population of subcellular particles. Cytochemical observations on the FA/FB fraction supported this finding and revealed the association of acid phosphatase reaction product with subcellular elements resembling multivesicular bodies.A detailed study of the hydrolytic activity toward both ortho- and pyrophosphate substrates by the soluble, as compared to the FA/FB fractions, revealed the existence of distinct phosphohydrolase activities. It is proposed that these differences in properties are due to the existence of phosphohydrolase isoenzymes and that they in turn relate to a differentiation in the cellular digestive functions of Crithidia.

scholarly journals Heterogeneous distribution of glucose 6-phosphatase in rat liver microsomal fractions as shown by adaptation of a cytochemical technique

1973 ◽  
Vol 134 (1) ◽  
pp. 69-78 ◽  
Author(s):  
John A. Lewis ◽  
Jamshed R. Tata
Keyword(s):  
Phosphatase Activity ◽  
Density Gradient ◽  
Rat Liver ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Lead Phosphate ◽  
Sucrose Density Gradient Centrifugation ◽  
General Applicability ◽  
Heterogeneous Distribution ◽  
Low Concentrations

1. A novel technique for the subfractionation of rat liver smooth and rough microsomal fractions according to their content of glucose 6-phosphatase is described. This technique, based on the Gomori lead histochemical procedure, involves incubation of smooth and rough microsomal fractions with low concentrations of Pb(NO3)2 and glucose 6-phosphate. Control experiments, in which enzyme was assayed in the presence of various amounts of Pb(NO3)2 or in which microsomal fractions were reisolated after incubation with low concentrations of Pb(NO3)2 and glucose 6-phosphate, showed that lead does not interfere with glucose 6-phosphatase activity. 2. Discontinuous sucrose-density-gradient centrifugation of microsomal fractions which had previously been incubated with various amounts of Pb(NO3)2 and glucose 6-phosphate showed that it is possible to subfractionate both smooth- and rough-microsomal fractions into several bands, owing to a differential modification of the density of the microsomal vesicles by the trapping of lead phosphate within them. 3. When the material in the bands obtained by density-gradient centrifugation of incubated microsomal fractions was assayed for glucose 6-phosphatase activity, it was found that the modification of the density of the microsomal fractions was directly related to their relative enrichment in glucose 6-phosphatase activity. Control experiments, in which microsomal fractions were incubated with Pb(NO3)2 and glucose 6-phosphate and then treated with EDTA, showed that the subfractionation was not due to aggregation of microsomal vesicles, lead and glucose 6-phosphate. Thus the resolution of microsomal preparations into subfractions with different glucose 6-phosphatase activities is interpreted as indicating heterogeneity of glucose 6-phosphatase distribution in the microsomal vesicles. 4. Electron micrographs of both smooth- and rough-microsomal subfractions show deposits of lead phosphate within the microsomal vesicles. The frequency and extent of these deposits correlate with the different amounts of glucose 6-phosphatase activity measured biochemically. 5. The nature of the heterogeneous distribution of glucose 6-phosphatase is discussed and the more general applicability of the technique for studying membrane fractions containing a heterogeneous distribution of phosphatases is indicated.

SP-A, SP-B, and SP-C in surfactant subtypes around birth: reexamination of alveolar life cycle of surfactant

1994 ◽  
Vol 266 (4) ◽  
pp. L436-L447 ◽  
Author(s):  
A. Baritussio ◽  
A. Alberti ◽  
D. Quaglino ◽  
A. Pettenazzo ◽  
D. Dalzoppo ◽  
...  
Keyword(s):  
Life Cycle ◽  
Density Gradient ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Lavage Fluid ◽  
Surfactant Protein ◽  
Lamellar Bodies ◽  
Alveolar Surfactant

Transformations of surfactant after secretion are incompletely understood. To clarify them, we lavaged lungs in fetuses and in newborn rabbits, fractionated the lavage fluid by differential and density gradient centrifugation, and analyzed the distribution of surfactant protein (SP) phospholipids, SP-A, SP-B, and SP-C. Furthermore, we administered into trachea of newborn rabbits labeled surfactant and compared the alveolar clearance of SP-A, SP-B, SP-C and saturated phosphatidylcholine. We found that, in the fetus, secreted lamellar bodies contain all components of surfactant, except a small amount of SP-A. As breathing starts and new surfactant subtypes are generated, the proteins are mostly associated with dense subtypes, but SP-B and SP-C are especially concentrated in dense materials that contain minor amounts of phospholipids and SP-A. Furthermore, we found that, during breathing, alveolar surfactant is degraded into more than one type of remnant, that the lavage fluid contains a pool of SP-A not associated with membranes, and that SP-A, SP-B, and SP-C are all turned over at a faster rate than saturated phosphatidylcholine.

Uranyl Acetate and Rotary Shadowing to Increase the Contrast of Small Aggregated Virus Particles

1969 ◽  
Vol 27 ◽  
pp. 424-425
Author(s):  
Lee F. Ellis ◽  
Richard M. Van Frank ◽  
Walter J. Kleinschmidt
Keyword(s):  
Electron Microscopy ◽  
Tissue Culture ◽  
Density Gradient ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Uranyl Acetate ◽  
Interferon Inducer ◽  
Virus Particles ◽  
Staining Methods ◽  
Rotary Shadowing

The extract from Penicillum stoliniferum, known as statolon, has been purified by density gradient centrifugation. These centrifuge fractions contained virus particles that are an interferon inducer in mice or in tissue culture. Highly purified preparations of these particles are difficult to enumerate by electron microscopy because of aggregation. Therefore a study of staining methods was undertaken.

Identification of neutrophil chemotactic factors in bronchoalveolar lavage fluid of asthmatic patients

1997 ◽  
Vol 27 (4) ◽  
pp. 396-405 ◽  
Author(s):  
L. M. TERAN ◽  
M. G. CAMPOS ◽  
B. T. BEGISHVILLI ◽  
J.-M. SCHRODER ◽  
R. DJUKANOVIC ◽  
...  
Keyword(s):  
Bronchoalveolar Lavage ◽  
Bronchoalveolar Lavage Fluid ◽  
Lavage Fluid ◽  
Asthmatic Patients ◽  
Chemotactic Factors

The Combining Ability of Glycoproteins IIb, IIIa and Ca2+ in EDTA-Treated Nonaggregable Platelets

1983 ◽  
Vol 50 (04) ◽  
pp. 848-851 ◽  
Author(s):  
Marjorie B Zucker ◽  
David Varon ◽  
Nicholas C Masiello ◽  
Simon Karpatkin
Keyword(s):  
Density Gradient ◽  
Combining Ability ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Electrophoretic Pattern ◽  
Sucrose Density Gradient ◽  
Irreversible Loss ◽  
Sucrose Density Gradient Centrifugation ◽  
Crossed Immunoelectrophoresis ◽  
Triton X

SummaryPlatelets deprived of calcium and incubated at 37° C for 10 min lose their ability to bind fibrinogen or aggregate with ADP when adequate concentrations of calcium are restored. Since the calcium complex of glycoproteins (GP) IIb and IIIa is the presumed receptor for fibrinogen, it seemed appropriate to examine the behavior of these glycoproteins in incubated non-aggregable platelets. No differences were noted in the electrophoretic pattern of nonaggregable EDTA-treated and aggregable control CaEDTA-treated platelets when SDS gels of Triton X- 114 fractions were stained with silver. GP IIb and IIIa were extracted from either nonaggregable EDTA-treated platelets or aggregable control platelets with calcium-Tris-Triton buffer and subjected to sucrose density gradient centrifugation or crossed immunoelectrophoresis. With both types of platelets, these glycoproteins formed a complex in the presence of calcium. If the glycoproteins were extracted with EDTA-Tris-Triton buffer, or if Triton-solubilized platelet membranes were incubated with EGTA at 37° C for 30 min, GP IIb and IIIa were unable to form a complex in the presence of calcium. We conclude that inability of extracted GP IIb and IIIa to combine in the presence of calcium is not responsible for the irreversible loss of aggregability that occurs when whole platelets are incubated with EDTA at 37° C.

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