NUTRITION OF ANIMAL CELLS IN TISSUE CULTURE: VIII. DESOXYRIBONUCLEIC ACID PHOSPHORUS AS A MEASURE OF CELL MULTIPLICATION IN REPLICATE CULTURES

1954 ◽  
Vol 32 (1) ◽  
pp. 319-326
Author(s):  
George M. Healy ◽  
Dorothy C. Fisher ◽  
Raymond C. Parker
Keyword(s):  
Horse Serum ◽  
Cell Multiplication ◽  
Desoxyribonucleic Acid ◽  
Present Communication ◽  
Animal Cells ◽  
Defined Media ◽  
Chick Embryo Extract ◽  
Material Good ◽  
Cell Strains

A chemical method of estimating the size of cell populations in replicate cultures has been devised for use with certain cell strains from the mouse and rat. The method is based upon the determination of desoxyribonucleic acid phosphorus (DNAP). Although the method may be used as an alternative to the procedure of counting isolated, stained nuclei in a hemocytometer, it actually supplements such measurements and provides useful information on the DNA-content per cell. The method consists of a modified Schmidt and Thannhauser separation of ribonucleic acid (RNA) and desoxyribonucleic acid (DNA) followed by the spectrophotometric estimation of the orthophosphate. The present communication describes the procedures in detail and reports recoveries of RNAP and DNAP from mixtures of highly polymerized nucleic acids and various types of biological material. Good correlation was found between nuclear counts and DNAP values in natural media (containing horse serum and chick embryo extract) and in synthetic (chemically-defined) media. Finally, the efficiency of the dispensing apparatus most frequently employed in preparing replicate cultures from continuously stirred, washed cell suspensions has been determined by measuring the DNAP content of serially dispensed samples.

NUTRITION OF ANIMAL CELLS IN TISSUE CULTURE: VIII. DESOXYRIBONUCLEIC ACID PHOSPHORUS AS A MEASURE OF CELL MULTIPLICATION IN REPLICATE CULTURES

1954 ◽  
Vol 32 (3) ◽  
pp. 319-326 ◽  
Author(s):  
George M. Healy ◽  
Dorothy C. Fisher ◽  
Raymond C. Parker
Keyword(s):  
Horse Serum ◽  
Cell Multiplication ◽  
Desoxyribonucleic Acid ◽  
Present Communication ◽  
Animal Cells ◽  
Defined Media ◽  
Chick Embryo Extract ◽  
Material Good ◽  
Cell Strains

A chemical method of estimating the size of cell populations in replicate cultures has been devised for use with certain cell strains from the mouse and rat. The method is based upon the determination of desoxyribonucleic acid phosphorus (DNAP). Although the method may be used as an alternative to the procedure of counting isolated, stained nuclei in a hemocytometer, it actually supplements such measurements and provides useful information on the DNA-content per cell. The method consists of a modified Schmidt and Thannhauser separation of ribonucleic acid (RNA) and desoxyribonucleic acid (DNA) followed by the spectrophotometric estimation of the orthophosphate. The present communication describes the procedures in detail and reports recoveries of RNAP and DNAP from mixtures of highly polymerized nucleic acids and various types of biological material. Good correlation was found between nuclear counts and DNAP values in natural media (containing horse serum and chick embryo extract) and in synthetic (chemically-defined) media. Finally, the efficiency of the dispensing apparatus most frequently employed in preparing replicate cultures from continuously stirred, washed cell suspensions has been determined by measuring the DNAP content of serially dispensed samples.

scholarly journals DETERMINATION OF INHERITED TRAITS OF H. INFLUENZAE BY DESOXYRIBONUCLEIC ACID FRACTIONS ISOLATED FROM TYPE-SPECIFIC CELLS

1951 ◽  
Vol 93 (4) ◽  
pp. 345-359 ◽  
Author(s):  
Hattie E. Alexander ◽  
Grace Leidy
Keyword(s):  
Population Size ◽  
Small Proportion ◽  
Growth Period ◽  
Cell Multiplication ◽  
Acid Fraction ◽  
Desoxyribonucleic Acid

Change of non-typable (R) strains of H. influenzae into the specific type of origin or new types (S) has been effected in vitro in a predictable fashion within a single 24 hour growth period, by a desoxyribonucleic acid-containing fraction isolated from type-specific cells of the type desired. Only a small proportion of the population of R cells are susceptible to the change induced in inherited characteristics by the desoxyribonucleic acid fraction. The data suggest that the number of susceptible cells present in any given population size varies with the specific type of origin of the R cells; a lesser degree of variation in different independent cultures of the same strain and population size has been demonstrated. The results suggest that the R H. influenzae cells which are susceptible to transformation arise as the result of mutation. It has been demonstrated that the reaction necessary for transformation takes place virtually immediately if susceptible cells are present. Furthermore it has been shown that the change which enables an R cell to form a colony of type-specific organisms has been completed within 15 minutes in an environment which does not permit cell multiplication.

scholarly journals Experimental Testing of a Size-Exclusion Chromatography Method Used for Evaluation of Molecular Parameters of Equine Anti-Ebola Immunoglobulin

2019 ◽  
Vol 19 (4) ◽  
pp. 261-267
Author(s):  
Е. Yu. Mishalova ◽  
E. V. Gordeev ◽  
V. N. Lebedev ◽  
S. A. Melnikov ◽  
S. A. Nimirskaya ◽  
...  
Keyword(s):  
Specific Activity ◽  
Experimental Testing ◽  
Horse Serum ◽  
Test Procedure ◽  
Size Exclusion ◽  
Array Detector ◽  
Test Procedures ◽  
Molecular Parameters ◽  
Hplc System

Haemorrhagic fever caused by the Ebola virus is a highly hazardous infectious disease with a mortality rate of 50– 90 %. Heterologous immunoglobulins with a high virus-neutralizing titer are an important element of the WHO-endorsed set of measures for emergency prevention and treatment of the disease. Specific activity of these products is largely determined by their fractional composition, and, in particular, by molecular mass distribution (MMD). The size-exclusion-high-performance liquid chromatography (SEC-HPLC) has traditionally been used for determination of the MMD of the target protein in human immunoglobulin-based products. The use of this method for evaluation of molecular parameters of heterologous immunoglobulin requires confirmation of its specificity, accuracy and precision, and establishment of the chromatographic system suitability criteria in the context of a new test object.The aim of the study was to test the applicability of the SEC-HPLC method to the assessment of molecular parameters of anti-Ebola immunoglobulin derived from horse serum.Materials and methods: three batches of purified equine anti-Ebola immunoglobulin were used in the study. Normal equine and human immunoglobulins of the IgG isotype were used as reference standards. The HPLC test procedures described in the European Pharmacopoeia 9.6 and State Pharmacopoeia of the Russian Federation, 14th ed., were used for determination of monomers and other immunoglobulin fractions. An Agilent 1260 Infinity (Agilent, USA) HPLC system with a diode array detector and an Agilent Bio SEC-3 HPLC column were used for quality evaluation of the tested products.Results: the resolution factor between IgG monomer and dimer peaks (1.69 and 2.10), and the chromatographic column efficiency (>2000) make it possible to use the SEC-HPLC system for evaluation of molecular parameters of heterologous immunoglobulin. The study demonstrated reproducibility of the test procedure.Conclusions: the study confirmed the applicability of the SEC-HPLC procedure for evaluation of molecular parameters of anti-Ebola immunoglobulin derived from horse serum. It demonstrated the compliance of the purified immunoglobulin to the national and international quality requirements in terms of «Molecular parameters».

scholarly journals Amplification of Color Yield Obtained Using a Chromogenic Substrate for Determining Plasminogen Activator

1977 ◽  
Author(s):  
Robert B. Friedmaa ◽  
Hau C. Kwaan ◽  
Marija Szczecinski
Keyword(s):  
Absorption Maximum ◽  
Enzyme Assay ◽  
Amide Bond ◽  
Chromogenic Substrate ◽  
Present Communication ◽  
Red Color ◽  
Enzyme Digest ◽  
Color Yield ◽  
Ammonium Sulfamate

A synthetic chromogenic substrate has recently been reported for use in the specific measurement of urokinase. This material (Chromozym UK, marketed by Pentapharm, Ltd., Basle) is a synthetic tripeptide linked through an amide bond to the chromogen para-nitroaniline (PNA). The release of free PNA can be monitored spectrophotometrically by measuring the increase of absorbance of light in the range of 380 to 410 nm. The present communication describes our efforts at enhancing the color yield of the assay as well as changing the absorption maximum to a wavelength which can more readily and accurately be measured on inexpensive laboratory spectrophotometers .Bratton and Marshall have developed a method which can be used for the determination of aromatic amines in biological systems. We have modified this method for use in semimicro applications and have adapted it to the measurement of free PNA in solution. After acidification of an aliquot of the enzyme digest with trichloroacetic acid (TCA), the liberated PNA is diazotized with sodium nitrite. Excess nitrous acid is removed with ammonium sulfamate, and the diazotized PNA is reacted with 1-naphthylethylenediamine reagent. The product has a magenta-red color with an absorption maximum of 545 nm. The substrate is not affected by exposure to TCA, and when fresh reagents are used, a low background is obtained. The color yield during actual enzyme assay is amplified twenty fold. Other coupling agents are also currently under investigation.

Development of a rapid and sensitive method for determination of cysteine/cystine ratio in chemically defined media

2010 ◽  
Vol 1217 (24) ◽  
pp. 3863-3870 ◽  
Author(s):  
Hassan Alwael ◽  
Damian Connolly ◽  
Leon Barron ◽  
Brett Paull
Keyword(s):  
Chemically Defined Media ◽  
Defined Media ◽  
Sensitive Method

scholarly journals The thermal and electrical conductivity of a single crystal of aluminium

1927 ◽  
Vol 115 (770) ◽  
pp. 236-241 ◽  
Keyword(s):  
Electrical Conductivity ◽  
Single Crystals ◽  
Test Specimen ◽  
Average Diameter ◽  
National Physical Laboratory ◽  
Present Communication ◽  
Uniform Size ◽  
The Third ◽  
Physical Laboratory

The recent work of Carpenter and Elam on the growth of single crystals of large dimensions has rendered possible the study of the physical constants of single crystals of the commoner metals, and the present communication describes the determination of the thermal and electrical conductivity of aluminium in the form of an isolated crystal. The form of the crystal investigated is shown in fig. 1. This crystal had been prepared at the National Physical Laboratory employing the technique described by Carpenter in “Nature,” p. 266, August 21, 1926, which briefly is as follows:— The test specimen is machined and subjected to three treatments, thermal, mechanical, and thermal. The first treatment is necessary to soften the metal completely and produce new equiaxed crystals of so far as possible uniform size, the average diameter being 1/150 inch. The second consists in straining these crystals to the required amount, and the third in heating the strained crystals to the requisite temperature, so that the potentiality of growth conferred by strain could be brought fully into operation.

scholarly journals A MICROPHOTOMETRIC STUDY OF THE SYNTHESES OF DESOXYRIBONUCLEIC ACID AND NUCLEAR HISTONE

1955 ◽  
Vol 1 (1) ◽  
pp. 17-28 ◽  
Author(s):  
David P. Bloch ◽  
Gabriel C. Godman
Keyword(s):  
Cell Division ◽  
Nucleic Acids ◽  
Standard Error ◽  
Basic Protein ◽  
Desoxyribonucleic Acid ◽  
Fast Green ◽  
Nuclear Histone ◽  
Source Of Error ◽  
Formalin Fixed

1. The fast green stain of Alfert and Geschwind for nuclear basic protein is shown to obey the Beer-Lambert laws when used on purified histone. Interference from acid substances other than nucleic acids as a possible source of error is indicated. 2. Use of this technique after a modified Feulgen stain enables determination of relative amounts of desoxyribonucleic acid and histone in the same individual cells. 3. DNA and histone are shown to have the same distribution in formalin-fixed nuclei. 4. The syntheses of DNA and histone proceed simultaneously resulting in the doubling of both these substances prior to cell division. 5. The standard error for histone values is greater than that for DNA; however, the source of this variability is not known.

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