Application of simultaneous determination of 3H, 14C, and 22Na by liquid scintillation counting to the measurement of cellular ion-transport.

A liquid scintillation counting method for simultaneous determination of three radioactive nuclides (3H, 14C, and 22Na) of biological interest was studied. By comparing the beta spectra of the three nuclides, their counting energy ranges, A, B, and C, were determined. 22NA was set high enough to avoid any spillover counts from lower-energy nuclides. Region A for 3H was set to maximize the counting efficiency. A good correlation between the counting efficiency for 22Na in region C and the counting efficiency of other nuclides in all regions was obtained. Prior to 3H and 14C dpm calculations, the 22Na counts spilled down in regions A and B were subtracted from the total counts in regions A and B. A simple linear equation was then used to compute 3H and 14C dpm. Findings show that the method presented is adaptable for highly quenched samples up to quenching indices of tSIE = 100. The method is useful for studying the biological transport coupled to Na+.     

A sonar fish counter

The basic principles of a new technique for counting fish moving up or down stream are given. The counter is based on the ability to detect the presence of fish in an acoustic beam directed across the river. The counter is capable of counting several fish moving up or down stream simultaneously and is designed for use in wide open channels. The results of field trials with the equipment at two different sites are discussed together with the problems encountered. The results obtained are in good agreement with other methods of counting fish but the totals obtained were insufficient to put confidence limits on the counter.     

Use of the IUL Countermat Automatic Colony Counter for Spiral Plated Total Viable Counts

An IUL Countermat automatic colony counter was used to enumerate colonies on spiral total viable count plates made with a wide variety of foods. The counter results exhibited a correlation with manual counting results similar to the reproducibility obtained with manually counted spiral plates. Use of this machine results in a large time saving compared with the conventional counting method and is recommended as a generally suitable method for counting spiral total viable count plates.     

Sizing of Bdellovibrio During Growth

A technique for the counting and relative sizing of host-independent bdellovibrio during growth, with the aid of a modified Coulter counter, is described.     

Automated counting of bacterial colony forming units on agar plates

Manual counting of bacterial colony forming units (CFUs) on agar plates is laborious and error-prone. We therefore implemented a colony counting system with a novel segmentation algorithm to discriminate bacterial colonies from blood and other agar plates.A colony counter hardware was designed and a novel segmentation algorithm was written in MATLAB. In brief, pre-processing with Top-Hat-filtering to obtain a uniform background was followed by the segmentation step, during which the colony images were extracted from the blood agar and individual colonies were separated. A Bayes classifier was then applied to count the final number of bacterial colonies as some of the colonies could still be concatenated to form larger groups. To assess accuracy and performance of the colony counter, we tested automated colony counting of different agar plates with known CFU numbers of S. pneumoniae, P. aeruginosa and M. catarrhalis and showed excellent performance.     

Development of a Quantitative Polyacrylamide Gel Electrophoresis Analysis Using a Multichannel Radioactivity Counter for the Evaluation of Oligonucleotide-Bound Drug Carrier

A quantitative polyacrylamide gel electrophoresis (PAGE) analysis using a multichannel radioactivity counter was designed for the evaluation of³³P-labeled antisense oligonucleotide associated with polymeric drug carrier (nanoparticles). The proposed analytical method was first validated. The criteria of specificity, linearity, reliability, detection and quantification limits, and resolution power were determined. Results were compared to those obtained using liquid scintillation counting of crude samples or after solubilization of gel slices. The proposed method gave a better linearity and reliability than liquid scintillation counting of solubilized gel slices. In comparison with the liquid scintillation counting of crude samples, the method presented the advantage of being able to directly separate oligonucleotides differing by only one nucleotide in length. This method was applied for the separation of free oligonucleotides and oligonucleotides bound onto nanoparticles, allowing quantification of the amount of free and bound oligonucleotides without any further separation steps. Thus, because it is easy and rapid, the quantitative PAGE analysis using a multichannel radioactivity counter offers interesting possibilities for the characterization of oligonucleotide nanoparticles.     

Quantitative autoradiography of dot blots using a microwell densitometer

We have established conditions for the quantitation of DNA hybridization by "reading" dot blot autoradiographs with a microwell plate densitometer. This method is more convenient, as accurate, and more sensitive than counting the spots in a liquid scintillation counter.     

Evaluation of leukocyte number by using an automated blood cell counter and a traditional hematological method in animals irradiated with gamma rays

Female mice were irradiated with a single whole body dose of 7 Gy of gamma-rays. Leucocyte numbers were monitored in the peripheral blood using automated blood cell counter Coulter counter and a traditional hematological method with a light microscope in the Bürker chamber. Reticulocyte numbers, RNA blood concentration, spleen weight and morphological changes in spleen and bone marrow were also studied. In the period between 15th-19th days after irradiation the numbers of leucocytes obtained by CC counting were manifold higher than those obtained by microscope counting. Since this period is characterised by a steep increase in the reticulocyte number and RNA concentration in blood as well as by increased weight of spleen as the result of marked regeneration of extramedullar erythropoiesis, leukocytes as well as reticulocytes are assumed to be additionally registered by the automated counter CC in this period, probably due to a higher resistance of reticulocytes to the lysing agent Zapoglobine.     

A simple electronic apparatus for the analysis of radioactively labeled gel electrophoretograms

Radioactively labeled gel electrophoretograms are generally analyzed either by slicing the gel and dissolving the individual pieces in a fluor for scintillation counting or by exposure to photographic emulsions. In this report, we describe a technique for fast quantitative analysis of such electrophoretograms, based on counting individually β-or γ-particles emerging from the gel surface and locating their position to ±0.6 mm. The apparatus which employs a position-sensitive single-wire gas proportional counter is simple to construct and operate with readily available electronics.     

Improved analysis of hemagglutination assays for quantitation of lectin activity

A method to quantitate lectin activity based on hemagglutination assay in microtiter plates is described. In addition to the normal method of visual titer evaluation an electronic particle counter is used for counting of nonagglutinated cells in the microtiter wells; this allows a rapid, quantitative determination of the amount of lectin required to agglutinate 50% of the countable single cells. It is also recommended that counting results should be related to a standard curve of concanavalin A to improve the reproducibility of the assay.     

Application of an automated colony counter for evaluation of the viability of a yeast culture

Application of an automated colony counter for evaluation of the viability of microbial cultures was investigated with yeast cultures as a model. Statistical comparison of the results of automated and visual (“manual”) colony counting is presented, as well as the results of the application of the bundled software to digital images obtained by light microscopy for determination of the cell concentration in suspensions. Automated counting is concluded to significantly accelerate the evaluation of culture viability by colony-forming capacity, provided that a certain requirements of sample preparation and analysis are observed.     

On the Correlation of the Volume of a Prolate-Spheroidal Cell, Tetrahymena as Determined by Electronic Particle Counters and by Morphometry

ABSTRACT. The cell volume provided by electronic particle counters may be incorrect. As a particle, or cell, passes the counting device, its volume is calculated as a sphere. The electronically derived, mean cell volume (electronic MCV) of a population of Tetrahymena (prolate spheroid) is smaller than the volume (morphometric MCV) calculated from measured cell length and width. This discrepancy was studied using a Coulter Multisizer particle counter and cell morphometry. The electronic MCV averaged 0.70 of the morphometric MCV (1.00) but changed from 0.72 (fast growth) to 0.63 and 0.76 (slow or no growth) for cells having a mean length/width of 2.05, 2.33, and 1.61, respectively. The measured diameter of latex particles (used for calibration) was identical to that stated, but the diameter of the electronic MCV was larger than the width of the cells which related to wehther the length/width of the cells was above, or below, 2.00. Hence, electron particle counters register primarily the width of a prolate-spheroidal cell, oriented with its long axis in the direction of flow, and uses this value as diameter for the calculated sphere, whereas for more spherical cells, tumbling without any orientation, a mean of the axes is used. Factors for correction of the electronic MCV of Tetrahymena are provided.     

Statistical evaluation of electronic and plate counts for estimating bacterial populations

Bacteria from cultures of Staphylococcus aureus, Pseudomonas geniculata, and Rhodotorula glutenis were counted by use of an electronic counter and by plate counts from broth cultures at selected periods from 0 to 24 hr. Variations in the two methods were noted, and the results were compared after calculating correlations, coefficients of variation, and nested analyses of variance. It was not possible to determine the absolute accuracy of the two methods; however, the precision of the results obtained with an electronic counter was better than that obtained with plate counting. It appears that electronic counting, because of ease and convenience, provides better comparative counts at a particular stage of growth, especially during the early stages of the growth period, whereas plate counting yields the best results in determining growth curves.     

Counting of p32 in plant tissues using the Cerenkov effect

Summary A procedure for counting p³² in plant tissues is presented. The method, based on the use of Cerenkov radiation, involves practically no sample preparation. Plant tissue are placed into vials containing water or hexane and counted with a liquid scintillation counter. Counts obtained, using this procedure were found to be linearily related to that obtained with a G.M. tube. The counting efficiency was, however, higher with the proposed method. The use of hexane is advantageous if leakage of p³² from the tissue is possible, or when higher counting efficiency is desireable. The use of different liquids may also enable a discriminative count of different beta emitters. As suggested recently⁸ use of wavelength shifter may further increase efficiency of counting Cerenkov radiation.     

Evaluation of an Automated Colony Counter

An automated colony counter was found to readily detect surface and subsurface bacterial colonies of 0.3-mm size or greater with a high degree of precision. On a logarithmic scale, counting efficiency consistently ranged from 89 to 95% of corresponding manual count determinations for plates containing up to 1,000 colonies. In routine application, however, automated plate counts up to approximately 400 colonies were selected as a more practical range for operation. The automated counter was easily interfaced with an automated data acquisition system.     

The effect of sample composition and vial type on Ĉerenkov counting in a liquid scintillation counter

The effect of sample vial type and sample composition on the ?erenkov count rate detected from ³²P and ³⁶Cl was studied using a liquid scintillation counter. When counting was done in the noncoincident mode, glass vials allowed higher counting efficiency than plastic vials. In the coincident mode light scattering caused by polyethylene and polyproplyene vials allowed higher counting efficiency than glass vials. Highest coincident counting efficiency was from plastic minivials in a glass carrier vial. Increased solute concentration in samples caused increased counting efficiency due to changes in the refractive index of the solution. This can cause significant counting efficiency changes with no sample channel ratio change in density gradient fractions. The use of wavelength shifters is shown to be inappropriate when the sample pH varies, as this can change the fluorescent properties of the shifters and thereby the observed count rate.     

Liquid scintillation counting of radioactive monomeric and polymeric carbohydrates on paper chromatograms

A simple, improved scintillation counting procedure was developed for the assay of radioactive mono- and polysaccharides on paper chromatograms. Segments of chromatograms are placed in scintillation vials and soaked in water to completely elute the carbohydrate before addition of Aquasol, a xylene-based scintillation fluid. The resulting water-Aquasol solution is counted in a liquid scintillation counter. Evaluation of numerous experimental variables revealed optimal conditions for complete elution of mono- and polysaccharides with water before counting in Aquasol.The water elution-Aquasol procedure allows water-soluble substances (¹⁴C- and ³H-labeled) on paper to be assayed with increased accuracy and sensitivity (three- to fivefold improvement in counting efficiency of tritiated samples). The simplicity of the procedure allows entire radiochromatograms to be assayed readily.     

Detection of reagent errors by internal quality control of the Coulter counter S]

By a simple program of quality control for the Coulter counter model S two deficient reagents could be detected. To high values of PCV associated with a decrease of MCHC were caused by a faulty Isoton batch. This error was revealed by native blood samples but not by the stabilized 4 C standard. Falsely elevated leukocyte counts due to insufficiently lysed giant platelets and platelet aggregates were produced by modified Lyse-S batches. Comparisons made with a Coulter counter model F using Zaponin and with a counting chamber yielded values at lower levels.     

Improved synchronized accumulating radioisotope detector for gas chromatography

A synchronized accumulating radioisotope detector for radio gas chromatography was developed. It comprised seven gas-flow proportional counters each with an inner volume of 10 ml. Every counter tube was connected by a mutual anti-coincidence circuit to reduce the background. The transit time of gas particles in one counter tube could be set to an optimal value between 1 and 4 s by regulating the flow-rate of the counting gas, according to analytical requirements. The improved detector maintained high chromatographic resolution, which suggested the applicability of the apparatus to capillary gas chromatography.