Laboratory Workstation Research Articles

A semi-automated, microplate version of the SOS Chromotest for the analysis of complex environmental extracts

Environmental monitoring for genotoxicity requires that a large number of measurements be made across space and time. This requirement demands a rapid and efficient bioassay system. The SOS Chromotest is a rapid, efficient bacterial system for the detection of DNA damaging agents. Over 100 publications have described its use on a variety of samples. Relatively few studies have used the test to examine complex mixtures. Effective testing of complex samples poses a variety of problems. Although solutions have been proposed, few have validated the resulting protocol. In this work we present a semi-automated microplate version of the SOS Chromotest for the examination of complex mixtures. Experiments were conducted to determine the optimal cell concentration, exposure time, substrate conversion time and S9 enzyme concentration. The performance of the method was evaluated using 6 reference genotoxins and 3 complex mixtures. The complex mixtures examined are extracts of diesel particulate matter, urban dust and coal tar. The results obtained indicate that optimal responses often require fewer cells (∼ 5–10 × 10 6 CFU/ml) and a longer exposure (3 h) than that recommended in the original protocol. Interfering effects of colored and turbid samples are removed using centrifugation and initial optical density readings taken 60 min after cell resuspension and lysis. The performance of the established protocol was evaluated using mitomycin C and benzo[ a]pyrene results for 207 microplates and solvent control results for 293 microplates. The results indicate that the established method is accurate, sensitive and precise. Coefficient of variation on mean SOSIP values for mitomycin C and benzo[ a]pyrene are <5%. Solvent control data indicate that the standard threshold for determination of a positive response (induction factor > 1.5) is excessively conservative. All liquid transfers were automated using the Biomek TM automated laboratory workstation. Automation permits a throughput of up to 72 samples per day and maintains excellent precision and accuracy.

Laboratory Workstation and Experiments on Discretization, Identification and Computer Control

The paper deals with the implementation of a basic workstation and the development of experiments for use in a Control Laboratory at educational institutions. The station consists of a digital computer, an A/D-D/A interface, some simulated or real processes and computer programs. The programs were written to interact with a well-known scientific computing environment, being the lab experiments referred to: Discretization of Continuous Systems, to obtain discrete dynamical equivalents tor continuous filters and compensators; Process Identification, to identify nonparametric and parametric models for dynamical processes; and Digital Control, to design, simulate and implement dynamical compensators, state-space and optimum controllers.

Supercomputer algorithms for efficient linear octree encoding of three-dimensional brain images

We designed and implemented algorithms for three-dimensional (3-D) reconstruction of brain images from serial sections using two important supercomputer architectures, vector and parallel. These architectures were represented by the Cray YMP and Connection Machine CM-2, respectively. The programs operated on linear octree representations of the brain data sets, and achieved 500–800 times acceleration when compared with a conventional laboratory workstation. As the need for higher resolution data sets increases, supercomputer algorithms may offer a means of performing 3-D reconstruction well above current experimental limits.

Chromatography in DNA radiolabeling: hands-off automation using a robotic workstation.

The experiments described in the present paper were performed in order to determine whether the Biomek-1000 (Beckman Instruments, Fullerton, CA, USA) automated laboratory workstation can be used in a fully automated DNA labeling method followed by automated gravity-driven size exclusion purification of molecular probes. To this aim, we performed random oligodeoxyribonucleotide priming of a HIV-1 LTR probe that was used for molecular hybridization to Southern blotted polymerase chain reaction products. The results obtained demonstrate that the automatically labeled probe can be efficiently purified by automated and gravity-driven Sephadex G-50 chromatography, without any major changes in hybridization property. This robotic methodology can be used in several procedures employing radioisotope labeling.

A chromatographic procedure for fully automated isolation of DNA from human whole blood

Isolation of DNA from peripheral blood constitutes a fundamental step in the molecular diagnosis of genetic disorders, analysis of mutations of oncogenes and detection of nucleic-acid sequences of pathogenic organisms in clinical samples. We investigated whether dextran sedimentation of blood followed by the chromatographic isolation of DNA could be applied to the Biomek-1000 automated laboratory workstation in order to develop a fully automated method of DNA isolation from whole blood. Under our experimental conditions, the Biomek-1000 could perform fully automated DNA isolation from seven human whole blood samples in less than 1.5 h. Our data that the DNA isolated from human blood in this way is suitable for polymerase chain reaction.

The laboratory workstation: a data management model for a small laboratory section.

Many small sections of clinical chemistry laboratories fail to benefit fully from laboratory information system (LIS) installation because the volume does not justify cost of interfacing the instruments in these sections to the LIS. The opportunity for manual data entry errors remains a problem in these sections. This paper describes the design of a laboratory workstation that serves as a data hub that makes it feasible to use one instrument interface line from the LIS to acquire data from four instruments. This approach reduces the likelihood of data entry error and improves the efficiency of personnel in the laboratory section. Further, the tools necessary to create such a workstation are commercially available and do not depend on microcomputer programming and support personnel within the chemistry laboratory.

Atomic level simulations on a million particles: The cell multipole method for Coulomb and London nonbond interactions

The N2 computations implicit in the Coulomb and other long range interactions remain the critical bottleneck in atomic-level simulations of the structure and dynamics of large systems. We report here the cell multipole method which scales linearly with N and requires only modest memory. To demonstrate the feasibility of this approach, we report systematic calculations on realistic polymer systems with up to 1.2 million atoms on a laboratory workstation. The method becomes faster than the exact method for systems of 300 atoms, and for a 1.2 million-atom polymer, it is 2377 times faster. The method treats a class of interactions of the form qiqj/rijp, which includes Coulomb (p=1), London dispersion (p=6), or shielded Coulomb (p=2) interactions. This method is well suited for highly parallel and vector computers.

The reduced cell multipole method for Coulomb interactions in periodic systems with million-atom unit cells

Standard methods for calculating Coulomb interactions of periodic systems use Ewald-type formulations or minimum image approximations, neither of which is practical for large (million-atom) systems. We describe the reduced cell multipole method which is 38 times faster than the Ewald method for a 4816-atom (unit cell) system. The method scales linearly with the number of atoms in the cell, making simulations of million-atom systems practical, as demonstrated in a series of applications on a laboratory workstation.

Evaluation of video gray‐scale display

Setting up and maintaining video display monitors properly will help to reduce display variation and improve overall presentation of the radiological image. Display monitor gray-scale characteristics were examined using the SMPTE test pattern. This test pattern may be used as a standard for adjusting brightness and contrast. The controls should be adjusted to display the full dynamic range so that the 5% and 95% signal levels in the pattern are visible. Measured luminance on a laboratory workstation used for radiological perceptual experiments, and on the Siemens CT gray-scale monitor was determined to range from 0.17 to 76.0 nit, and 0.17 to 24.66 nit, respectively. These were compared with the range of approximately 17 to 514 nit for a typical film-viewbox combination. Characteristic curves were determined for both monitors, and CRT gammas were 3.34 and 2.48 for the perceptual workstation and CT console, respectively. The display gamma was determined from fitting luminance data to a log-log plot of luminance versus input gray level. The usefulness of the SMPTE test pattern for visual presentation as well as photometric measurement is demonstrated.

Studies on the quantification of proteoglycans by the dimethylmethylene blue dye-binding method. Specificity, quantitation in synovial lavage fluid, and automation.

The dimethylmethylene blue (DMMB) dye-binding technique is widely used for the quantification of sulfated glycosaminoglycans (sGAG) and proteoglycans. We conducted further studies on this technique in our laboratory and found that concentrations of DNA and RNA in excess of 20 micrograms/ml interfered negatively with the detection of sGAGs; interference was eliminated by using DNase and RNase. Hyaluronan at 40 micrograms per ml did not interfere with the detection of sGAG. However, because of the higher concentrations of hyaluronan in synovial lavage fluid, it was necessary to treat this fluid with Streptomyces hyaluronidase in order to quantify sGAG. The DMMB assay was automated with a laboratory work station and compared to the standard method.

A microplate version of the SOS/ umu-test for rapid detection of genotoxins and genotoxic potentials of environmental samples

The umu-microtest is a miniaturized automated short-term test version proposed for screening of umuC-dependent mutagenic potentials of chemicals relevant to environmental pollution, river water and industrial waste water. The test is based on the SOS/ umu-test and has been modified in order to allow extensive testing of environmental samples. Genetically engineered Salmonella typhimurium (TA1535/pSK1002) are incubated on a microplate rotor in a sloping position for 2 h with the test samples, followed by addition of fresh culture medium to reach a 10-fold dilution of the incubation medium. 2 h later, the activity of the β-galactosidase, which reflects umuC induction, is determined colorimetrically. The incubation of the bacteria in the presence of the test compounds as well as the assessment of β-galactosidase activity takes place in 96-well microplates, thus enabling simultaneous screening of large numbers of samples. Data of the genotoxic potentials are available within 8 h. Computer-controlled automation is possible by using a laboratory workstation.

High-throughput fluorescent DNA sequence analysis: Methods and automation

A strategy for high-throughput DNA sequence analysis using the fluorescent dye-primer chemistry is described. This strategy combines rapid preparation of template DNA using a modification of the polymerase chain reaction (PCR), automation of the DNA sequencing reactions using a robotic laboratory workstation, and subsequent analysis of the fluorescent-labeled reaction products on a commercial automated fluorescent sequencer. Asymmetric PCR provides a reliable means of simultaneously producing sufficient and uniform quantities of several template DNAs directly from bacterial colonies or bacteriophage plaques. The automation scheme for subsequent processing and analysis of DNA samples allows a DNA sequencing facility to operate the fluorescent sequencer twice daily at its full capacity of 24 samples. Furthermore, the methods and instruments described eliminate much of the labor required by technicians to obtain a high level of data output. As described here, a DNA sequencing laboratory equipped with a single automated fluorescent sequencer can perform analysis of up to 48 fluorescent-labeled reactions every day, with an average output of over 10,000 bp per sequence run.

Computer Laboratory Workstation Dimensions:

No abstract available for this article.

The biomek 1000 ? A laboratory workstation which, due to its modular structure, becomes an adapted system i.e. for ELISA assays in feed and food analysis

The biomek 1000 ? A laboratory workstation which, due to its modular structure, becomes an adapted system i.e. for ELISA assays in feed and food analysis

Automation of a chloramphenicol acetyltransferase assay

Accurate quantification of chloramphenicol acetyltransferase (CAT) enzyme activity in a large number of samples has been achieved through robotization of a CAT assay on a laboratory workstation (Biomek 1000). The basic principle of this CAT assay relies on the selective diffusion of [ 3H]acetylchloramphenicol into a water-immiscible liquid scintillation cocktail. This methodology gives unique characteristics to this robotized protocol by allowing complete control over the kinetics of the CAT enzymatic reaction which is a critical parameter in the CAT assay. Thus it has been possible to optimize the CAT assay for every processed sample, through real time monitoring of the enzymatic reaction, and to achieve maximum accuracy in CAT quantification. Moreover the sensitivity of this automated assay is high (detection threshold; 10 −4 CAT unit), and the sample processing is fast (≈ 125 samples per hour). Compared to other CAT assay protocols currently used, our robotized technique offers major advantages in terms of CAT quantification, and sets new standards for CAT assay productivity.

A microplate version of the DNA-synthesis inhibition test for rapid detection of DNA-alteration potentials

A microplate version of the DNA-synthesis inhibition test (DIT) for fast detection of DNA-alteration potentials has been developed. The DIT is based on the concept that DNA damage causes inhibition of DNA synthesis that becomes detectable some time after replicating cells have been in contact with genotoxic agents. In this test procedure human tissue culture cells (HeLa S3), prelabeled with [ 14C]thymidine, are exposed for 90 min to the substances in question. After the cells are rinsed, they are allowed to recover for 2 1 2 h in fresh culture medium, thereby unspecific interactions interfering with DNA replication are practically eliminated. Next, [ 3H]thymidine is added for 30 min, and then the cells are harvested and thoroughly rinsed. Finally, incorporated radioactivity is determined by liquid scintillation counting for measurement of the 3H 14C ratio. This allows for the evaluation of DNA synthesis during the 3H-labeling period and of the extent of genotoxic damage. This microplate version of the DIT can be carried out fully automated in a laboratory workstation. The test is compared to other tests for genotoxicity. Its advantages are discussed.

Development of an automated procedure for fluorescent DNA sequencing

We describe here the development of a procedure for complete automation of the dideoxynucleotide DNA sequencing chemistry using fluorescent dye-labeled oligonucleotide primers. This procedure combines rapid preparation of template DNA using a modification of the polymerase chain reaction, automation of the DNA sequencing reactions using a robotic laboratory workstation, and subsequent analysis of the fluorescent-labeled reaction products on a commercial automated fluorescent sequencer. Using this procedure, we were able to produce sufficient quantities of template DNA directly from bacterial colonies or bacteriophage plaques, perform the DNA sequencing reactions on these templates, and load the reaction products on the fluorescent DNA sequencer in a single work day. This scheme for automation of the fluorescent DNA sequencing method allows the fluorescent sequencer to be run at its full capacity every day and eliminates much of the labor required to obtain a high level of data output. Currently, we are able to perform and analyze 16 fluorescent-labeled reactions every day, with an average output of over 7000 bp per sequencer run.

Utilization of a computer-controlled laboratory workstation (Biomek 1000) in routine radioimmunoassay laboratory

The suitability and performance characteristics of a recently introduced computer-controlled laboratory workstation (Biomek 1000) for use in automating sample transfer and reagent additions in radioimmunoassay techniques were assessed. The system is based on the use of disposable tips and, therefore, reduces any possible sample carry-over and eliminates the need for priming with the subsequent reduction in cost of reagents. However, the machine lacks the useful option of liquid-level sensing facility. Acceptable technical performance in terms of precision, accuracy and throughput was obtained with the Biomek 1000 which complied with international recommendations on the safety of hospital laboratory equipment.

Automated determination of excitatory amino acid neurotoxicity in cortical culture

We used a commercially available robotic laboratory workstation to quantitatively study excitotoxic neuronal injury in cell culture. A Beckman Instruments Biomek 1000 was programmed to perform both timed exposures to excitatory amino acid agonists, and kinetic assay of the resultant efflux of lactic dehydrogenase from damaged neurons, using 96-well culture plates. Examination of homocysteate neurotoxicity utilizing this automated method produced results similar to those obtained earlier using manual techniques. The method described here may facilitate the characterization of neurotoxic agonist or antagonist activity.

ワークステーションＢｉｏｍｅｋ１０００の輸血検査への適用

In order to promote rationalization and economization of blood transfusion test, the utility of automated laboratory workstation Biomek 1000 for blood grouping, irregular antibody screening, hepatitis B antigen and antibody screening, etc. of patients' specimens was compared with manual operation. Processing was much faster with Biomek 1000 than that with manual operation in each test. The volume of reagent used with Biomek 1000 was smaller that that with manual operation at blood grouping and antibody screening. The results with both procedures were correlated well in each experiment. Then, Biomek 1000 is very useful with respect to rationalization and economization of blood transfusion test and protection of laboratory staff against infectious diseases.