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PubMedDate not supplied

Flexible automation of cell culture and tissue engineering tasks.

Until now, the predominant use cases of industrial robots have been routine handling tasks in the automotive industry. In biotechnology and tissue engineering, in contrast, only very few tasks have been automated with robots. New developments in robot platform and robot sensor technology, however, make it possible to automate plants that largely depend on human interaction with the production process, e.g., for material and cell culture fluid handling, transportation, operation of equipment, and maintenance. In this paper we present a robot system that lends itself to automating routine tasks in biotechnology but also has the potential to automate other production facilities that are similar in process structure. After motivating the design goals, we describe the system and its operation, illustrate sample runs, and give an assessment of the advantages. We conclude this paper by giving an outlook on possible further developments.

Artificial Intelligence↗
PubMed2003

Smartnotebook: a semi-automated approach to protein sequential NMR resonance assignments.

Complete and accurate NMR spectral assignment is a prerequisite for high-throughput automated structure determination of biological macromolecules. However, completely automated assignment procedures generally encounter difficulties for all but the most ideal data sets. Sources of these problems include difficulty in resolving correlations in crowded spectral regions, as well as complications arising from dynamics, such as weak or missing peaks, or atoms exhibiting more than one peak due to exchange phenomena. Smartnotebook is a semi-automated assignment software package designed to combine the best features of the automated and manual approaches. The software finds and displays potential connections between residues, while the spectroscopist makes decisions on which connection is correct, allowing rapid and robust assignment. In addition, smartnotebook helps the user fit chains of connected residues to the primary sequence of the protein by comparing the experimentally determined chemical shifts with expected shifts derived from a chemical shift database, while providing bookkeeping throughout the assignment procedure.

Amino Acid Sequence↗
PubMed2003

Influence of the completeness of chemical shift assignments on NMR structures obtained with automated NOE assignment.

Reliable automated NOE assignment and structure calculation on the basis of a largely complete, assigned input chemical shift list and a list of unassigned NOESY cross peaks has recently become feasible for routine NMR protein structure calculation and has been shown to yield results that are equivalent to those of the conventional, manual approach. However, these algorithms rely on the availability of a virtually complete list of the chemical shifts. This paper investigates the influence of incomplete chemical shift assignments on the reliability of NMR structures obtained with automated NOESY cross peak assignment. The program CYANA was used for combined automated NOESY assignment with the CANDID algorithm and structure calculations with torsion angle dynamics at various degrees of completeness of the chemical shift assignment which was simulated by random omission of entries in the experimental 1H chemical shift lists that had been used for the earlier, conventional structure determinations of two proteins. Sets of structure calculations were performed choosing the omitted chemical shifts randomly among all assigned hydrogen atoms, or among aromatic hydrogen atoms. For comparison, automated NOESY assignment and structure calculations were performed with the complete experimental chemical shift but under random omission of NOESY cross peaks. When heteronuclear-resolved three-dimensional NOESY spectra are available the current CANDID algorithm yields in the absence of up to about 10% of the experimental 1H chemical shifts reliable NOE assignments and three-dimensional structures that deviate by less than 2 A from the reference structure obtained using all experimental chemical shift assignments. In contrast, the algorithm can accommodate the omission of up to 50% of the cross peaks in heteronuclear- resolved NOESY spectra without producing structures with a RMSD of more than 2 A to the reference structure. When only homonuclear NOESY spectra are available, the algorithm is slightly more susceptible to missing data and can tolerate the absence of up to about 7% of the experimental 1H chemical shifts or of up to 30% of the NOESY peaks.

Algorithms↗
PubMed1992

Do computerised visual fields and automated optic disc analysis assist in the choice of therapy in glaucoma?

The choice of therapy in patients with glaucoma is determined by the presence of damage in the visual field or optic disc or by change in the appearance of the optic disc or visual field over time. Computerised visual fields and automated optic disc analysis can assist in this determination. Automated optic disc analysis permits the measurement of neuroretinal rim area which is more closely related to the presence of pathology than cup/disc ratio. Simple ophthalmoscopic techniques may also be used to estimate the size of the neuroretinal rim area. Automated perimetry allows accurate diagnostic information to be obtained in a reliable manner. This permits easier diagnosis of glaucoma as well as easier long term follow up for change. The availability of automated techniques in ophthalmology has helped the clinician in the choice of therapy. The use of astute clinical judgement remains however of the utmost importance.

Diagnosis, Computer-Assisted↗
PubMed2002

Automated subcellular localization and quantification of protein expression in tissue microarrays.

The recent development of tissue microarrays-composed of hundreds of tissue sections from different tumors arrayed on a single glass slide-facilitates rapid evaluation of large-scale outcome studies. Realization of this potential depends on the ability to rapidly and precisely quantify the protein expression within each tissue spot. We have developed a set of algorithms that allow the rapid, automated, continuous and quantitative analysis of tissue microarrays, including the separation of tumor from stromal elements and the sub-cellular localization of signals. Validation studies using estrogen receptor in breast carcinoma show that automated analysis matches or exceeds the results of conventional pathologist-based scoring. Automated analysis and sub-cellular localization of beta-catenin in colon cancer identifies two novel, prognostically significant tumor subsets, not detected by traditional pathologist-based scoring. Development of automated analysis technology empowers tissue microarrays for use in discovery-type experiments (more typical of cDNA microarrays), with the added advantage of inclusion of long-term demographic and patient outcome information.

Algorithms↗
PubMed2006

Effect of posterior capsular opacification removal on automated perimetry.

PURPOSE: Although posterior capsular opacification (PCO) is a common phenomenon in a considerable number of ophthalmologic patients, no prospective controlled trials assessing its influence on automated perimetry exist. This technique continues as a standard in the diagnosis of glaucoma and neuro-ophthalmological diseases. The aim of the present report is to investigate the effect of PCO on automated visual field examination. METHODS: A total of 26 PCO affected eyes of 26 patients had Humphrey SITA standard (program 24-2) immediately before, and between 1 and 8 weeks after Neodymium : YAG capsulotomy. The effect of learning associated with repeated testing was controlled with automated perimetry before enrollment and visual fields of the fellow eye. Best-corrected visual acuity (BCVA), intraocular pressure (IOP), and global perimetric pre- and post-laser indices were compared using the Student's t-test for paired samples. Correlation and linear regression analyses were also performed. RESULTS: BCVA and mean deviation (MD) improved following capsulotomy. Pattern standard deviation (PSD), an indicator of localized defects in the field, also improved significantly when PCO was solved. Moreover, a strong association among BCVA, MD, and PSD was shown both prior to and after capsulotomy. CONCLUSION: PCO is a heterogeneous mean opacity. This polymorphism may alter visual field results, and may even simulate the perimetric behaviour of other pathologies such as glaucoma. Consequently, the presence of PCO should be considered in the interpretation of any automated perimetry in pseudophakic patients. In addition, the values obtained before capsulotomy may partially predict the values obtained after capsulotomy.

Aged↗
PubMed1997

The impact of physician vs automated blood pressure readings on office-induced hypertension.

Blood pressure (BP) readings in the doctor's office are frequently higher than home or ambulatory values. This study examines the role of the physician in the aetiology of the 'white coat' effect, by comparing standard readings taken by the family physician of 27 treated hypertensive patients with readings taken by an automated BP recording device, with the patient alone in the examining room during the same office visit. The physician and automated readings were each compared to the mean awake ambulatory BP. Mean (+/-s.e.m.) routine office BP (mm Hg) recorded by the patient's physician (155+/-4/80+/-2) was similar to the mean value obtained using the automated BP recording device (157+/-3/83+/-2). The mean awake ambulatory BP was 145+/-3/78+/-2 with the systolic value lower (P < 0.05) than either the physician or automated reading. Self-measurement of BP by the patient in the office setting does not reduce the magnitude of the white coat effect.

Aged↗
PubMed1997

Automated fed-batch fermentation with feed-back controls based on dissolved oxygen (DO) and pH for production of DNA vaccines.

A fermentation process in Escherichia coli for production of supercoiled plasmid DNA for use as a DNA vaccine was developed using an automated feed-back control nutrient feeding strategy based on dissolved oxygen (DO) and pH. The process was further automated through a computer-aided data processing system to regulate the cell growth rate by controlling interactively both the nutrient feed rate and agitation speed based on DO. The process increased the total yield of the plasmid DNA by approximately 10-fold as compared to a manual fed-batch culture. The final cell yield from the automated process reached 60 g L-1 of dry cell weight (OD600 = 120) within 24 h. A plasmid DNA yield of 100 mg L-1 (1.7 mg g-1 cell weight) was achieved by using an alkaline cell lysis method. Plasmid yield was confirmed using High Performance Liquid Chromatography (HPLC) analysis. Because cells had been grown under carbon-limiting conditions in the automated process, acetic acid production was minimal (below 0.01 g L-1) throughout the fed-batch stage. In contrast, in the manual process, an acid accumulation rate as high as 0.36 g L-1 was observed, presumably due to the high nutrient feed rates used to maintain a maximum growth rate. The manual fed-batch process produced a low cell density averaging 10-12 g L-1 (OD600 = 25-30) and plasmid yields of 5-8 mg L-1 (approximately 0.7 mg g-1 cells). The improved plasmid DNA yields in the DO- and pH-based feed-back controlled process were assumed to be a result of a combination of increased cell density, reduced growth rate (mu) from 0.69 h-1 to 0.13 h-1 and the carbon/nitrogen limitation in the fed-batch stage. The DO- and pH-based feed-back control, fed-batch process has proven itself to be advantageous in regulating cell growth rate to achieve both high cell density and plasmid yield without having to use pure oxygen. The process was reproducible in triplicate fermentations at both 7-L and 80-L scales.

Acetic Acid↗
PubMed2000

Computer-automated clinical technique for tinnitus quantification.

This study addresses the need for uniformity in techniques for clinical quantification of tinnitus. Because automation can be an effective means to achieve standardization, this laboratory is developing techniques to perform computer-automated tinnitus testing. The present study was conducted to demonstrate the feasibility of obtaining reliable tinnitus measures using a fully automated system. A computer-controlled psychoacoustical system was developed to quantify tinnitus loudness and pitch using a tone-matching technique. Hearing thresholds were also obtained as part of the procedure. The system generated test stimuli and simultaneously controlled a notebook computer positioned in the sound chamber facing the patient. The notebook computer displayed instructions for responding and relayed response choices through on-screen "buttons" that the patient touched with a pen device. Twenty individuals with tinnitus were evaluated with the technique over two sessions, and responses were analyzed for test-retest reliability. Analyses revealed good reliability of thresholds, loudness matches, and pitch matches. These results demonstrate that use of a fully automated system to obtain reliable measurements of tinnitus loudness and pitch is feasible for clinical application.

Adult↗
PubMed2001

Evaluation of the BacT/Alert automated blood culture system for detecting bacteria and measuring their growth kinetics in leucodepleted and non-leucodepleted platelet concentrates.

BACKGROUND AND OBJECTIVE: To evaluate the BacT/Alert automated blood culture system for the detection of bacteria in platelet concentrates, and to determine bacterial growth kinetics in leucodepleted and non-leucodepleted units. MATERIALS AND METHODS: Apheresis (Cobe Leucocyte Reduction System [LRS]) and pooled buffy coat-derived (Optipress) platelet concentrates (PCs) were tested. Six organisms were used for spiking the PCs: Clostridium perfringens, Bacillus cereus, Group B Streptococcus, Staphylococcus epidermidis, Staphylococcus aureus and Escherichia coli. Units were inoculated to give a final concentration of approximately equal to 1 and 50 colony-forming units (CFU)/ml. On days 0, 2 and 5, BacT/Alert standard aerobic and anaerobic bottles were inoculated with a 5-ml fill volume and bacteria were enumerated. RESULTS: The BacT/Alert Automated blood culture system gave rapid determination times of spiked units, with all positives detected within 48 h and 98.1% detected within 24 h. In general, as the inoculum concentration increased, the detection time decreased. Rapid growth was obtained with all organisms tested except for B. cereus, which failed to grow on four occasions. Bacterial numbers on day 2 ranged from 10(5) to 10(11) CFU/ml and on day 5 ranged from 10(4) to 10(12) CFU/ml. Growth was not significantly greater in leucodepleted units. CONCLUSIONS: The study confirmed that PCs are an excellent growth medium for bacteria. Rapid and substantial growth was obtained with all organisms under test. Leucodepletion does not appear to enhance bacterial proliferation. The BacT/Alert automated blood culture system could rapidly detect contamination of units. Bacterial screening using an automated blood culture system is therefore a potential option.

Bacteria↗
PubMed1999

Interference of Hb-H disease in automated reticulocyte counting.

Hb-H disease is a form of alpha-thalassemia commonly found in south-east Asia. In this condition, numerous Hb-H bodies are found in erythrocytes using reticulocyte staining preparations. This is the first study addressing the possible interference of Hb-H inclusions in automated reticulocyte counting. In this study, seven Hb-H disease samples were tested. Results obtained by the visual method and the Technicon H*3 automated method agreed relatively well. This was in contrast to the Coulter STKS automated method which generally gave lower results. Furthermore, when the incubation time was extended to 180 min in the Coulter STKS method, two Hb-H disease samples gave results several-fold higher than those obtained using the 60 min incubation time. These discrepant results were likely to have been caused by the analyser using the wrong threshold to separate reticulocytes and mature erythrocytes. High and low interference in Hb-H disease samples was observed in the Coulter STKS automated reticulocyte method. Laboratories should be aware of this potential problem, particularly when samples are from patients of Asian origin.

Autoanalysis↗
PubMed2002

Automated screening of blood donations for hepatitis C virus RNA using the Qiagen BioRobot 9604 and the Roche COBAS HCV Amplicor assay.

BACKGROUND AND OBJECTIVES: In order to reduce the potential for transmission of hepatitis C virus (HCV) from an RNA-positive, anti-HCV-negative blood donation, the National Blood Service (NBS) introduced nucleic acid amplification technology (NAT) testing for HCV in England and Wales. The objective of this study was to develop an automated assay using commercial components for the detection of HCV RNA in blood donations for transfusion. MATERIALS AND METHODS: The Qiagen QIAamp 96 'Viral RNA' and 'Virus' BioRobot kits for HCV RNA extraction, and the Roche COBAS HCV Amplicor v2.0 and AmpliScreen v2.0 assays for polymerase chain reaction (PCR) amplification and detection, were investigated. RESULTS: QIAamp technology and the BioRobot 9604 allow automation of the viral RNA extraction process. By combining the automated silica-membrane based QIAamp 96 Virus extraction and automated reverse transcription-polymerase chain reaction (RT-PCR) set-up with COBAS HCV AmpliScreen v2.0 amplification and detection it is possible to achieve a 95% detection level for HCV of 12.8 IU/ml. Cross-contamination studies have shown that use of the BioRobot 9604 does not pose a detectable contamination risk. Between 1999 and 2001, approximately 6.8 x 106 donations were tested in England and Wales, of which only four were found to contain RNA without anti-HCV. CONCLUSIONS: This combination of methods results in an assay with a high sample throughput, little 'hands-on' time and fast turnaround time that is also sufficiently sensitive to allow testing of pools of up to 96 samples at a time. These methods have been successfully introduced into routine use within the NBS for release of blood components with a shelf-life of longer than 24 h.

Blood Donors↗
PubMedDate not supplied

The status of automation and data processing in the United States blood banking community.

A survey of the extent of automation and data processing in the blood banking community was conducted by questionnaire. The likelihood that a facility had some type of automation was related to the volume of blood products produced or transfused. Facilitates that collected blood or collected and transfused blood were more likely to have automation for ABO and Rh testing than those that only transfused blood. Automation for many other blood bank tests is unavailable. Many facilities had data processing capability, which is used primarily for accounting.

Blood Banks↗
PubMedDate not supplied

An automated screening test for antibodies to human immunodeficiency virus-1.

An automated pretransfusion testing system was applied to HIV antibody screening by a gelatin particle agglutination (HIV-PA) test. The test conditions for the test, such as diluent, plate shape, and incubation time, were applicable not only for the HIV-PA test but also for other routine tests, including those for hepatitis B surface antigen and HTLV-1 antibodies. After a 60-minute incubation, the plates were read automatically and then were assessed visually. Tests with seropositive samples obtained from hemophiliacs and an HIV enzyme immunoassay (EIA) familiarization panel showed that the automated HIV-PA test was more sensitive than the EIA and did not show a false-negative result. Of 11,300 blood donors, 44 were positive by the automated test, and 51 were positive by the EIA. None of the blood donors was confirmed to be seropositive. Using the automated test, a large number of samples could be screened in a relatively short time without a significant increase of nonspecific reactions.

Agglutination Tests↗
PubMedDate not supplied

Density gradient separation of peripheral blood stem cells: comparison of an automated cell processing device and manual methods.

Peripheral blood stem cells were collected from normal donors by leukapheresis on a cell separator. The leukapheresis product contained 1.5 x 10(10) mononuclear cells (MNCs) and was divided into two aliquots that underwent either automated or manual density gradient separation with ficoll-hypaque and subsequent washing. In the automated process, recovery of MNCs was 85 percent, reduction in platelet content was 64 percent, and the final hematocrit (Hct) was less than 1 percent. The manual separation resulted in 76-percent MNC recovery, a 79-percent reduction in platelet content, and a final Hct of less than 1 percent. The purified MNCs were then placed in methylcellulose culture at a concentration of 4 x 10(5) MNCs per mL. Quadruplicate 1-mL aliquots were cultured, and colonies were counted and classified on Day 14. Comparison of automated and manual ficoll-hypaque separations demonstrated no differences in the total, erythroid, or granulocyte-macrophage colony numbers. The cell processor used is fast, reliable, uncomplicated, and provides a sterile product containing progenitor cells that are not adversely affected by the automated ficoll-hypaque separation.

Cell Separation↗
PubMed1998

Evaluation of an automated microbiologic blood culture device for detection of bacteria in platelet components.

BACKGROUND: Automated culture methods have been used by several investigators to detect bacterial contamination of cellular blood components. We investigated several factors affecting detection by automated culture of bacteria in platelet concentrates (PCs). These factors included the initial contamination level in PCs, the PC sample volume, the PC sample time, and the white cell level in relation to bacteria levels in the PCs. STUDY DESIGN AND METHODS: Staphylococcus epidermidis or Escherichia coli was inoculated into freshly prepared PCs or white cell-reduced PCs to yield colony-forming unit (CFU) levels of 10, 1, or 0.1 per mL. At the time of inoculation (t=0) and at t=6, t=24, and t=48 hours, 0.5, 1.0, and 2.0 mL samples of the contaminated PCs were transferred into culture bottles. The presence of bacteria in the culture bottles was subsequently monitored by an automated blood culturing instrument. Bacteria levels in the PC at the time of first automated culture detection were determined by quantitative plating. RESULTS: E. coli was detected in 92 percent of experiments when 1.0- or 2.0-mL samples were taken at t=6 hours. At t=24 hours, 100-percent detection was observed with all tested inoculation volumes; however, by that time, >10(7) CFU per mL of bacteria were present in every PC. For S. epidermidis, 89 percent and 83 percent of contaminated PCs were detected with a t=24 hour sampling time and 2.0- or 1.0-mL sampling volume. Seven of 36 PCs with a 2.0-mL sampling volume and 10 of 36 PCs with a 1.0-mL sampling volume contained >10(6) CFU per mL of S. epidermidis at the time of first detection. CONCLUSION: Data from this preliminary evaluation suggest that sampling times of 24 hours or more would be necessary to provide confidence in detection of E. coli or S. epidermidis in PCs using this culture method.

Bacterial Infections↗
PubMed2001

Automation of nucleic acid extraction for NAT screening of individual blood units.

BACKGROUND: Automation of NAT for single units of blood is currently hampered by the labor-intensive steps involved in the extraction of nucleic acids from samples before the amplification procedures. A new method has been developed for the automation of these steps using hydrophilic polyvinylidene fluoride (PVDF) filter plates. STUDY DESIGN AND METHODS: Quantitative nucleic acid recoveries from sera containing HCV, HIV, HBV, HAV, and human parvovirus B19 and from 3H-labeled HCV RNA were determined in parallel by the semi-automated PVDF method and a single-column method (Qiagen). Quantitative PCR was performed. RESULTS: Similar recoveries of HCV, HIV, and HBV (with silica beads) were observed with the PVDF method and with the Qiagen single-column method. The sensitivity of the PVDF-based PCR assay for HCV, HIV, and HBV in serially diluted serum samples was always within two serial dilutions of that obtained when the Qiagen single-column method was used in the same assays. With the use of 3H-labeled HCV RNA, recoveries of approximately 70 percent were found by both methods. CONCLUSION: The PVDF method will permit full automation of the simultaneous extraction of nucleic acid from sera containing HCV, HIV, and HBV. This procedure will permit NAT screening of individual units of blood, will replace the current screening of pools, and will achieve improved blood safety with reduced labor and costs.

Blood Transfusion↗
PubMed2003

Impact of tissue harmonic imaging in patients with distorted left ventricles: improvement in accuracy and reproducibility of visual, manual and automated echocardiographic assessment of left ventricular ejection fraction.

AIMS: The aim of this study was to assess the incremental value of tissue harmonic imaging vs conventional echocardiography for evaluating left ventricular ejection fraction by manual and automated quantitation as well as visual estimation in patients with distorted left ventricles. METHODS AND RESULTS: In 25 patients unselected for image quality and with distorted left ventricles who underwent a nuclear study, digital cineloops of standard apical views were acquired by both tissue harmonic imaging and conventional echocardiography and sent to six observers for analysis of visual and quantitative left ventricular ejection fraction. Tissue harmonic imaging improved both the correlation and agreement of all echo techniques with nuclear measures, compared with conventional echocardiography echo, reducing standard errors (SE) to below 10%: for the visual estimate SE=7.5%, for manual tracing SE=6.3% and for automated tracing SE=8%. Tissue harmonic imaging decreased inter-observer variability compared with conventional echocardiography echo for both visual assessment (12.4% vs 18.4%, P<0.05) and quantitative measures (for manual tracing, 8.2% vs 11.8%, P<0.05; for automated tracing, 7.8% vs 16.8%, P<0.05). CONCLUSIONS: In patients with distorted left ventricles unselected for image quality, tissue harmonic imaging improves accuracy and reproducibility of both visual and quantitative echocardiographic assessment of left ventricular ejection fraction. In particular, it promotes automated quantitation by reducing its high standard error into a clinically reasonable range.

Aged↗