Quality assessment of Affymetrix GeneChip data

Affymetrix GeneChips are one of the best established microarray platforms. This powerful technique allows users to measure the expression of thousands of genes simultaneously. However, a microarray experiment is a sophisticated and time consuming endeavor with many potential sources of unwanted variation that could compromise the results if left uncontrolled. Increasing data volume and data complexity have triggered growing concern and awareness of the importance of assessing the quality of generated microarray data. In this review, we give an overview of current methods and software tools for quality assessment of Affymetrix GeneChip data. We focus on quality metrics, diagnostic plots, probe-level methods, pseudo-images, and classification methods to identify corrupted chips. We also describe RNA quality assessment methods which play an important role in challenging RNA sources like formalin embedded biopsies, laser-micro dissected samples, or single cells. No wet-lab methods are discussed in this paper.     

Quality assessment of multiple alignment programs

A renewed interest in the multiple sequence alignment problem has given rise to several new algorithms. In contrast to traditional progressive methods, computationally expensive score optimization strategies are now predominantly employed. We systematically tested four methods (Poa, Dialign, T-Coffee and ClustalW) for the speed and quality of their alignments. As test sequences we used structurally derived alignments from BAliBASE and synthetic alignments generated by Rose. The tests included alignments of variable numbers of domains embedded in random spacer sequences. Overall, Dialign was the most accurate in cases with low sequence identity, while T-Coffee won in cases with high sequence identity. The fast Poa algorithm was almost as accurate, while ClustalW could compete only in strictly global cases with high sequence similarity.     

Quality of life assessment in patients with dementia

Quality of life (QoL) is one of the most important outcome variables in the study of the efficacy of interventions with people with dementia. However, its assessment is difficult 1) because it is a complex construct for which there is no unified theoretical or conceptual approach, and 2) because of the inherent difficulties in the cognitive impairments of the people under study. In this work different methods and instruments to this end are reviewed, and related findings are discussed. It is important to take into account the subjective view of the assessed person, as assessments done by proxies tend to underestimate QoL. In spite of the need for further development in this field, it is concluded that the instrument of choice is the QOL-AD, as it is change-sensitive, it correlates with health measurements, it is translated into several languages and it can be administered to people with low MMSE scores.     

Non-invasive measurement of hemoglobin: assessment of two different point-of-care technologies

Background

Measurement of blood hemoglobin (Hb) concentration is a routine procedure. Using a non-invasive point-of-care device reduces pain and discomfort for the patient and allows time saving in patient care. The aims of the present study were to assess the concordance of Hb levels obtained non-invasively with the Pronto-7 monitor (version 2.1.9, Masimo Corporation, Irvine, USA) or with the NBM-200MP monitor (Orsense, Nes Ziona, Israel) and the values obtained from the usual colorimetric method using blood samples and to determine the source of discordance.

Methods and Findings

We conducted two consecutive prospective open trials enrolling patients presenting in the emergency department of a university hospital. The first was designed to assess Pronto-7™ and the second NBM-200MP™. In each study, the main outcome measure was the agreement between both methods. Independent factors associated with the bias were determined using multiple linear regression. Three hundred patients were prospectively enrolled in each study. For Pronto-7™, the absolute mean difference was 0.56 g.L⁻¹ (95% confidence interval [CI] 0.41 to 0.69) with an upper agreement limit at 2.94 g.L⁻¹ (95% CI [2.70;3.19]), a lower agreement limit at -1.84 g.L⁻¹ (95% CI [-2.08;-1.58]) and an intra-class correlation coefficient at 0.80 (95% CI [0.74;0.84]). The corresponding values for the NBM-200MP™ were 0.21 [0.02;0.39], 3.42 [3.10;3.74], -3.01 [-3.32;-2.69] and 0.69 [0.62;0.75]. Multivariate analysis showed that age and laboratory values of hemoglobin were independently associated with the bias when using Pronto-7™, while perfusion index and laboratory value of hemoglobin were independently associated with the bias when using NBM-200MP™.

Conclusion

Despite a relatively limited bias in both cases, the large limits of agreement found in both cases render the clinical usefulness of such devices debatable. For both devices, the bias is independently and inversely associated with the true value of hemoglobin.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01321580","term_id":"NCT01321580"}}NCT01321580 {"type":"clinical-trial","attrs":{"text":"NCT01321593","term_id":"NCT01321593"}}NCT01321593     

Quality assessment and analysis of Biogen Idec compound library

A subset of the compound repository for lead identification at Biogen Idec was characterized for its chemical stability over a 3-year period. Compounds were stored at 4 degrees C as 10 mM DMSO stocks, and a small subset of compounds was stored as lyophilized dry films. Compound integrity of 470 discrete compounds (Compound Set I) and 1917 combinatorial chemistry-derived compounds (Compound Set II) was evaluated by liquid chromatography/mass spectrometry from the time of acquisition into the library collection and after 3 years of storage. Loss of compound integrity over the 3 years of storage was observed across the 2 subsets tested. Of Compound Set I, 63% of samples retained > 80% purity, whereas 57% of samples from Compound Set II had purity greater than 60%. The stability of the lyophilized samples was superior to the samples stored as DMSO solution. Although storage at 4 degrees C as DMSO solution was adequate for the majority of compounds, the authors observed and quantified the level of degradation within the compound collection. Their study provides general insight into compound storage and selection of library subsets for future lead identification activities.     

Quality assessment of cryopreserved biospecimens reveals presence of intact biomolecules

Recapitulation of tumor features in isolated biomolecules is preeminently dependent on obtaining reliable quality biospecimen. Moreover, quality assessment of biobanked specimens at regular intervals is an essential intervention for carrying out effective translational and clinical research. In the current study, genomic DNA was extracted from 140 fresh frozen tissues of oral, breast and colorectal specimens cryopreserved over a period of 3 to 8 months (short term) and 3 to 4 years (long term). Quantification of genomic DNA by absorption and fluorescence spectroscopy confirmed high concentration while qualitative analysis by gel electrophoresis showed intact bands for 94% and 87% of short‐ and long‐term cohorts, respectively. PC‐LDA based classification of Raman spectra showed overlapping groups of both cohorts suggesting the quality of DNA being preserved irrespective of storage period. To the best of our knowledge this is the first Indian biobank study reporting quality analysis of biospecimens cryopreserved at different time periods.     

Quality assessment of confocal microscopy slide-based systems: instability.

BACKGROUND: All slide-based fluorescence cytometry detections systems basically include an excitation light source, intermediate optics, and a detection device (CCD or PMT). Occasionally, this equipment becomes unstable, generating unreliable and inferior data. METHODS: A number of tests have been devised to evaluate equipment performance and instability. The following four instability tests are described: galvanometer scanning, stage drift, correct wavelength spectral detection, and long-term laser power. RESULTS: Quality assurance tests revealed that a confocal microscope can become unstable in the following parameters, yielding inaccurate data: laser power, PMTs functionality, spectrophotometer accuracy, galvanometer scanning and laser stability, and stage drift. Long-term laser power stability has been observed to vary greatly. CONCLUSIONS: Confocal systems can become unstable in the following parameters: long-term laser power, galvanometer scanning, spectrophotometer accuracy, and stage stability. Instability in any of these parameters will affect image quality. Laser power fluctuations result from either a defective Acousto-optic tunable filter or improper heat dissipation. Spectrophotometer instability will generate unreliable spectra data, extra light reflections, and poor image quality. Galvanometer scanning instability yields poor image quality while microscope stage drift results in a sample going out of the plane of focus. With minor modifications, these tests may be applicable to other slide-based systems.     

Performance assessment of protein multiple sequence alignment algorithms based on permutation similarity measurement

Protein multiple sequence alignment is an important bioinformatics tool. It has important applications in biological evolution analysis and protein structure prediction. A variety of alignment algorithms in this field have achieved great success. However, each algorithm has its own inherent deficiencies. In this paper, permutation similarity is proposed to evaluate several protein multiple sequence alignment algorithms that are widely used currently. As the permutation similarity method only concerns the relative order of different protein evolutionary distances, without taking into account the slight difference between the evolutionary distances, it can get more robust evaluations. The longest common subsequence method is adopted to define the similarity between different permutations. Using these methods, we assessed Dialign, Tcoffee, ClustalW and Muscle and made comparisons among them.     

Quality assessment of confocal microscopy slide based systems: performance.

BACKGROUND: All fluorescence slide-based cytometry detections systems basically include the following components: (1) an excitation light source, (2) intermediate optics, and (3) a detection device consisting of a CCD camera or a PMT. The optical principles employed is slide-based systems are similar to those of confocal microscopes (CLSM). METHODS: The following tests evaluated confocal equipment performance: dichroic reflectivity, field illumination, lens performance, laser power output, spectral registration, axial resolution, PMT reliability, and system noise. RESULTS: Quality assurance tests provide a basis to determine if the equipment is operating correctly. Laser power, PMTs function, dichroic reflection, spectral registration, axial registration, system noise and sensitivity, lens performance and laser stability were tested colocalization of UV and visible peaks of a bead should be less than 210 nm. Interference contrast optics decrease fluorescence resolution. CONCLUSIONS: QA tests that assess CLSM system performance are also applicable to other slide-based systems. By utilization this type of testing approach, the subjective nature of assessing the CLSM may be eliminated. These tests serve as guidelines for other investigators to ensure that their machines are providing data that is accurate with the necessary resolution, sensitivity and precision.     

Quality assessment of compost prepared from fly ash and crop residue

Fly ash was co-composted with wheat straw and 2% rock phosphate (w/w) for 90 days and different chemical and microbiological parameters monitored to evaluate its effect on the composting process. Fly ash addition at 20% level resulted in the lowest C/N of 16.4:1 and highest available and total phosphorus. Increasing the addition of fly ash from 40 to 60% (w/w) did not exert any detrimental effect on either C:N or the microbial population.     

Hydrogen clearance: assessment of technique for measurement of skin-flap blood flow in pigs

The hydrogen clearance technique has been used for many years by investigators to determine brain blood flow and has been partially validated in this setting using other methods of blood flow measurement. The method has been modified to allow blood flow measurements in skin, but the accuracy of H2 clearance for measuring skin blood flow has not been determined. Multiple blood flow measurements were performed using H2 clearance and radioactive microspheres on skin flaps and control skin in pigs. On 12 pigs, a total of 117 flap and 42 control skin measurements were available for analysis. There was no significant difference between the two techniques in measuring mean control skin blood flow. In skin flaps, H2 clearance was significantly correlated to microsphere-measured blood flow, but it consistently gave an overestimate. Sources of error may include injury to the tissues by insertion of electrodes, consumption of H2 by the electrodes, or diffusion of H2 from the relatively ischemic flap to its well-vascularized bed. Further studies are necessary to determine the cause of this error and to measure the technique's accuracy in skeletal muscle and other flaps.     

Quality assessment of tandem mass spectra based on cumulative intensity normalization

A large proportion of MS/MS spectral analyses do not result in significant matches because their spectral quality is too poor to produce meaningful identification. Throughput of peptide identification can be greatly improved, if one can filter out, in advance, spectra that would lead to wrong identification. We introduce here an innovative approach to assess spectral quality utilizing a new spectral feature called Xrea, based on cumulative intensity normalization.     

A novel measurement of allele discrimination for assessment of allele-specific silencing by RNA interference

Allele-specific silencing by RNA interference (ASP-RNAi) is an atypical RNAi that is capable of discriminating target alleles from non-target alleles, and may be therapeutically useful for specific inhibition of disease-causing alleles without affecting their corresponding normal alleles. However, it is difficult to design and select small interfering RNA (siRNAs) that confer ASP-RNAi. A major problem is that there are few appropriate measures in determining optimal allele-specific siRNAs. Here we show two novel formulas for calculating a new measure of allele-discrimination, named “ASP-score”. The formulas and ASP-score allow for an unbiased determination of optimal siRNAs, and may contribute to characterizing such allele-specific siRNAs.