Inconclusive decisions and error rates in forensic science.

In recent years, there has been discussion and controversy relating to the treatment of inconclusive decisions in forensic feature comparison disciplines when considering the reliability of examination methods and results. In this article, we offer a brief review of the various viewpoints and suggestions that have been recently put forth, followed by a solution that we believe addresses the treatment of inconclusive decisions. We consider the issues in the context of method conformance and method performance as two distinct concepts, both of which are necessary for the determination of reliability. Method conformance relates to an assessment of whether the outcome of a method is the result of the analyst's adherence to the procedures that define the method. Method performance reflects the capacity of a method to discriminate between different propositions of interest (e.g., mated and non-mated comparisons). We then discuss implications of these issues for the forensic science community.

Therapeutic drug monitoring of anti-epileptic drugs - a clinical verification of volumetric absorptive micro sampling.

Background Therapeutic drug monitoring (TDM) of antiepileptic drugs (AEDs) can serve as a valuable tool in optimising and individualising epilepsy treatment, especially in vulnerable groups such as pregnant women, the elderly and children. Unfortunately, TDM is often performed suboptimally due to limitations in blood collection. Therefore, we investigated volumetric absorptive micro sampling (VAMS) - a new home-sampling technique. We aimed to evaluate VAMS to determine and quantify the different AEDs and concentrations of 16 different AEDs in whole blood collected by VAMS. Methods Patient blood samples (n = 138) were collected via venepunctures at the Academic Centre for Epileptology Kempenhaeghe. AED concentrations were determined, and these concentrations were used to compare the VAMS method (whole blood) with the conventional method (serum). In addition, the recovery was examined as well as the impact of haematocrit. Finally, AED-spiked blood was used to test the stability of the AEDs inside the micro-sampler devices over a period of time and whether temperature had an effect on the stability. Results VAMS allows for an accurate detection of 16 different AEDs within 2 days after sampling. Deviation in recovery was less than 10% and high correlations were found between VAMS and conventional sampling. Moreover, haematocrit does not have an effect with values between 0.3 and 0.5 (L/L). Finally, although storage temperature of VAMS does affect some AEDs, most are unaffected. Conclusions VAMS enables an accurate detection of a wide variety of AEDs within 2 days after sampling.

Reconstruction of defects involving the Achilles tendon and local soft tissues: a quick solution for a lingering problem.

A total of seven patients (six men and one woman) with a defect in the Achilles tendon and overlying soft tissue underwent reconstruction using either a composite radial forearm flap (n = 3) or an anterolateral thigh flap (n = 4). The Achilles tendons were reconstructed using chimeric palmaris longus (n = 2) or tensor fascia lata (n = 2) flaps or transfer of the flexor hallucis longus tendon (n = 3). Surgical parameters such as the rate of complications and the time between the initial repair and flap surgery were analysed. Function was measured objectively by recording the circumference of the calf, the isometric strength of the plantar flexors and the range of movement of the ankle. The Achilles tendon Total Rupture Score (ATRS) questionnaire was used as a patient-reported outcome measure. Most patients had undergone several previous operations to the Achilles tendon prior to flap surgery. The mean time to flap surgery was 14.3 months (2.1 to 40.7). At a mean follow-up of 32.3 months (12.1 to 59.6) the circumference of the calf on the operated lower limb was reduced by a mean of 1.9 cm (sd 0.74) compared with the contralateral limb (p = 0.042). The mean strength of the plantar flexors on the operated lower limb was reduced to 88.9% of that of the contralateral limb (p = 0.043). There was no significant difference in the range of movement between the two sides (p = 0.317). The mean ATRS score was 72 points (sd 20.0). One patient who had an initial successful reconstruction developed a skin defect of the composite flap 12 months after free flap surgery and this resulted in recurrent infections, culminating in transtibial amputation 44 months after reconstruction. These otherwise indicate that reconstruction of the Achilles tendon combined with flap cover results in a successful and functional reconstruction.

Finite-element modelling reveals force modulation of jaw adductors in stag beetles.

Male stag beetles carry large and heavy mandibles that arose through sexual selection over mating rights. Although the mandibles of Cyclommatus metallifer males are used in pugnacious fights, they are surprisingly slender. Our bite force measurements show a muscle force reduction of 18% for tip biting when compared with bites with the teeth located halfway along the mandibles. This suggests a behavioural adaptation to prevent failure. We confirmed this by constructing finite-element (FE) models that mimic both natural bite situations as well as the hypothetical situation of tip biting without muscle force modulation. These models, based on micro-CT images, investigate the material stresses in the mandibles for different combinations of bite location and muscle force. Young's modulus of the cuticle was experimentally determined to be 5.1 GPa with the double indentation method, and the model was validated by digital image correlation on living beetles. FE analysis proves to be a valuable tool in the investigation of the trade-offs of (animal) weapon morphology and usage. Furthermore, the demonstrated bite force modulation in male stag beetles suggests the presence of mechanosensors inside the armature.

Denitrification of high nitrate concentration wastewater using alternative carbon sources.

The use of different organic carbon sources in the denitrification of wastewater containing 2500 mg nitrates/L in a SBR was studied. Three alternative sources of carbon were tested: wastewater from a sweet factory, a residue from a soft drinks factory and a residue from a dairy plant. The first two are sugar-rich, whereas the third presents a high content in lactic acid. Maximum specific denitrification rates of between 42 and 48 mg NO(3)-N/g VSS h were obtained. The effluents were nitrate-free and very low COD concentrations were obtained in 4-6h reaction time, especially with the sugar-rich carbon sources. The values of the denitrifier net yield coefficient were higher than when using methanol (0.93-1.75 g VSS(formed)/g NO(x)-N(reduced)). The lowest value was obtained using the lactic acid-rich residue. The optimum COD/N ratios varied between 4.6 for the lactic acid-rich carbon source and 5.5-6.5 for the sugar-rich carbon sources.

Denitrification of wastewater containing high nitrate and calcium concentrations.

The removal of nitrate from rinse wastewater generated in the stainless steel manufacturing process by denitrification in a sequential batch reactor (SBR) was studied. Two different inocula from wastewater treatment plants were tested. The use of an inoculum previously acclimated to high nitrate concentrations led to complete denitrification in 6h (denitrification rate: 22.8mg NO3- -N/gVSSh), using methanol as carbon source for a COD/N ratio of 4 and for a content of calcium in the wastewater of 150mg/L. Higher calcium concentrations led to a decrease in the biomass growth rate and in the denitrification rate. The optimum COD/N ratio was found to be 3.4, achieving 98% nitrate removal in 7h at a maximum rate of 30.4mg NO3- -N/gVSSh and very low residual COD in the effluent.

Information Model for Machine-Tool-Performance Tests.

This report specifies an information model of machine-tool-performance tests in the EXPRESS [1] language. The information model provides a mechanism for describing the properties and results of machine-tool-performance tests. The objective of the information model is a standardized, computer-interpretable representation that allows for efficient archiving and exchange of performance test data throughout the life cycle of the machine. The report also demonstrates the implementation of the information model using three different implementation methods.

Comparative study of pressure- and nutrient-induced germination of Bacillus subtilis spores.

Germination experiments with specific germination mutants of Bacillus subtilis, including a newly isolated mutant affected in pressure-induced germination, suggest that a pressure of 100 MPa triggers the germination cascades that are induced by the nutrient germinant alanine (Ala) and by a mixture of asparagine, glucose, fructose, and potassium ions (AGFK), by activating the receptors for alanine and asparagine, GerA and GerB, respectively. As opposed to germination at 100 MPa, germination at 600 MPa apparently short-cuts at least part of the Ala- and AGFK-induced germination pathways. Inhibitors of nutrient-induced germination (HgCl(2) and Nalpha-P-tosyl-L-arginine methyl ester) also inhibit pressure-induced germination at 600 MPa, suggesting that germination at 600 MPa involves activation of a true physiological germination pathway and is therefore not merely a physico-chemical process in which water is forced into the spore protoplast.

Similar alterations of lymphocyte subpopulations in type I and type II diabetes.

Peripheral blood lymphocytes of diabetes mellitus patients were analyzed by flow-cytometry using monoclonal antibodies directed against cell surface markers present in T- and B-cells, monocytes and natural killer cells. The lymphocyte subsets were quantified and expressed in an absolute amount. The study included 17 patients with type I (insulin-dependent), 21 patients with type II (non-insulin-dependent) diabetes and 40 age-matched control subjects. Quantification of the cells present within different lymphocyte subsets revealed a general increase in both patient groups compared to their controls, with the exception of activated T-cells. However, no significant difference was found in the relative amount of T-helper cells and T-suppressor/cytotoxic cells of the diabetes patients when they were compared with their control groups. The fact that we found similar changes in lymphocyte subsets in both type I and type II diabetes suggests that the altered immunological state is secondary to the diabetes mellitus in general.

A time-resolved fluoroimmuno assay of the IgM-rheumatoid factor.

A time-resolved fluoroimmuno assay of the IgM-rheumatoid factor is described. Aggregated rabbit IgG was coated to microtitre plates to serve as the target protein. F(ab')2-fragments from antibodies, raised in rabbits against human IgM, were labelled with Eu3+ and used in the assay to mark the bound IgM-rheumatoid factor. The labelling procedure is easy to perform, and there is no need for special equipment. The shelf life of the label at -20 degrees C is at least one year. The lower detection limit of the assay is 1.3 x 10(3) IU/l. The range over which the IgM-rheumatoid factor can be measured at a within-run precision of less than 5% without varying the dilution (working range) is 5-1200 x 10(3) IU/l. Linearity in serum dilutions is good. There is good correlation with existing methods for the assay of IgM-rheumatoid factor. This correlation is better with an assay using rabbit IgG as the target than with one using human IgG. Comparison of methods shows that standardization, despite the use of the WHO Reference Preparation as the first calibrator, remains problematic. The 95th percentile in normal bloodbank donors is 8 x 10(3) IU/l. The costs for the reagents were about 0.5 Dutch florin (ca 0.30 US-$) per well. In conclusion, the method described here is analytically at least comparable with other methods, in precision, linearity, working range etc. Finally, it is easy to perform.

Haemodialysis-induced changes in red blood cell size-distribution histograms.

Treatment by haemodialysis may result in changes of the red blood cell size-distribution histograms in most subjects. After maintenance haemodialysis a statistically significant shift to the left is observed for the mean size distribution histogram in a group consisting of sixteen women. The shift is in agreement with the finding of significantly higher values for the microcyte fraction, as well as with significantly lower mean cell volume (MCV) values. No significant changes were observed for the macrocytic fractions of red blood cells.

Changes in size and zinc protoporphyrin/haemoglobin ratio in red cells of infants during the first months of life.

A longitudinal investigation of changes in red blood cell (RBC) size distribution and zinc protoporphyrin/haemoglobin (ZPP/Hb) ratio was performed. In the course of the first few weeks after birth RBC size distribution histograms show a loss of macrocytic RBCs demonstrated by a rapid decrease of the fraction of macrocytes, whereas the degree of RBC volume dispersion decreases simultaneously. Within this period a significant change in the fraction of microcytes was not observed. At about 6 months of age, a marked shift of RBC size distribution histograms towards lower volumes occurs. Concomitant with the lower MCV values, the values for the absolute distribution width at half peak height (ADW0.5) also showed an obvious decrease. At birth, ZPP/Hb ratios are about three times higher than those of adult subjects and decrease slowly during infancy. This suggests that iron deficiency is unlikely in the first months of life.

Investigations on red cell size distribution histograms in subjects treated by maintenance haemodialysis.

In forty-nine subjects on maintenance haemodialysis quantitative information with respect to changes in the erythron was derived from the red blood cell size distribution histogram. Increased values for the fraction of microcytes were observed in seventeen subjects, whereas in three subjects increased fractions of macrocytes were established. Both fractions were found to be increased simultaneously in only one subject. In the case of normal mean corpuscular volume (MCV) values, the fractions of microcytes were shown to be increased in four male and three female subjects. In contrast, MCV values were increased in four male subjects while the fractions of macrocytes did not exceed the reference range. Several causes which may give rise to abnormalities in red blood cell size distribution histograms in subjects treated by haemodialysis are discussed. From the interpretation of results for serum ferritin concentration, serum iron concentration, total iron binding capacity, and iron saturation percentage it is difficult to establish unequivocally whether the iron supply to the patient is appropriate.

Investigation of red cell size-distribution histograms related to folate, vitamin B12 and iron state in the course of pregnancy.

Effects of gradually reduced stores of iron, folate or vitamin B12 on the erythron were investigated in the course of pregnancy in 23 subjects. Significant decreases in serum concentrations of the compounds above were shown at 20 weeks of gestation with a further decline later. An increased value for the microcyte fraction was established in two subjects, whereas seven other subjects showed increased values for the macrocyte fraction. As pregnancy proceeded a significant intra-individual change could be detected in the microcyte fraction in only one subject. In three of the seven above-mentioned subjects, the values measured for the macrocyte fraction demonstrated a further slight increase during the course of pregnancy. Values established for the absolute distribution width at half peak height (ADW0.5) did not reveal significant changes in the course of pregnancy. If compared with the mean cell volume (MCV) and ADW0.5 values, the microcyte and macrocyte fraction respectively may yield a sensitive and specific indication of anomalies of the erythron. Changes in successively determined haemocytometric values in several pregnant women might indicate decreased availability of nutrients for erythropoiesis. However, one may not draw definite conclusions because it is doubtful whether the condition of the stores in these circumstances will be reflected accurately by the corresponding serum concentrations.

Interference of plasma fluorophores on the red blood cell zinc protoporphyrin: haemoglobin ratio as determined on a haematofluorimeter.

Direct measurement of the zinc protoporphyrin:haemoglobin ratio (ZPP:Hb ratio) in blood samples is performed by using a haematofluorimeter. Interference by non-specific fluorophores can be eliminated by removing the plasma and making the measurement on washed red blood cells (RBCs). After re-suspending RBCs in isotonic saline, haematofluorimeter readings for the ZPP:Hb ratios revealed higher stability in the course of time whereas a good relationship was found with results obtained by application of an extraction method. Separate reference ranges were established for adult male and female subjects. After washing, the mean values calculated for ZPP:Hb ratios of subjects belonging to the reference groups demonstrated a reduction of 0.04 mumol ZPP mol Hb, corresponding with approximately 30%. In the patients' group, application of washing resulted in a variable decrease of ZPP:Hb ratios.

Investigation of red cell size distribution histograms of normal male and female subjects.

Size distribution histograms characterizing the red blood cell (RBC) populations of normal subjects are provided. Separate haemocytometric reference ranges were established for apparently healthy male and female adults by using an Ortho ELT 800 Hematology Analyzer, which is based on the principle of measuring light scattering intensity (LSI). Effects of sphering RBCs on some characteristics of size distribution histograms were also investigated. The absolute distribution width at half peak height (ADW0.5) of RBC size distribution histograms of sphered RBCs is shown to be approximately 40% lower than for unsphered RBCs. The mean RBC size distribution histogram determined for the group of female subjects was shown to be significantly shifted to the right in comparison with the mean size distribution histogram calculated for males, indicating a sex-related difference. The shift is concordant with a significantly higher fraction of microcytes in male subjects, as well as with lower mean corpuscular volume (MCV) values in males compared with females.

Some effects on light scattering intensity and red blood cell size distribution histograms due to sphering.

At present most haematology blood cell analysers routinely provide red blood cell (RBC) size distribution histograms. Sophisticated improvements of the instruments have re-awakened interest in the study of size histograms. The quantitative information derived from the histograms may be applied more fruitfully if insight is available, with respect to some essential principles of sizing technology and methods for treatment of RBCs before measurement. In this study the consequences of sphering RBCs are investigated in relation to the generation of size distribution histograms by means of methods based on light scattering intensity (LSI). Sphering of RBCs results in considerably narrower histograms than upsphered RBCs. The overall signal to noise ratio increases and there is a broader gap between large platelets and microcytic RBCs. Narrower size distribution ranges will enable closer modes to be separated. Compared to unsphered RBCs, microcytic sphered RBCs yield increased LSI whereas macrocytic sphered RBCs yield decreased LSI.

Evaluation of red blood cell size distribution histograms: the fraction of microcytes.

A method is described for the assessment of the fraction of microcytes from the red blood cell (RBC) size distribution histogram. For quantification of the fraction of microcytes an upper threshold corresponding with a RBC volume of 70 fL is established. The reference interval for the fraction of microcytes covers a range from 0.01 to 0.09. In order to examine the relationship between the fraction of microcytes and the mean cell volume (MCV) values computer simulation studies were performed. The computer simulations are based on a Gaussian distributed reference RBC histogram as generated on a Technicon H 6000/H 601 Hematology Analyser. From our studies it is shown that determination of the fraction of microcytes yields a higher sensitivity than MCV values for detecting small populations of microcytes. In particular, measurement of the fraction of microcytes is very sensitive to minor changes in the MCV values if a normocytic RBC population combined with a microcytic RBC fraction shows a wide dispersion in their cell volumes.

Measurements on the forward light scattering intensities of monodisperse latex particles and sphered or unsphered red blood cells.

Characteristics with respect to light scattering intensity (LSI) were measured on latex spheres and on sphered and unsphered red blood cells (RBC). From the discrepancies in LSI signals from polymer spheres and RBCs it is obvious that latices cannot be used for absolute calibration of RBC sizes. Sphering of RBCs did not give rise to a closer resemblance. Monodisperse latex spheres produce extremely narrow size distribution histograms. Regarding these sharp peaks and the long-term stability, polymer spheres offer definite advantages if compared with blood controls to which preservatives were added. A shift from the RBC size distribution histogram, caused by gradually developing instrument drift, can be easily detected and corrected at an early stage.