Measurement Of Analyte Concentration Research Articles

Semiquantitative analysis on microfluidic thread-based analytical devices by ruler

This paper describes a simple and semiquantitative method for analytical detection by measuring the length of colour change on indicator treated threads using a ruler. Thread-based analytical devices (μTADs), fabricated from two types of threads (cotton and polyester), provide an easy-to-use platform for the rapid measurement of analyte concentrations in aqueous solutions. Multiple threads were fixed to a supporting polymer film to allow capillary wicking of liquid samples, free from contact with outside surfaces. In this method, interaction between deposited reagents and analytes within samples produces coloured zones of differing lengths on the threads. The length of the coloured zones correlates with the concentrations of the analytes in the samples. The diagnostic application of the length measurement method, using μTADs, was demonstrated through the successful testing of two different bioassays, for protein and nitrite, in simulated human urine samples. Additionally, it was shown that this method can be used for the rapid analysis of environmental samples such as Ni2+ in water. Furthermore, the length measurement method can be applied to perform simultaneous assays to quantify the concentrations of different biomarkers present within the same sample. Finally, it was also compared with currently used colorimetric methods on thread, which were reported by us and others previously, and was shown to have higher accuracy and simplicity.

Interference in Antiphospholipid Antibody Assays

Antiphospholipid antibodies (aPL) are detected with two types of laboratory tests: first, antigen-specific immunoassays for the determination of antibodies against cardiolipin, β2-glycoprotein I and other phospholipids, and phospholipid-protein complexes; second, functional (coagulation) assays for the detection of lupus anticoagulants. Both aPL immunoassays and coagulation assays are prone to interferences, and clinicians need to be aware of the limitations of these assays. Interference is a clinically significant bias in the measured analyte concentration due to the effect of another component or property of the sample. Besides immune-mediated interferences (such as heterophile or human anti-animal antibodies, rheumatoid factor, high immunoglobulin levels, or factor inhibitors), aPL assays are uniquely affected by anticoagulants and the presence of residual platelets in test plasma. Interferences are usually analyte- and assay-specific and may go unrecognized in routine laboratory practice. Despite advances in our knowledge on the mechanisms of interferences in aPL assays, it is unlikely that total elimination will be possible.

Selective pressurized liquid extraction of three classes of halogenated contaminants in fish

Pressurized liquid extraction (PLE) combined with in-cell clean-up of co-extracts, so-called selective-PLE (S-PLE), is a fast and accepted method for the analysis of halogenated organic contaminants in fish. However, many of the existing methods were optimized for use with single classes of contaminants. The main objective of this research was to develop an S-PLE method that elutes a minimal amount of fats while simultaneously extracting halogenated pesticides, polychlorinated biphenyls (PCBs), and polybrominated diphenyl ethers (PBDEs) from fat-rich fish. The optimized method uses n-hexane:dichloromethane (75:25, v/v) as the extraction solvent, a 0.0078 fat to fat-retainer ratio (FFR) with Florisil as the fat retainer, three individual 5-min extractions with flush volumes of 150% and a selection of labeled surrogate standards (isotope dilution). This method resulted in a mean recovery of 77% for all target analytes in spiked samples and an average relative standard deviation of 6.3%. The method was validated with a certified reference material; the mean measured analyte concentrations agreed with the reference values except in the case of individual endosulfan isomers. It is likely that interconversion from the beta- to alpha-endosulfan isomer had occurred in the CRM, resulting in low measured concentrations for beta-endosulfan and high measured concentrations for alpha-endosulfan when compared with the reference values. Finally, the method was tested on three fish species with varying fat content. Different contaminant patterns were observed in the various species.

Winemaking Process Monitoring Based on a Biosensor Automatic System

This paper proposes an automatic measurement system for monitoring of biological processes using amperometric biosensors in flow injection analysis. Whereas this kind of measurement usually requires intervention of human experts, the proposed system allows full automation of the measurement process. Both flow injection and analyte concentration measurement processes are managed without human interactions. In this paper, a first prototype of the system customized for winemaking process monitoring is described and characterized.

Erroneous augmentation of multiplex assay measurements in patients with rheumatoid arthritis due to heterophilic binding by serum rheumatoid factor

Serum rheumatoid factor (RF) and other heterophilic antibodies potentially interfere with antibody-based immunoassays by nonspecifically binding detection reagents. The purpose of this study was to assess whether these factors confound multiplex-based immunoassays, which are used with increasing frequency to measure cytokine and chemokine analytes in patients with rheumatoid arthritis (RA). We performed multiplex immunoassays using different platforms to measure analyte concentrations in RA patient samples. Samples were depleted of RF by column-based affinity absorption or were exposed to agents that block heterophilic binding activity. In RA patients with high-titer RF, 69% of analytes demonstrated at least a 2-fold stronger multiplex signal in non-RF-depleted samples as compared to RF-depleted samples. This degree of erroneous signal amplification was less frequent in low-titer RF samples (17% of analytes; P < 0.0000001). Signal amplification by heterophilic antibodies was blocked effectively by HeteroBlock (≥ 150 μg/ml). In 35 RA patients, multiplex signals for 14 of 22 analytes were amplified erroneously in unblocked samples as compared to blocked samples (some >100-fold), but only in patients with high-titer RF (P < 0.002). Two other blocking agents, heterophilic blocking reagent and immunoglobulin-inhibiting reagent, also blocked heterophilic activity. All multiplex protein detection platforms we tested exhibited significant confounding by RF or other heterophilic antibodies. These findings have broad-reaching implications in the acquisition and interpretation of data derived from multiplex immunoassay testing of RA patient serum and possibly also in other conditions in which RF or other heterophilic antibodies may be present. Several available blocking agents effectively suppressed this erroneous signal amplification in the multiplex platforms tested.

Comparison of the Use of Volume Fractions with other Measures of Concentration for Quantitative Spectroscopic Calibration Using the Classical Least Squares Method

Since the commercial development of modern near-infrared spectroscopy in the 1970s, analysts have almost invariably used units of weight percent as the measure of analyte concentration, due largely to the historical precedent from other analytical methods, including other spectroscopic techniques. The application of the CLS algorithm to a set of binary and ternary liquid mixtures reveals that the spectroscopic measurement sees the sample differently; that the measured absorbance spectrum is in fact sensitive to the volume fraction of the various components of the mixture. Because there is not a one-to-one relationship between volume fraction and other measures of analyte concentration, nor is the relationship linear, this has important implications for the application of both the CLS algorithm and the various other, more conventional, calibration algorithms that are commonly used.

Confocal Fluctuation Spectroscopy and Imaging

Currently, work with subnanomolar concentrations is routine while femtomolar and even single-molecule studies are possible with some efforts getting high on single-molecule biophysics and biochemistry. Methodological breakthroughs, such as reducing the background light contribution in single-molecule studies, which has plagued many studies of molecular fluorescence in dilute solution, and particularly in live cells, have recently described by us. We first demonstrated how optimized time-gating of the fluorescence signal, together with time-correlated, single-photon counting, can be used to substantially boost the experimental signal-to-noise ratio about 140-fold, making it possible to measure analyte concentrations that are as low as 15 pM. By detection of femtomolar bulk concentrations, confocal microsopy has the potential to address the observation of one and the same molecule in dilute solution without immobilization or hydrodynamic/electrokinetic focusing at longer observation times than currently available. We present relevant physics. The equations are derived using Einstein's approach showing how it fits with Fick's law and the autocorrelation function. An improved technology is being developed at ISS for femtomolar microscopy. The general concepts and provided experimental examples should help to compare our approach to those used in conventional confocal microscopy.

Matrix solid-phase dispersion on column clean-up/pre-concentration as a novel approach for fast isolation of abuse drugs from human hair

A simple and fast sample pre-treatment method based on matrix solid-phase dispersion (MSPD) for isolating cocaine, benzoylecgonine (BZE), codeine, morphine and 6-monoacethylmorphine (6-MAM) from human hair has been developed. The MSPD approach consisted of using alumina (1.80 g) as a dispersing agent and 0.6 M hydrochloric acid (4 mL) as an extracting solvent. For a fixed hair sample mass of 0.050 g, the alumina mass to sample mass ratio obtained was 36. A previously conditioned Oasis HLB cartridge (2 mL methanol, plus 2 mL ultrapure water, plus 1 mL of 0.2 M/0.2 M sodium hydroxide/boric acid buffer solution at pH 9.2) was attached to the end of the MSPD syringe for on column clean-up of the hydrochloric acid extract and for transferring the target compounds to a suitable solvent for gas chromatography (GC) analysis. Therefore, the adsorbed analytes were directly eluted from the Oasis HLB cartridges with 2 mL of 2% acetic acid in methanol before concentration by N 2 stream evaporation and dry extract derivatization with N-methyl-tert-butylsilyltrifluoroacetamide (BSTFA) and chlorotrimethylsilane (TMCS). The optimization/evaluation of all the factors affecting the MSPD and on column clean-up procedures has led to a fast sample treatment, and analytes extraction and pre-concentration can be finished in approximately 30 min. The developed method has been applied to eight hair samples from poli-drug abusers and measured analyte concentrations have been found to be statistically similar (95% confidence interval) to those obtained after a conventional enzymatic hydrolysis method (Pronase E).

Electrically stretched capacitive membranes for stiffness sensing and analyte concentration measurement

A novel method for stiffness sensing is developed using an electret capacitive sensor. The electret membrane is coated with a recognition layer that responds with a change in its stiffness/elasticity in the presence of a target analyte. Since the electret membrane is stretched by electrostatic pulling towards a metallic base plate, a change in stiffness of the composite membrane results in deflection of the membrane. This deflection is measured as a change in capacitance of the sensor. The sensitivity of the sensor to stiffness changes depends on the strength of the preset electric field. The developed sensor operates in a quasi-static mode and eliminates the need for resonant monitoring. The sealed capacitive sensor is ideal for monitoring analytes in both gas and liquid environments. The final sensor package with the capacitance measurement circuitry has a low power consumption (<30 mW). A proof-of-concept carbon dioxide gas concentration sensor is developed by coating the electret sensor with a single-walled nanotube film whose stiffness changes in the presence of carbon dioxide. Experimental results prove the viability of the sensing technique.

Evaluation of Occupational Exposure to Antiblastic Drugs in an Italian Hospital Oncological Department

The determination of the current antiblastic drug contamination levels in an Italian hospital oncology ward was carried out. Statistical evaluation of data aiming to identify potential exposure causes was performed. Cyclophosphamide (CP), ifosfamide (IF) and 5-fluorouracil (5-FU) were determined by wipe tests, extracted with diatomaceous earths and quantified by GC/MSMS or HPLC/UV. Data were analysed with respect to the potential contamination levels of sampled surfaces, and various amounts of handled analyte. chi(2) tests and Spearman correlation coefficients were calculated. Median concentration levels of 0.086, 0.071, 2.363 microg/dm(2) were obtained, (CP, IF, 5-FU, respectively). 3.8 and 13.5% of investigated surfaces showed CP and IF concentrations higher than 1 microg/dm(2) (up to 26.96 microg/dm(2)) and 13.4% of samples contained 5-FU concentrations in the range 20-208.9 microg/dm(2). Analytes' concentration levels were dependent on sampling sites, with significant correlations showing a progressive contamination decrement going from workbenches, floor, hood planes and other surfaces. A diffuse contamination (traces of all the three analytes) was found on all investigated surfaces, even when analytes had not been used during the sampling days. A significant correlation (rho s=0.303, p=0.001) between the measured analyte concentration and the analyte handled amount was found only in the case of IF. The risk management strategy should be improved, as suggested by the measured and widespread levels of contamination. Since contamination also depends on other factors attributable to working modalities and cleaning procedures, the obtained results suggest that performance of specific training courses as well as scheduling environmental monitoring plans to achieve an actual decrement of the observed contamination levels should be implemented.

Quantitative, Multiplexed Assays for Low Abundance Proteins in Plasma by Targeted Mass Spectrometry and Stable Isotope Dilution

Biomarker discovery produces lists of candidate markers whose presence and level must be subsequently verified in serum or plasma. Verification represents a paradigm shift from unbiased discovery approaches to targeted, hypothesis-driven methods and relies upon specific, quantitative assays optimized for the selective detection of target proteins. Many protein biomarkers of clinical currency are present at or below the nanogram/milliliter range in plasma and have been inaccessible to date by MS-based methods. Using multiple reaction monitoring coupled with stable isotope dilution mass spectrometry, we describe here the development of quantitative, multiplexed assays for six proteins in plasma that achieve limits of quantitation in the 1-10 ng/ml range with percent coefficients of variation from 3 to 15% without immunoaffinity enrichment of either proteins or peptides. Sample processing methods with sufficient throughput, recovery, and reproducibility to enable robust detection and quantitation of candidate biomarker proteins were developed and optimized by addition of exogenous proteins to immunoaffinity depleted plasma from a healthy donor. Quantitative multiple reaction monitoring assays were designed and optimized for signature peptides derived from the test proteins. Based upon calibration curves using known concentrations of spiked protein in plasma, we determined that each target protein had at least one signature peptide with a limit of quantitation in the 1-10 ng/ml range and linearity typically over 2 orders of magnitude in the measurement range of interest. Limits of detection were frequently in the high picogram/milliliter range. These levels of assay performance represent up to a 1000-fold improvement compared with direct analysis of proteins in plasma by MS and were achieved by simple, robust sample processing involving abundant protein depletion and minimal fractionation by strong cation exchange chromatography at the peptide level prior to LC-multiple reaction monitoring/MS. The methods presented here provide a solid basis for developing quantitative MS-based assays of low level proteins in blood.

Paramagnetic particle detection for use with an immunoassay based biosensor

A biosensor and detection system that can provide measurement of analyte concentration within a test sample in four minutes is presented. The biosensor uses an immunoassay with micrometre-sized paramagnetic particle (PMP) labels for quantification. An immunoassay relies on the specific interaction between an antigen (analyte) and the corresponding antibody to immobilise the antigen to a substrate for measurement. In this system, the test samples and PMP labels are added to a reaction vessel on the biosensor and are attracted to a reaction surface at the base of the vessel using an electromagnet, where specific binding takes place. Unlike other immunoassay systems no extra washing or processing procedures are required. The bound labels, and associated antigen are detected, in situ, using a detector coil on the surface of the biosensor. The incorporation of a second coil under the biosensor allows for compensation for environmental changes such as thermal drift. The measurement of the resonant frequency of the detector coil in a parallel resonant circuit, prior to and after addition of the sample, provides a measure of the antigen in the sample. Good frequency stability is provided by the use of a phase locked loop arrangement. The resulting sensitivity was 1.17 µg of PMP labels per 1 Hz of frequency change and the minimum mass of PMPs detected was 62.5 µg. To demonstrate the utility of the technology and the rapidity of measurement, a model immunoassay for Troponin I (a cardiac marker) was used and discrimination at the clinically relevant value of 0.5 ng/ml Troponin I was achieved.

Microfluidic immunoassays as rapid saliva-based clinical diagnostics

At present, point-of-care (POC) diagnostics typically provide a binary indication of health status (e.g., home pregnancy test strip). Before anticipatory use of diagnostics for assessment of complex diseases becomes widespread, development of sophisticated bioassays capable of quantitatively measuring disease biomarkers is necessary. Successful translation of new bioassays into clinical settings demands the ability to monitor both the onset and progression of disease. Here we report on a clinical POC diagnostic that enables rapid quantitation of an oral disease biomarker in human saliva by using a monolithic disposable cartridge designed to operate in a compact analytical instrument. Our microfluidic method facilitates hands-free saliva analysis by integrating sample pretreatment (filtering, enrichment, mixing) with electrophoretic immunoassays to quickly measure analyte concentrations in minimally pretreated saliva samples. Using 20 microl of saliva, we demonstrate rapid (<10 min) measurement of the collagen-cleaving enzyme matrix metalloproteinase-8 (MMP-8) in saliva from healthy and periodontally diseased subjects. In addition to physiologically measurable indicators of periodontal disease, conventional measurements of salivary MMP-8 were used to validate the microfluidic assays described in this proof-of-principle study. The microchip-based POC diagnostic demonstrated is applicable to rapid, reliable measurement of proteinaceous disease biomarkers in biological fluids.

Integrated Microfluidic Platform for Oral Diagnostics

While many point-of-care (POC) diagnostic methods have been developed for blood-borne analytes, development of saliva-based POC diagnostics is in its infancy. We have developed a portable microfluidic device for detection of potential biomarkers of periodontal disease in saliva. The device performs rapid microfluidic chip-based immunoassays (<3-10 min) with low sample volume requirements (10 microL) and appreciable sensitivity (nM-pM). Our microfluidic method facilitates hands-free saliva analysis by integrating sample pretreatment (filtering, enrichment, mixing) with electrophoretic immunoassays to quickly measure analyte concentrations in minimally pretreated saliva samples. The microfluidic chip has been integrated with miniaturized electronics, optical elements, such as diode lasers, fluid-handling components, and data acquisition software to develop a portable, self-contained device. The device and methods are being tested by detecting potential biomarkers in saliva samples from patients diagnosed with periodontal disease. Our microchip-based analysis can readily be extended to detection of biomarkers of other diseases, both oral and systemic, in saliva and other oral fluids.

Empirical versus modelling approaches to the estimation of measurement uncertainty caused by primary sampling

Measurement uncertainty is a vital issue within analytical science. There are strong arguments that primary sampling should be considered the first and perhaps the most influential step in the measurement process. Increasingly, analytical laboratories are required to report measurement results to clients together with estimates of the uncertainty. Furthermore, these estimates can be used when pursuing regulation enforcement to decide whether a measured analyte concentration is above a threshold value. With its recognised importance in analytical measurement, the question arises of 'what is the most appropriate method to estimate the measurement uncertainty?'. Two broad methods for uncertainty estimation are identified, the modelling method and the empirical method. In modelling, the estimation of uncertainty involves the identification, quantification and summation (as variances) of each potential source of uncertainty. This approach has been applied to purely analytical systems, but becomes increasingly problematic in identifying all of such sources when it is applied to primary sampling. Applications of this methodology to sampling often utilise long-established theoretical models of sampling and adopt the assumption that a 'correct' sampling protocol will ensure a representative sample. The empirical approach to uncertainty estimation involves replicated measurements from either inter-organisational trials and/or internal method validation and quality control. A more simple method involves duplicating sampling and analysis, by one organisation, for a small proportion of the total number of samples. This has proven to be a suitable alternative to these often expensive and time-consuming trials, in routine surveillance and one-off surveys, especially where heterogeneity is the main source of uncertainty. A case study of aflatoxins in pistachio nuts is used to broadly demonstrate the strengths and weakness of the two methods of uncertainty estimation. The estimate of sampling uncertainty made using the modelling approach (136%, at 68% confidence) is six times larger than that found using the empirical approach (22.5%). The difficulty in establishing reliable estimates for the input variable for the modelling approach is thought to be the main cause of the discrepancy. The empirical approach to uncertainty estimation, with the automatic inclusion of sampling within the uncertainty statement, is recognised as generally the most practical procedure, providing the more reliable estimates. The modelling approach is also shown to have a useful role, especially in choosing strategies to change the sampling uncertainty, when required.

Application of a static solid-phase microextraction procedure combined with liquid–liquid extraction to determine poly(dimethyl)siloxane–water partition coefficients for selected polychlorinated biphenyls

A static solid-phase microextraction (SPME) procedure combined with liquid–liquid extraction (LLE) was used to determine the poly(dimethyl)siloxane (PDMS)–water partition coefficients ( K f) for selected polychlorinated biphenyl congeners (PCBs), including PCB 1, 15, 28, 47, 101, 153, 180, 202, 206, and 209. The accuracy for the measurements of analyte concentrations in the aqueous phase was ensured with a one-to-one recovery correction strategy employing one 13C-labeled PCB congener as a surrogate standard for each unlabeled PCB counterpart. The effects of coating thickness (7, 30, and 100 μm) and sample volume (130 mL and 2 L) on the K f values were examined experimentally and confirmed with paired t-tests. Significant dependence of K f values on coating thickness was found for a few heavily chlorinated congeners only, and was tentatively attributed to the use of the inaccurate effective coating volumes and the structural variation with these PDMS coatings. In addition, no significant differences in the log K f values of all analytes except for PCB 206 were found between the sample sizes of 130 mL and 2 L for both the 7- and 100-μm coatings. Overall, K f values obtained with 2-L sample containers were consistently higher than those reported in the literature, which is attributable to the selection of appropriate equilibrium times for SPME and direct measurements of aqueous analyte concentrations with LLE in the present study.

A Spatial-domain Fourier Transform Infrared Spectrometer: Application for Analyte Measurement in Cell Culture Media

A spatial-domain Fourier Transform (FT) infrared (IR) spectrometer coupled with a PtSi Schottky­barrier IR detector plane was developed in the spectral range of <TEX>$2.0-2.5{\mu}m$</TEX> for noninvasive measurement of analyte concentrations in cell culture media during cell culture processing. A key optical component of the spectrometer is a Savart plate which is a birefringent polarizer generating coherent two rays for interfering. The spectral resolution of the spectrometer was determined as <TEX>$71cm^{-1}$</TEX> (<TEX>${\~}0.05{\mu}m$</TEX> at <TEX>$2.5{\mu}m$</TEX>). Clear IR fringe patterns were imaged on the IR detector plane. The feasibility of the spectrometer for our application was investigated by measuring absorbance spectra of glucose and fetal bovine serum (FBS) which are important compounds in cell culture media. Experiment results show that the spectral quality of glucose and FBS was comparable with the standard spectra acquired with a commercial FT-IR spectrometer, presenting the feasibility of the spectrometer to perform analyte measurement in cell culture media.

Spatial contaminant heterogeneity: quantification with scale of measurement at contrasting sites

Material within the terrestrial environment is rarely homogeneously distributed, either spatially or temporally. One consequence of heterogeneity is that uncertainty is usually generated in measurements that are taken with the aim of characterising the environment. For example, a measurement of analyte concentration within soil taken from one sampling location on contaminated land can vary substantially when compared against another sample taken at effectively the same nominal location. The measurement uncertainty arising from the heterogeneity can substantially limit the reliability of the interpretations made upon environmental investigations. The sampling uncertainty usually outweighs the analytical uncertainty from the laboratory, often by a factor of 20 or more. One approach to reducing the uncertainty is to design a more suitable sampling strategy. This might be achieved by predicting the degree of heterogeneity prior to the investigation, but this is often difficult to achieve accurately. Another approach, which was investigated here, is to actually characterise the heterogeneity prior to the main investigation using rapid and inexpensive technology, such as in situ measurement techniques. In situ portable X-ray fluorescence (PXRF) and X-ray microprobe (XMP) techniques were employed to test the feasibility of this approach. Two contrasting contaminated land sites were chosen to characterise the two-dimensional spatial heterogeneity of heavy metal contamination in topsoil at a range of scales (50 m to 0.001 m). The spatial heterogeneity of contaminants, expressed as relative standard deviations, was found to differ between the two sites by a factor of two, largely due to the mode of deposition of pollution. The study also indicated that the heterogeneity did not change systematically with the scale of measurement between sampling locations at either site.

Chemical Applications of a Programmable Image Acquisition System

Image analysis is widely used in chemistry, both for rapid qualitative evaluations using techniques such as thin layer chromatography (TLC) and for quantitative purposes such as well-plate measurements of analyte concentrations or fragment-size determinations in gel electrophoresis. This paper describes a programmable system for image acquisition and processing that is currently used in the laboratories of our organic and physical chemistry courses. It has also been used in student research projects in analytical chemistry and biochemistry. The potential range of applications is illustrated by brief presentations of four examples: (1) using well-plate optical transmission data to construct a standard concentration–absorbance curve; (2) the quantitative analysis of acetaminophen in Tylenol and acetylsalicylic acid in aspirin using TLC with fluorescence detection; (3) the analysis of electrophoresis gels to determine DNA fragment sizes and amounts; and, (4) using color change to follow reaction kinetics. Th...

Signal Amplification in Time-resolved Fluorometry

A frequent question among clinical biochemists is how sensitive analytical techniques should be in the context of clinical diagnostics. Currently, the analyte concentrations usually encountered in clinical chemistry range from 10−3 to 10−12 mol/L. By far, the most sensitive nonamplification techniques used in the clinical laboratory are based on noncompetitive immunological assays. Is there any need for measuring analytes at even lower concentrations? The answer is likely yes. Once the methodologies for measuring even lower concentrations of analytes are developed and our knowledge of the many new candidate biological markers that likely will be discovered through the Human Genome Project is more complete, we may be interested in or need to measure analyte concentrations that are 1/10th to 1/100th of those currently measured. Hence, we should continually pursue the development of methodologies that can reach the ultimate sensitivity, i.e., detection of single molecules. In other areas of laboratory medicine, e.g., microbiology, single pathogen particles (e.g., viruses and bacteria) have diagnostic significance. We should not forget that the measurement of a single molecule in a very small fraction of the total blood volume may mean that the whole organism could contain relatively large numbers of such pathogenic or abnormal constituents. When the analytes are nucleic acids (DNA …