Clinical detection limits

Zwelg, M. "Establishing Clinical Detection Limits of Laboratory Tests" - Chapt. 8 in this volume. 

Establishing Clinical Detection Limits of Laboratory Tests... 

ImmunO lSS iy. Chemiluminescence compounds (eg, acridinium esters and sulfonamides, isoluminol), luciferases (eg, firefly, marine bacterial, Benilla and Varela luciferase), photoproteins (eg, aequorin, Benilld), and components of bioluminescence reactions have been tested as replacements for radioactive labels in both competitive and sandwich-type immunoassays. Acridinium ester labels are used extensively in routine clinical immunoassay analysis designed to detect a wide range of hormones, cancer markers, specific antibodies, specific proteins, and therapeutic dmgs. An acridinium ester label produces a flash of light when it reacts with an alkaline solution of hydrogen peroxide. The detection limit for the label is 0.5 amol. 

It should be noted that the term sensitivity sometimes may alternatively be used, namely in analytical chemistry and other disciplines. Frequently the term sensitivity is associated with detection limit or detection capability. This and other misuses are not recommended by IUPAC (Orange Book [1997, 2000]). In clinical chemistry and medicine another matter is denoted by sensitivity , namely the ability of a method to detect truly positive samples as positive (O Rangers and Condon [2000], cited according to Trullols et al. [2004]). However, this seems to be more a problem of trueness than of sensitivity. 

This field is therefore at an exciting stage. Ion-selective electrodes have a proven track record in terms of clinical and biomedical analysis, with a well-developed theory and a solid history of fundamental research and practical applications. With novel directions in achieving extremely low detection limits and instrumental control of the ion extraction process this field has the opportunity to give rise to many new bioana-lytical measurement tools that may be truly useful in practical chemical analysis. 

After oral administration of 400 mg of rifaximin to fasted healthy volunteers blood drug concentration was found to be lower than the detection limit of the analytical method (i.e. 2.5 ng/ml) in half of them [102]. In the remaining subjects very low amounts were detected at some of the time intervals during the first 4 h after intake. Along the same lines, the urinary concentrations of the drug were very low and often undetectable. The effect of food on the absorption of the antibiotic was also evaluated [34] and a significant, albeit not clinically relevant, increase of bioavailabity was observed after a high-fat breakfast (table 5). 

Flame emission spectrometry is used extensively for the determination of trace metals in solution and in particular the alkali and alkaline earth metals. The most notable applications are the determinations of Na, K, Ca and Mg in body fluids and other biological samples for clinical diagnosis. Simple filter instruments generally provide adequate resolution for this type of analysis. The same elements, together with B, Fe, Cu and Mn, are important constituents of soils and fertilizers and the technique is therefore also useful for the analysis of agricultural materials. Although many other trace metals can be determined in a variety of matrices, there has been a preference for the use of atomic absorption spectrometry because variations in flame temperature are much less critical and spectral interference is negligible. Detection limits for flame emission techniques are comparable to those for atomic absorption, i.e. from < 0.01 to 10 ppm (Table 8.6). Flame emission spectrometry complements atomic absorption spectrometry because it operates most effectively for elements which are easily ionized, whilst atomic absorption methods demand a minimum of ionization (Table 8.7). 

For applications in the diagnostics and biomarker area, 8-oxo-7,8-dihydro-2,-deoxyguanosine (8-oxoGuo) was measured as an oxidation stress biomarker in urine samples from smokers and non-smokers (Hu et al. 2006). When 100, uL of samples (10 times dilution) were used, a detection limit of 5.7 pg/mL (2.0 fmol) was achieved. The cycle time was 10 min per sample. The application was used for clinical scale. A similar approach was used for the detection of N7-methylguanine, another carcinogen exposure biomarker in human urine (Chao et al. 2005). 

In this case, sodium emission is monitored at a wavelength of 589.6 nm and potassium at a wavelength of 769.9 nm. The intensity of emission is calibrated with appropriate standards for the samples to be analyzed. In this way it is possible to automatically determine 100 values of sodium and potassium for 100 samples/h using modern clinical instruments. Limits of detection are sub-ppm and for serum values 140 mg/m the range of reproducibility is on the order of 2-3%. 

Although these assay procedures for steroids were successfully applied to clinical samples, the sensitivity was not extensively improved, as shown by their femtomole range detection limits 2 fmoF/assay for E2, 0.32 nmol/liter (40 fmoF/assay) for progesterone, and 0.4 nmol/liter (8 fmol /assay) for Ei-3G, respectively. Difficulty in obtaining a higher sensitivity would come from the competitive substitution of the bound hapten with the /3 -type antibody and/or imperfect selectivity of the a-type antibody toward the hapten-antibody complex. 

Further progress of ECL probes immobilization methods should result in new robust, stable, reproducible ECL sensors. Especially, the use of electrochemilumi-nescent polymers may prove to be useful in this respect. There are also good prospects for ECL to be used as detection in miniaturized analytical systems particularly with a large increase in the applications of ECL immunoassay because high sensitivity, low detection limit, and good selectivity. One can believe that miniaturized biosensors based on ECL technology will induce a revolution in clinical analysis because of short analysis time, low consumption of reactants, and ease of automation. 

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