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Objective: Recent studies have shown that calculations of the percent free/total prostate-specific antigen (PSA) improves the specificity of PSA testing. Characterizing the variability of free PSA and total PSA is necessary to evaluate the utility of an isolated free/total PSA measurement. We investigated the total variation of free and total PSA levels to determine how the percent free/total PSA was affected., Design: Serum was obtained from nine urological patients on 5 different days over a 2-week period. Free and total PSA levels were measured on the day of collection. The total variation expressed in terms of percent coefficient of variation (%CV) was calculated, and the biological variation was derived taking analytical variation into consideration., Setting: Patients were from Seattle (Wash) Urological Associates, and samples were processed at the Dynacare Laboratory of Pathology, Seattle, Wash., Patients: Nine men (aged 48 to 69 years) were evaluated; three had been diagnosed with prostate cancer, three with benign prostatic hyperplasia, one with chronic prostatitis, one with high-grade prostatic intraepithelial neoplasia, and one was clinically normal., Main Outcome Measures: Total variation for free, total, and percent free/total PSA., Results: The average total variation was 13.9% CV, 7.5% CV, and 10.6% CV for free, total, and percent free/ total PSA, respectively. Biological variation was derived to be 13.0% CV, 5.6% CV, and 8.0% CV for free, total, and percent free/total PSA, respectively., Conclusions: When applied, these results suggest that there are significant random changes in the numerator and denominator of the free PSA-total PSA ratio that could result in clinical misinterpretation. Clinicians must be aware that free PSA and total PSA levels will fluctuate owing to nonpathologic variation.

We modified the Hybritech Tandem-E prostate-specific antigen (PSA) assay by increasing the sample volume, increasing enzyme-substrate incubation time, and using diethanolamine buffer. Our modified method has a detection limit of 0.009 microgram/L (P < 0.01). The assay curve is linear from 0.01 to 1.0 micrograms/L and has an overall assay time of about 4 h. Linear plots are obtained when the 1.0 micrograms/L standard is diluted with either matrix buffer or serum from men containing PSA < 0.01 microgram/L. Recovery of PSA (0.10 microgram/L) added to serum from men averaged 94%. Interassay CVs were 13%, 7%, and 4% at PSA concentrations of 0.04, 0.07, and 0.30 micrograms/L, respectively (n = 33). This assay should be useful in the detection of early recurrence of prostate cancer after radical prostatectomy.

The objective of this study was to examine quantitative differences between the two commonly used methods for determining serum albumin concentration, bromcresol green (BCG) and bromcresol purple (BCP), in normal subjects and in 235 unselected dialysis patients in view of recently established Health Care Financing Administration (HCFA) quality assurance review criteria. The mean of normal results by the BCG method was 4.4 g/dL, and 97.5% of values were 3.8 g/dL or higher. The mean of normal results by the BCP method was 3.9 g/dL, and 97.5% of values were 3.3 g/dL or higher. Serum albumin concentrations in samples from the dialysis patients had respectively lower mean values by both methods. For the BCG method, the mean was 3.8 g/dL, and 82% of values were 3.5 g/dL or higher; for the BCP method, the mean was 3.3 g/dL, and 82% of values were 3.0 g/dL or higher. Likewise, for the reference immunonephelometric procedure, the mean value for the dialysis patients was 3.3 g/dL, and 82% of values were 3.0 g/dL or higher. For the samples from the dialysis patients, in comparison with the immunonephelometric method, the BCG method exhibited both constant (intercept = 9.3 g/L) and proportional error (slope = 0.87). The mean albumin value for the BCG method was 3.8 g/dL, 15% higher. In contrast, the BCP method compared closely with the reference method: slope = 1.00, intercept = 0.8 g/L, mean x = 3.3 g/dL, mean y = 3.3 g/dL. The HCFA quality assurance criteria are valid only for the BCG method.(ABSTRACT TRUNCATED AT 250 WORDS)

The enzymatic cholesterol method used with the Du Pont aca has been modified to provide a reliable measurement of high-density lipoprotein cholesterol in serum after heparin/Mn2+ precipitation of the low- and very-low-density lipoproteins. Interference by Mn2+, equivalent to about 90 mg of cholesterol per liter, is decreased to less than 40 mg of cholesterol per liter by the presence of ethylenediaminetetraacetate (8 mmol/liter) in the diluent; the residual effect of Mn2+ is compensated by calibrating the aca with standards containing Mn2+ and heparin. With an 80-microliter sample, the sensitivity is 236 muA/mg per liter and linearity ranges from 50 to 1500 mg/liter. Average analytical recovery of cholesterol added to the high-density lipoprotein fraction was 103%. Diluted fractions give the expected results. Between-run reproducibility (CV) is 1.3 and 1.6% at 463 and 554 mg/liter. Correlation with the Lipid Research Clinics' procedure (25 samples) gave a regression line of y(aca) = 1.039x- 15, and a correlation coefficient of 0.997.

We describe the performance of a commercial (Steranti/EIR) RIA reagent kit for measuring 17 beta-estradiol directly in serum. Day-to-day precision data for control sera were as follows: mean = 102.8 ng/L, CV = 6.8%, n = 20; mean = 231.1 ng/L, CV = 5.3%, n = 21; mean = 747.7 ng/L, CV = 9.4%, n = 21. Analytical recovery of added estradiol from seven different serum pools from men, to which three different concentrations of estradiol had been added, was (mean +/- SD): 98.6 +/- 7.0% at 107.5 ng/L added; 98.8 +/- 4.7% at 322.5 ng/L added; 108.2 +/- 4.8% at 645 ng/L added. Overall recovery of estradiol in these experiments (mean +/- SD for 21 determinations) averaged 101.9 +/- 7.0%. Assay of 32 serum specimens from women by both the direct (y) and an extraction method (x) gave the following linear regression statistics: y = 1.12x - 1.3, r = 0.998, Sy/x = 30.2 ng/L, mean y = 438.2 ng/L, mean x = 391.4 ng/L. Hemoglobin, bilirubin, and moderate lipemia do not interfere. Sensitivity of the direct assay was 2.6 ng/L. Compared with the extraction assay, the direct estradiol assay has advantages of speed and simplicity.

We describe a completely automated enzymic system for measuring total cholesterol in serum. All reagents are contained in an analytical test pack and the test is performed on Du Pont's Automatic Clinical Analyzer (aca), which mixes the sample (20 microliter) and reagents and performs the necessary absorbance measurements and calculations. In the procedure, cholesterol oxidase oxidizes free cholesterol. The oxidation step produces cholest-4-en-3-one and hydrochloride peroxidesterase hydrolyzes cholesterol esters and cholesterol in direct proportion to the amount of cholesterol present. N,N-Diethylaniline hydrochloride and 4-aminoantipyrine react with the hydrogen peroxide to produce a quinoneimine dye (lambda max = 553 nm). Interacting reagents have been optimized simultaneously (coöptimization) utilizing response surface designs coupled with computer analysis of the data. Reagent efficiency is high and analytical performance reliable.

We describe the performance of an RIA for the measurement of prostate-specific antigen (PA). Between-assay precision (CV) for control sera with various analyte concentrations was as follows: mean = 1.67 micrograms/L, CV = 7.1%; mean = 4.47 micrograms/L, CV = 5.6%; mean = 7.15 micrograms/L, CV = 5.5% (n = 19 each). Analytical recovery of PA (nine concentrations ranging from 2.3 to 21.1 micrograms/L) added to a serum pool averaged 101.8% (range 96.1 to 116.1%). Sensitivity (detection limit) of the RIA was 0.25 micrograms/L. Cross reactivity of prostatic acid phosphatase (PAP) in this assay was less than 0.022%. The mean percent B/B0 for 74 specimens from women was 98.9%, not statistically different from that for the zero standard. The normal reference interval for men was 0-2.7 micrograms/L ( 99th percentile), as established by assay of specimens from 276 apparently normal men. Measurement of PA and prostatic acid phosphatase in 205 consecutive serum specimens from patients with clinical evidence of prostate disease similarly placed patients into normal (98) or abnormal groups (54) in 152 cases. However, in 49 cases only the concentration of PA in serum was abnormal. Sequential measurement of both tissue markers in specimens from several patients who were undergoing therapy for prostate cancer appeared to provide supplemental information regarding treatment success.

We describe an automated method for magnesium assay, for use with the Cobas-Bio centrifugal analyzer. Magnesium is reacted with calmagite, a dye, and the absorbance of the magnesium-dye complex at 520 nm is measured. EDTA is then added to break up the complex and the absorbance at 520 nm is re-measured to correct for sample color and turbidity. Including ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) and triethanolamine in the dye reagent eliminates interference by calcium and iron. Within-run CVs were less than 2.0% for concentrations of magnesium ranging from 18 to 40 mg/L, and less than 4% for a magnesium concentration of 9.4 mg/L. Day-to-day precision data, determined over five months were: mean = 21.0 mg/L, CV = 2.7%; mean = 43.0 mg/L, CV = 3.2% (n = 550 for both). Comparison of the Cobas-Bio method (y) with an atomic absorption spectrometric method (x) gave the following results: y = 0.968x + 0.448, r = 0.986, Sy/x = 0.34 mg/L, mean x = 19.8 mg/L, mean y = 19.6 mg/L, n = 44. Hemoglobin, bilirubin, and turbidity do not interfere. The standard curve is linear up to a magnesium concentration of 97 mg/L.

An 125I-radioimmunoassay is described which measures unconjugated estriol in the serum of pregnant women. Estriol is extracted into ethyl acetate/hexane, an aliquot is evaporated, and the residue is redissolved in phosphate buffer. The sample is incubated with tracer and antibody at 37 degrees C for 15 min and then at 4 degrees C for 1 h and 45 min. The antibody-bound fraction is then precipitated with polyethylene glycol and isolated by centrifugation. Because the antigen-antibody complex is stable in the presence of polyethylene glycol, the separation steps are not influenced by timing. Extraction recovery of 3H-labeled estriol added to a pool of sera from pregnant women averaged 96.7% (SD = 1.3, n = 10). Estriol-supplemented (5, 10, and 20 microgram/liter) serum from men, carried through the entire procedure, showed analytical recovery ranging from 94 to 106%. Structurally analogous steroids normally present in serum of pregnant women exhibit negligible cross reactivity. Day-to-day precision (CV) is 13.3% (3.1 microgram/liter), 6.4% (7.6 microgram/liter), and 5.6% (21.4 microgram/liter) for n = 21. The current reagent cost (about 17 cents per tube), and a total procedural time, including counting, of 5.5 h make this an acceptable assay for routine use.

We describe an automated method for calcium assay, for use with the Cobas-Bio centrifugal analyzer. Calcium is reacted with cresolphthalein complexone and the absorbance of the calcium--dye complex at 575 nm is measured. EDTA is then added to break up the calcium--dye complex and the absorbance at 575 nm is re-measured, to correct for endogenous color and turbidity. Day-to-day precision data, determined over four months, were as follows: mean = 92.9 mg/L, CV = 1.47%; n = 216; mean = 128.7 mg/L, CV = 1.72%; n = 216. Comparison of the Cobas-Bio method (y) with an atomic absorption spectrometric method (x) gave the following results: y = 1.012x--2.05, r = 0.991, Sy/x = 1.2, mean x = 92.63 mg/L, mean y = 91.69 mg/L, n = 74. Hemoglobin, bilirubin, or turbidity does not interfere. At the medical decision value (110 mg/L), the overall analytical error is 4.6 mg/L, which is less than the 5 mg/L allowable (95% confidence limit) error.

Prostatic acid phosphatase values in 98 patients with prostatic carcinoma were measured by a commercial radioimmunoassay (RIA) and by enzymatic assay. Forty-three carcinomas were staged by rigorous pathological criteria. Patients (N = 129) with benign prostatic hyperplasia were the control group. At 94% specificity, sensitivities of the RIA vs the enzymatic assay for clinically staged patients were as follows: stage A, 22% vs 6%; B, 29% vs 10%; C, 52% vs 38%; and D, 87% vs 80%. However, none of the seven patients with pathological stage A and B disease had a positive test result, and we suggest that variability in staging criteria accounts for the discrepant sensitivity claims reported. Prostatic acid phosphatase RIA should not be used for screening but as an adjunct for staging known prostatic carcinoma.