Evaluation of urinary oestrogen assays after the menopause and their potential for screening

Urinary oestrone-3-glucuronide, oestradiol-3-glucuronide, oestrone and oestradiol were measured by radioimmunoassay methods adapted for the very low levels found in postmenopausal women. Oestrogen concentrations related to creatinine in morning urine samples from ten postmenopausal women were found to correlate well with total oestrogen excreted in 24 h (r = 0.934, 0.867, 0.947, 0.909, respectively; p less than 0.002). Day to day variation in five individuals, measured over 5 weeks, showed random fluctuations, with no obvious cyclical variation. Reference ranges, based on two consecutive morning urine samples from 131 postmenopausal women, were 0.73-4.57 mumol/mol creatinine for oestrone-3-glucuronide, 0.66-3.00 mumol/mol creatinine for oestradiol-3-glucuronide, 4.8-30.9 nmol/mol creatinine for oestrone and 3.8-16.8 nmol/mol creatinine for oestradiol. We suggest that such assays may have a part to play in the screening for women at greatest risk of developing osteoporotic fractures.     

Comparison of immunoturbidimetric and immunonephelometric assays for specific proteins

Immunoturbidimetric assays for specific proteins are available on “open system” clinical chemistry analyzers. The analytical performance of nine immunoturbidimetric specific protein assays (C3, C4, CRP, Haptoglobin, IgA, IgG, IgM, RF, and Transferrin) was compared to immunonephelometry. Testing was performed on the Abbott ARCHITECT ci8200 and the Dade Behring BNII nephelometer and evaluated for precision, linearity, limit of detection, prozone phenomenon, method comparison, workflow, and proficiency testing survey comparison. Immunoturbidimetric assays performance was satisfactory for total precision, linearity, limit of detection and the prozone effect was not observed. Method comparison was acceptable for the immunoglobulins, CRP and transferrin but less favorable for the other assays, likely due to methodology and antibody specificity differences. Immunourbidimetric specific protein assays allow for efficient test consolidation on a general purpose clinical chemistry analyzer.     

Methodological evaluation and comparison of five urinary albumin measurements

Background: Microalbuminuria is an indicator of kidney damage and a risk factor for the progression kidney disease, cardiovascular disease, and so on. Therefore, accurate and precise measurement of urinary albumin is critical. However, there are no reference measurement procedures and reference materials for urinary albumin. Methods: Nephelometry, turbidimetry, colloidal gold method, radioimmunoassay, and chemiluminescence immunoassay were performed for methodological evaluation, based on imprecision test, recovery rate, linearity, haemoglobin interference rate, and verified reference interval. Then we tested 40 urine samples from diabetic patients by each method, and compared the result between assays. Results: The results indicate that nephelometry is the method with best analytical performance among the five methods, with an average intraassay coefficient of variation (CV) of 2.6%, an average interassay CV of 1.7%, a mean recovery of 99.6%, a linearity of R=1.00 from 2 to 250 mg/l, and an interference rate of <10% at haemoglobin concentrations of <1.82 g/l. The correlation (r) between assays was from 0.701 to 0.982, and the Bland–Altman plots indicated each assay provided significantly different results from each other. Conclusion: Nephelometry is the clinical urinary albumin method with best analytical performance in our study. J. Clin. Lab. Anal. 25:324–329, 2011. © 2011 Wiley‐Liss, Inc.     

Reactivity of urinary albumin (microalbumin) assays with fragmented or modified albumin

BACKGROUND: Controversy exists regarding occurrence and measurement of structural variants of albumin in urine. In this study, we examined cross-reactivity of in vitro modified albumins in assays for urine albumin (microalbumin). METHODS: We analyzed albumin modified by reagents, trypsin, or physical treatments or differing in primary sequence (animal albumins) with an immunoturbidimetric assay (Beckman LX20) using goat antiserum and a competitive immunoassay (Siemens Immulite) using a monoclonal antibody. We assessed occurrence of albumin fragments in urine by use of Western blotting of 24 specimens. RESULTS: Chemical modification, modest sequence substitution (gorilla albumin), or cleavage of albumin by cyanogen bromide (CNBr) had little effect on reactivity in the LX20 assay. Albumin extensively cleaved with trypsin retained partial reactivity. The Immulite assay generally was affected more severely by albumin modifications and sequence changes. Western blots of fresh urine specimens or specimens stored at -80 degrees C showed little albumin fragmentation, but some specimens stored for 3 years at -20 degrees C had extensively fragmented albumin that was detected by the LX20 but not the Immulite assay. CONCLUSIONS: Nearly equivalent reactivity of intact albumin and CNBr fragments in the immunoturbidimetric assay indicates reactivity of antibodies with multiple epitopes throughout albumin. Therefore, it is difficult to abolish reactivity of albumin in this type of urine albumin assay. Differential sensitivity of 2 assays to albumin modification identifies a potential source of assay nonequivalence in measuring urinary albumin, particularly for specimens stored at -20 degrees C.     

Evaluation of a new radioimmunoassay for urinary albumin

We have evaluated a new commercially available radioimmunoassay kit for albumin determination in urine. The assay is precise; within-run precision (CV) in the clinically significant ranges is 1.8-3.5%, between-run, 1.2-8.5%. The minimum detection limit was 0.6 micrograms/ml. Analytic recovery of different concentrations of albumin added to urine ranged from 98% to 103%. Samples, stored in plastic containers, were stable at room temperature for periods up to 7 days. Mean albumin excretion rates, measured in 20 normal volunteers for 3-h and 24-h periods during the same day were similar (7.1 +/- 4.6 [SD] versus 6.5 +/- 5.0 micrograms/min). In 8 normal subjects, 3-h excretion rates measured daily for 5 days showed no significant variability. In eight insulin-dependent diabetic subjects, albumin excretion measured in short periods of urine collection (3 h) were also in close agreement with 24-h collections (24.7 +/- 28.9 versus 17.6 +/- 18 micrograms/min). From these results it appears that this commercially available kit is suitable for conveniently monitoring microalbuminuria in large numbers of patients in research studies as well as for office practice. Such widespread use should make it possible to better determine the clinical usefulness of this test in the management of diabetic patients.     

Evaluation of five high-sensitivity American thyrotropin assays

We compared five commercial immunometric kits for thyrotropin (TSH) manufactured in the United States--Abbott, Diagnostic Products, Hybritech, Nichols, and Becton Dickinson--and one sensitive "in-house" radioimmunoassay for their ability to monitor TSH levels in patients with hyperthyroidism and during thyroid hormone suppression therapy. With these five methods, we measured TSH concentrations is serum specimens from the following categories of patients: euthyroid (N = 99), hyperthyroid (N = 51), hypothyroid (N = 50), and thyroid cancer and nodular goiter on thyroid hormone suppression therapy (N = 33). The immunometric methods were similar in assay sensitivity and precision; the in-house radioimmunoassay was less sensitive than all other assays studied. The simultaneous correlation of TSH values obtained from all diagnostic categories was more than 90% among the assays studied. The immunometric assays clearly distinguished hyperthyroid from euthyroid patients and quantitated TSH levels in the subnormal range.     

免疫比浊法和放射免疫法测定尿微量白蛋白的比较

目的比较免疫比浊法和放射免疫法测定尿微量白蛋白的临床应用价值。方法分别采用免疫比浊法与放射免疫法测定50份新鲜尿标本中微量白蛋白浓度。结果免疫比浊法与放射免疫法测定结果无统计学意义，相关性良好（r=89．3％，P=0．062，P〉0．05）。结论免疫比浊法测定微量白蛋白浓度比放射免疫法结果更准确、及时，方法更简便、快速，无创伤性，更能满足临床需要。     

免疫比浊法和放射免疫法测定尿微量白蛋白的比较

目的比较免疫比浊法和放射免疫法测定尿微量白蛋白的临床应用价值。方法分别采用免疫比浊法与放射免疫法测定50份新鲜尿标本中微量白蛋白浓度。结果免疫比浊法与放射免疫法测定结果无统计学意义,相关性良好(r=89.3%,P=0.062,P>0.05)。结论免疫比浊法测定微量白蛋白浓度比放射免疫法结果更准确、及时,方法更简便、快速,无创伤性,更能满足临床需要。     

五种常用轮状病毒检测方法的评价及应用策略

目的比较胶体金免疫层析法(GICA)、酶联免疫吸附试验(ELISA)、反转录-聚合酶链反应(RT-PCR)、实时荧光定量-反转录聚合酶链反应(real-time RT-PCR)和聚丙烯酰胺凝胶电泳(polyacrylamide gelelectrophoresis,PAGE)5种常用的轮状病毒检测方法的准确性。方法取A组轮状病毒阳性标本,经1×101~1×10810倍系列稀释,采用上述5种方法平行检测,比较检测限;选择分别含有以下9种病原体且轮状病毒证实为阴性的粪便标本作为阴性对照:脊髓灰质炎病毒疫苗株Ⅰ~Ⅲ型、肠道病毒EV71、柯萨奇病毒A16、ECHO病毒6、诺如病毒Ⅰ型、诺如病毒Ⅱ型和星状病毒,采用上述5种方法平行检测,比较特异性;采用上述方法,分别对184例婴幼儿腹泻粪便标本进行检测,以PAGE检测结果作为参考标准,比较其余4种方法的灵敏度和特异性。结果上述5种方法检测A组轮状病毒的检测限相差5个数量级,分别为:real-time RT-PCR法10-6稀释度,胶体金法10-3稀释度,ELISA和PAGE方法均为10-2稀释度,RT-PCR法10-1稀释度;5种方法对9种对照病原体均无交叉反应;以PAGE方法检测结果作为标准,real-time RT-PCR、ELISA、胶体金和RT-PCR灵敏度分别为95.45%、96.97%、93.94%和62.12%,特异性分别为82.2%、86.44%、82.20%和94.07%。结论 PAGE作为检测轮状病毒的经典方法,具有高特异性,可用于确认实验;real-time RT-PCR方法检测A组轮状病毒的灵敏度优于其他4种方法,可用于轮状病毒的高通量检测;ELISA和胶体金方法具有良好的灵敏度与特异性,且胶体金方法成本低,可用于初筛检测;行业标准推荐的RT-PCR方法特异性较高,但灵敏度较低,有待改进。     

Development of immunoturbidimetric assays for fourteen human serum proteins on the Hitachi 912.

Many laboratories rely on dedicated nephelometers or turbidimeters and commercial reagent kits for the evaluation of serum proteins. However, with growing emphasis on cost containment, laboratories are forced to seek additional operational efficiencies by capitalizing on the use of existing analyzers whenever possible. In the present paper we describe the development of immunoturbidimetric assays for routine analysis of 14 human serum proteins (alpha1-antitrypsin, alpha2-macroglobulin, albumin, apolipoproteins Al and B, complement components 3 and 4, haptoglobin, immunoglobulins A, G, and M, orosomucoid, prealbumin, and transferrin) on the Hitachi 912, a general chemistry analyzer. With this system, we obtained excellent precision at levels corresponding to low, normal, and high physiologic concentrations of each protein (within-run imprecision CVs < or = 3.4%, total imprecision CVs < or = 4.1%). Linearity for each method was within 5% of the expected value throughout the calibration range, and method comparisons with either the Roche turbidimetric or Dade Behring nephelometric assays were in good agreement (r >0.97). We observed no significant interference from bilirubin (up to 718 micromol/l), hemoglobin (up to 8 g/l), triglyceride (up to 14.7 mmol/l) or rheumatoid factor (up to 4,140 IU/ml). Calibration for the 14 protein assays was stable for at least 7 days and onboard refrigerated reagents were stable for at least 3 months. The instrument's automated sample re-run feature minimized sample handling and helped to conserve specimens. In conclusion, the newly developed assays on the Hitachi 912 offer high throughput (>250 tests per hour) without the associated cost of a dedicated instrument for protein assays.     

人群调查中尿白蛋白检测方法的探讨

目的 探讨人群调查中尿白蛋白标本留取及检测方法.方法 选取659名北京市居民,收集其24 h尿测定UAER;并留取次日随机尿和晨尿,分别采用尿ACR及半定量尿白蛋白试纸法测定尿白蛋白.以24 h UAER作为标准,建立应用两种方法检测晨尿和随机尿白蛋白的ROC曲线,比较敏感度、特异度及ROC曲线下面积.结果 晨尿和随机尿ACR分别为9.36(5.12～33.29)mg/g及11.29(6.34～41.29)mg/g,组间比较差异无统计学意义(t=-1.382,P＞0.05),相关分析显示两者高度相关(r=0.932,P＜0.01).晨尿和随机尿ACR与24 h UAER具有高度相关性,相关系数分别为0.853及0.874,P均＜0.01.随机尿ACR诊断白蛋白尿的敏感度为77.9%,特异度为91.0%,晨尿ACR诊断白蛋白尿的敏感度为78.4%,特异度为95.7%.随机尿白蛋白试纸法诊断白蛋白尿的敏感度为90.3%,特异度为41.1%,特异度明显低于ACR法.随机尿与晨尿ACR的ROC曲线下面积分别为0.918±0.012及0.929±0.015,组间比较差异无统计学意义(χ2=2.13,P＞0.05).随机尿白蛋白试纸法ROC曲线下面积0.661±0.021,低于随机尿ACR(χ2=248.41,P＜0.01).结论 随机尿的ACR兼具简便及准确的特点,是人群调查中诊断白蛋白尿的良好指标.

Abstract：

Objective To evaluate the spot urine sample collection method and value of urinary albumin measurement in population survey. Methods Six hundred and fifty-nine Beijing residents were requested to collect 24 h urine for detection of UAER, as well as random spot urine samples and morning urine samples in the next day. Rapid semi-quantitative urinary albumin-specific dipstick and ACR were measured in each spot urine specimen. The 24 h UAER was measured as golden standard to generate ROC curves and evaluate the sensitivity, specificity and AUC of each method. Results The value of ACR in the morning spot urine samples and random spot urine samples were 9. 36(5. 12-33.29) mg/g and 11.29(6. 34-41.29) mg/g respectively and there was no significant difference between these two groups (t = - 1. 382,P＞0.05). The correlation was significant in the two groups (r = 0.932, P ＜ 0.01). The correlation coefficient between ACR in the morning spot urine samples and UAER was 0. 853 (P ＜ 0. 01). The correlation coefficient between ACR in the random spot urine samples and UAER was 0. 874 (P ＜0. 01).The sensitivity and specificity of ACR for diagnosis of albuminuria in the random urine samples were 77. 9% and 91.0%. The sensitivity and specificity of ACR for diagnosis of albuminuria in the morning urine samples were 78. 4% and 95.7%. Concerning the semi-quantitative urinary albumin-specific dipstick, sensitivity and specificity were 90. 3% and 41.1% , respectively. The specificity was much lower than that of ACR. The area under the ROC curves of ACR in the random urine specimens and the morning urine specimens was 0. 918 ±0. 012, 0. 929 ± 0. 015, respectively. There was no statistical difference between these two groups (χ2 =2. 13, P＞0. 05). The area under the ROC curves of semi-quantitative urinary albumin-specific dipstick in the random urine specimens was 0.661 ±0.021, lower than that of ACR (χ2 = 248.41, P＜0.01).Conclusion Measurement of ACR in random urine samples is a reasonable method with simplicity and accuracy for the detection of albuminuria in general population screening program.     

Assessment of new hyaluronic acid assays and their impact on FibroMeter scores

BackgroundWe compared three hyaluronic acid (HA) assays and analyzed the impact of their variations on FibroMeter scores.MethodsIn a test group of 165 patients, HA levels were assessed with the commonly used ELISA assay from Corgenix, a new ELISA assay from Teco and an immunoturbidimetry assay from Wako, this latter tested across three different instruments. Five different FibroMeter scores were calculated.ResultsCorrelation across the three assays (r_s between 0.969 and 0.995) was very good. Means of differences (d) were lower when the immunoturbidimetry assay was compared on different instruments: d between ?3.4 and 2.0 μg/L. However, a higher value for HA measurement was observed with Corgenix assay, compared to the other two assays (Teco and Wako): d between 27.1 and 36.4 μg/L. The assessment also demonstrated that HA variations had very little impact on FibroMeter scores: 0.0117 for virus and 0.0416 for alcoholic fibrosis scores, and between 0.58 and 1.71 for the area of fibrosis (expressed in percentage).ConclusionsThe two new assays found lower values of HA, as compared to the Corgenix assay. However, these differences had very little impact on FibroMeter scores and had no impact on clinical evaluation of liver fibrosis.     

A simple immunoturbidimetric method for IgG and albumin quantitation in cerebrospinal fluid and serum

We describe a simple immunoturbidimetric method for measuring both IgG and albumin in CSF and serum, which enables the calculation of CSF indices. For each protein, only one calibration curve is used for both CSF and serum samples. The assay protocol is simple and similar for both tests. Sensitivity and versatility of the method afford measurements over a very wide range of concentrations (approx. 0.007 to 94 g/l for IgG and 0.06 to 92.40 g/l for albumin). Precision studies (triplicates for 6 runs over 15 days) gave overall CVs: less than or equal to 2.9 and 4.9% for IgG in CSF (11.5 mg/l) and serum (10.28 g/l); less than or equal to 1.3 and 1.1% for albumin in CSF (115 mg/l) and serum (76.89 g/l). Comparison studies showed good correlation with radial immuno-diffusion (r greater than or equal to 0.995 and 0.976 for IgG and albumin) and rate nephelometry (r greater than or equal to 0.967 and 0.982 for IgG and albumin). Thus, the method under investigation proved to be reliable and appears to be particularly suitable for the routine work.     

Evaluation of a quantitative immunoturbidimetric assay for rheumatoid factors

A quantitative immunoturbidimetric assay for rheumatoid factors (RF) is described, based on the immunoprecipitation between aggregated human IgG and rheumatoid factors in serum. The resulting turbidity is measured photometrically at 340 nm. The method is standardized against the WHO international reference preparation and the results are expressed in international units per milliliter (int. units/mL). Results correlate well with those by different latex-agglutination techniques (r = 0.80-0.96). The correlation with Waaler-Rose test modifications were 0.75 and 0.92. The within-run and between-run coefficients of variations were respectively from 1.2 to 2.6% and 1.3 to 1.9% for high and low RF concentrations. Quantitative and reproducible results, together with high throughput of samples and compatibility with most clinical chemistry analyzers and photometers, make this new assay well suited for routine screening and monitoring of rheumatoid factors.     

Effect of plasma triglyceride concentrations on the accuracy of immunoturbidimetric assays of apolipoprotein B

We have studied the influence of triglyceride-rich particles on the analytical bias of apolipoprotein B measurements by various immunoturbidimetric methods. Three commercially available methods grossly overestimate apolipoprotein B in samples with even moderately above-normal triglyceride concentrations. This effect is due to the increased relative reactivity of very-low-density lipoproteins in these reagent systems, and can be eliminated by including Tween 20 (2 g/L) in the reagent buffer. We have developed, and describe, an automated immunoturbidimetric method that allows the accurate determination of apolipoprotein B in the plasma of patients with hypertriglyceridemia.     

Albumin excretion rate, albumin concentration, and albumin/creatinine ratio compared for screening diabetics for slight albuminuria

Slight albuminuria, an overnight albumin excretion rate (AER) greater than 30 micrograms/min in an "Albustix"-negative sample, predicts development of diabetic nephropathy. This study compares the AERs for 261 timed overnight urine collections with the albumin concentrations and albumin/creatinine ratios for the same specimens (equivalent to first morning specimens). Thirty-one specimens (11.9%) had AERs greater than 30 micrograms/min. Use of an albumin/creatinine ratio greater than 3.0 mg/mmol to predict an AER greater than 30 micrograms/min gave a sensitivity of 96.8%, a specificity of 93.9%, and a predictive value of 68.2%, with a correlation coefficient of 0.921. Use of an albumin concentration greater than 17 mg/L gave a sensitivity of 96.8%, a specificity of 90.9%, a predictive value of 58.8%, and a slightly poorer correlation (r = 0.904). Evidently either method is acceptable as an initial screening procedure, but determination of albumin concentration alone would be preferable because of lesser cost.     

Evaluation of Cobas 8000® for the quantification of albumin and IgG in serum and cerebrospinal fluid

According to the 2017 revised McDonald criteria, the presence of oligoclonal bands (OCB) at isoelectric focusing (IEF) is useful for the diagnosis of Multiple Sclerosis (MS), including relapsing-remitting MS and primary progressive MS. In this context, the quantification of IgG in serum and CSF is required for IEF execution (to deposit the same amount of IgG in serum and CSF), while the quantification of albumin in serum and CSF allows the calculation of the albumin quotient.We have evaluated the analytical performances of Cobas 8000® analyzer for the quantification of albumin and IgG in serum and CSF. Coefficients of variation were below 3.3% for within-run precision and below 3.1% for between-run precision. Results were similar or better than those obtained on nephelometer Immage 800® and turbidimeter SPAPLUS®. The uncertainty of quantification of IgG in CSF was 9% and that of albumin in CSF was 12%. IgG and albumin measured on Cobas 8000® in serum and CSF showed good agreement with results obtained on the nephelometer Immage 800®, including for the classification of albumin quotient and CSF IgG index as normal or pathological. Therefore, Cobas 8000® is a valuable tool for the quantification of IgG and albumin in CSF, in the context of diagnosis of MS and other inflammatory disease affecting the central nervous system.     

Screening for albuminuria: a case for estimation of albumin in urine.

The usefulness of bromcresol green for estimating albumin in urine was evaluated by comparison with the Laurell "rocket" technique. In contrast to the bromcresol green method applied for urinary albumin, rather doubtful results were obtained with conventional (Microzone) electrophoresis for albumin and with precipitation techniques for total protein estimation. Albumin estimation with bromcresol green is recommended as a more reliable substitute for total-protein estimations in urine. Limitations of bromcresol green are also pointed out.