Increased plasma concentrations of tumour markers in the absence of neoplasia

Tumour markers are a very heterogeneous group of molecules that are generally found in very small concentrations in the plasma and serum of healthy individuals. In the process of neoplastic differentiation the cell can synthesize, release, or induce synthesis of other cells, thus increasing their concentration in plasma and serum. These substances may also increase their plasma concentration in patients without cancer due to processes that increase the release or reduce catabolism, and so give rise to false positives. An understanding of the main physiopathological processes that increase the concentrations of these substances could improve our interpretation of tumour markers and their clinical application. In this study we review the physiopathological processes that may increase the plasma concentrations of tumour markers. We performed a bibliography review in PubMed, searching for causes of false positives for the following tumour markers: α-Fetoprotein, CA 125, CA 15-3, CA 19-9, CA 72-4, carcinoembryonic antigen, CYFRA 21-1, squamous cell carcinoma, prostatic specific antigen, β(2)-microglobulin, choriogonadotropin (β chain), chromogranin A, neuron specific enolase, HER2-neu, progastrin releasing peptide, S-100, and thyroglobulin. The results favour the use of tests which can identify pathological processes that may increase tumour marker concentrations.     

Enzyme immunoassay for simultaneous measurement of autoantibodies against thyroglobulin and thyroid microsome in serum

A combined enzyme immunoassay (micro-ELISA) technique was established for measuring autoantibodies against thyroglobulin and thyroid microsome, involving the immuno-dot blot technique. Thyroglobulin and thyroid microsome antigens (1 g/L each) prepared from normal thyroids were spotted separately onto nitrocellulose membrane filter discs. Results by this method and those by immunofluorescence correlated well. The percentages of confirmed positives were 30% and 48% and the negatives were 58% and 46% (n = 50) for thyroglobulin and microsome, respectively. Testing these samples by gelatin agglutination gave a high percentage of false positives (up to 20%, n = 128) and hemagglutination testing yielded a high percentage of false negatives (up to 20%, n = 45). The titer of autoantibodies by the micro-ELISA technique was greater than by agglutination. This technique is highly specific and sensitive.     

Digoxygenin-labelled DNA-probe: a rapid non-radioactive method for hepatitis B virus DNA detection in serum.

The sensitivity and specificity of two non-radioactive spot hybridization assays for hepatitis B virus DNA (HBV-DNA) using biotin and digoxygenin-labelled DNA probes were investigated in parallel in 122 serum samples from patients with chronic hepatitis B and 50 controls. The results were compared with an isotopic technique using a 32P-labelled probe. HBV-DNA was detected in 56 (80%) out of 70 hepatitis B "e" antigen (HBeAg)-positive cases and in 4 (8%) out of 52 antibody to hepatitis B "e" antigen (anti-HBe)-positive cases using the digoxygenin or 32P-labelled probes. No false positives were found with either method. Using the biotin-labelled probe, 16% of sera gave discordant results, which were considered to be false positive. The time required for detection of serum HBV-DNA was 2 hours for the non-radioactive probes and 16 hours for the isotopic probes. This study suggests that the digoxygenin-labelled probe for detection of HBV-DNA is the most rapid and sensitive method for routine diagnosis of viral replication in clinical laboratories.     

Rapid detection of group B streptococcal antigen from vaginal specimens using a new Optical ImmunoAssay technique

A total of 531 vaginal specimens were used to evaluate a new Optical ImmunoAssay (OIA) screening technique for the rapid detection of group B streptococcal antigen. The results of the OIA test, the ICON Strep B membrane immunoassay (Hybritech ICON), and conventional culture on sheep blood agar (direct TSA) were compared to broth enhanced culture. Results obtained from the OIA test, ICON, and direct TSA yielded 72, 39, and 68 positives, respectively, as compared to 100 positives using the Lim broth culture method as the standard. The Optical ImmunoAssay technique is as sensitive as the conventional plating method and is capable of providing results in 30 minutes.     

False positives in plasma ammonia measurement and their clinical impact in a pediatric population

OBJECTIVE: Plasma ammonia measurement is greatly influenced by pre-analytical conditions, which may lead to false positives. We wanted to evaluate the prevalence and clinical impact of plasma ammonia false positives in a pediatric population. DESIGN AND METHODS: Over a 28-month period, charts of patients with elevated ammonemia were retrospectively reviewed to identify false positives defined as elevated concentrations that subsequently normalized without plausible explanation for the elevations. RESULTS: 1880 Ammonia measurements were available in 479 patients. Elevated results that subsequently normalized were found in 86 patients. Forty-one (48%) of these patients had most likely falsely elevated ammonemia. Additional blood sampling and laboratory testing were the most frequent consequences of false positives. CONCLUSION: There is a high proportion of false positives among elevated plasma ammonia measurements. Capillary samples and delay between sampling and centrifugation are possible contributing factors. Clinical consequences of false positives were most often limited.     

Development of a Salmonella-specific biotinylated DNA probe for rapid routine identification of Salmonella

Classical microbiological techniques used in the detection and identification of Salmonella spp. in foods, drinking water and clinical samples are relatively lengthy. Immunoassays, on the other hand, have the major disadvantage of often generating false positives and false negatives. Recombinant DNA technology offers more efficient alternatives to the detection of a specific organism by employing cloned DNA sequences unique to the organism. Demonstration of a presence of complementary sequences among a heterogeneous population of molecules of DNA isolated from bacteria can be made by using a DNA-DNA hybridization technique. We have obtained a fragment of DNA from Salmonella typhimurium chromosomal DNA, cloned it in Escherichia coli plasmid and tested it in colony hybridization tests with 57 strains of Salmonella and other enterobacteriaceae. In all tests, the fragment was found to be Salmonella-specific in that it gave a positive reaction with all strains of Salmonella tested and was negative when tested against other Enterobacteriaceae.     

The diagnostic accuracy of three recommended methods for serum aspartate aminotransferase assays in patients suspected of myocardial infarction and hepatobiliary diseases

Serum aspartate aminotransferase (AST) activity was measured by the methods recommended by the Scandinavian Committee on Enzymes (SCE) and by the International Federation of Clinical Chemistry (IFCC) with pyridoxal phosphate (PLP) and without (-PLP) in one laboratory at 37 degrees C with the Abbott ABA-100 and in another at 30 degrees C with the IL Multistat III. Reference ranges were determined on 195 healthy hospital staff. Sera from 102 patients with suspected hepatobiliary disease (HBD) and 104 with suspected myocardial infarction (MI) were assayed at both laboratories by all three methods. Based on the above reference ranges, all assays with each method at both hospitals were abnormal in 59 of 67 cases with HBD and 53 of 55 with MI. In aggregate, all three methods yielded comparable rates of misclassification (20-23). The SCE method gave highest false negatives (18) and lowest false positives (5); the IFCC method gave lowest false negatives (1) and highest false positives (20); intermediate values of 8 false positives and 12 false negatives were given by the IFCC (-PLP) method. Using receiver operating characteristic (ROC) curves, the SCE method was clearly superior at 30 degrees C, and the IFCC (-PLP) method was marginally superior at 37 degrees C. However, when the decision threshold corresponded with a 2.5% false positive rate in the non-HBD, non-MI patients, the SCE method gave the lowest false negatives at both temperatures and, on the basis of the present data, must be considered to be the method of choice for AST activity determinations.     

CA 19-9 in early cancers of the bronchi. Comparison with the carcinoembryogenic antigen

CA 19-9 (monosialoganglioside) isolated from adenocarcinomas of the gastrointestinal tract can be measured in biological fluids using a monoclonal antibody assay. A radioimmunometric assay kit produced by ORIS Industrie has been used to measure the serum level of CA 19-9 in lung cancer and the results have been compared to that of carcinoembryonic antigen. The combination of the 2 markers increase by 10% the number of subjects with a raised marker. There is no significant relationship between the levels of CA 19-9, the type of tumour or the tumour stage. The recognition of the sialic acid as an epitope by the monoclonal antibody appears to be responsible for the false positive results in non-malignant lung disease.     

Rapid large scale screening of blood donor plasma for IgA deficiency,confirmation and quantitation

Plasma from each of 20,068 blood donor samples collected for routine ABO and Rhesus grouping were screened for IgA deficiency using a rapid latex agglutination inhibition technique. 83 donors gave positive reactions with the latex screening reagent used. After elimination of the various false positives using further latex tests, 24 donors were eventually classified as true IgA deficients with an IgA level of less than 0.05 g/L, giving an incidence of 1:836.     

Extraction of human plasma or sera by heat treatment for a solid-phase radioimmunoassay of carcinoembryonic antigen.

Heat treatment and a solid-phase radioimmunoassay are combined to give a relatively simple and rapid procedure for assay of carcinoembryonic antigen in plasma or serum. The new way we describe to extract this antigen is an alternative to the conventional method of extraction with perchloric acid. Heating plasma or serum samples in acetate buffer (0.16 mol/L, pH 5.0) at 70 degrees C for 15 min precipitates out most of the heat-labile, nonspecific plasma proteins, but leaves most of the antigen in solution, with its immunochemical properties apparently unaffected. Comparison between the heat treatment and the perchloric acid extraction yielded comparable values when tested either by solid-phase radioimmunoassay or by the zirconyl phosphate precipitation method. An added advantage of our method is that it gives the same assay values for both plasma and serum. Results for a group of pathological plasma samples, assayed by both our method and the perchloric acid-zirconyl phosphate precipitation method, gave a correlation coefficient of 0.90.     

Evaluation of a latex agglutination screening test for the detection of anti-streptolysin O antibodies

A latex agglutination test (Mercia ASL Latex Kit) for the rapid detection of anti-streptolysin O (ASO) antibodies was compared with the conventional macrotitration method (Rantz-Randall) in patients with suspected streptococcal disease. Of the 64 serum specimens tested 37 were positive (≥ 200 IU ml⁻¹) for ASO antibodies by the macrotitration procedure. The latex agglutination test gave seven false positives but no false negatives and had a sensitivity of 100%, a specificity of 74·1%, a predictive positive value of 84·1% and a predictive negative value of 100%. Our results indicate that the Mercia ASL Latex Kit provides a rapid, simple, convenient and reliable test for the detection of anti-streptolysin O antibodies.     

Comparison of patient questionnaires, medical records, and plasma assays in assessing exposure to benzodiazepines in elderly subjects.

OBJECTIVE: Exposure in pharmacoepidemiologic studies can rely on various sources such as medical records, patient questionnaires, or plasma samples, which do not always concur. This study endeavored to compare sources of information on current exposure to benzodiazepines in elderly subjects. METHODS: In a study in a hospital admissions department, 1136 elderly subjects included in a case-control study each completed a structured questionnaire. In addition, an inspection of the medical records of each subject was performed, as well as screening of a plasma sample (high-pressure liquid chromatography--diode array detector) for current exposure to benzodiazepines. RESULTS: Benzodiazepines were found in the plasma of 33% of 1013 patients, in the records of 31% of patients, and in the questionnaires of 36% of 797 respondents. With use of the plasma results as a standard, questionnaires had 11% false positives and 28% false negatives; medical records had 14% false positives and 23% false negatives. The kappa for concordance between questionnaires and records was 0.63. Most of the errors were related to the unexpected presence in plasma of clorazepate, commonly used as a hypnotic agent. CONCLUSIONS: Patient recall and medical records are not reliable measures of current exposure to benzodiazepines in elderly persons, although this unreliability may be more marked with certain drugs used as hypnotic agents.     

Modification of neonatal screening test for erythrocyte glucose-6-phosphate dehydrogenase deficiency.

An improved technique is proposed for detecting G6PD deficiency on dried blood. A simple and rapid ascending chromatography of NADPH makes it possible to differentiate the false positives, too frequently found with Beutler's spot test (1968).     

The factor VIII inhibitor assays can be standardized: results of a workshop

Summary. Background: The Bethesda and the Nijmegen assays are commonly used for the measurement of inhibitor levels in hemophilia A patients. Despite test innovations, the between‐laboratory coefficient of variation (CVb) of factor VIII inhibitor test data in external quality surveys remains very high (40–60%) with a high degree of false‐negative and false‐positive results resulting in undesired effects on treatment. Objectives: Organization of a workshop in order to address the causes of this phenomenon and to suggest ways to improve the assays. Methods: Fifteen laboratories showing a high CVb in regular surveys and using a variety of methods participated in the wet workshop, which included four different sessions where variables probably contributing to the high CVb (e.g. use of [non‐]buffered plasma, FVIII‐deficient plasma, sample dilution and APTT reagents) were investigated. Results: The CVb varied from 30% to 70% in the first session of the workshop when the participants used their own test settings and reagents. The use of buffered normal pooled plasma and FVIII‐deficient plasma as a reference sample by all participants did not significantly alter the CVb (35–50%) but decreased the number of false positives. However, the use of buffered pooled plasma in combination with standardized sample dilution procedures by all participants showed a significant improvement (CVb, 10–20%). Conclusions: These results may contribute to improvement of FVIII inhibitor testing. However, improved inter‐laboratory comparison of factor VIII inhibitor assay results can only be reached when further local standardization is implemented.     

Comparison of plasma and serum for antibody detection using DiaMed microtubes

Atypical antibody detection in DiaMed microtubes using a low-ionic-strength saline (LISS) indirect antiglobulin technique (IAT) was assessed using both serum and plasma. During the first period of the study all atypical antibodies originally detected in serum were also detected in EDTA plasma with comparable reaction strengths. Two of 73 antibodies were not detected in citrated plasma.   During the second period of the study all routine antibody screens were performed in both serum and EDTA plasma. More clinically significant antibodies were detected in EDTA plasma than in serum. More false positives and non-specific antibodies were also detected in EDTA plasma. It is concluded that EDTA plasma is a suitable medium for antibody detection using LISS IAT in DiaMed gel microtubes.     

胃肠道恶性肿瘤患者血浆肿瘤型M2-丙酮酸激酶的测定及其临床价值

目的评估肿瘤型M2丙酮酸激酶（TUM2-PK）作为一种肿瘤标志物在胃肠道恶性肿瘤患者临床诊疗中的应用价值。方法用酶联免疫吸附试验（ELISA）检测106例胃肠道恶性肿瘤患者、20例胃肠道良性疾病患者以及40名健康体检者的血浆TUM2-PK，并结合癌胚抗原（CEA）、糖类抗原724（CA72-4）、糖类抗原50（CA50）等指标进行比较分析。结果胃肠道恶性肿瘤患者血浆中TUM2-PK水平显著高于胃肠道良性疾病组和健康对照组（P〈0．01），其总体敏感性为67．0％，总体特异性为78．3％。随着肿瘤病程进展，中、晚期患者血浆中TUM2-PK水平明显高于早期患者（P〈0．01）；淋巴结转移患者TUM2-PK水平显著高于未转移者（P〈0．05）。此外，联合CEA、CA72-4及CA50测定时，胃肠道恶性肿瘤诊断敏感度可有所提高。结论TUM2-PK测定可提高对胃肠道肿瘤诊断的有效性，也是胃肠道肿瘤患者病情监测及预后评估的有效指标。     

Resident-performed Compression Ultrasonography for the Detection of Proximal Deep Vein Thrombosis: Fast and Accurate

OBJECTIVES: To assess whether emergency medicine residents (EMRs) could quickly perform accurate compression ultrasonography (CUS) for the detection of proximal lower extremity deep vein thromboses (PLEDVTs) with minimal training. METHODS: A prospective, observational study using a convenience sample of patients presenting with signs and/or symptoms for PLEDVT. Vascular laboratory and department of radiology studies were considered the criterion standard. CUS of the femoral vessels was performed. Incompressibility or visualized thrombus was considered "positive." RESULTS: Eight residents with limited ultrasound (US) experience and no prior experience with deep vein thrombosis (DVT) US volunteered to participate in this study, enrolling 72 patients. Their average scan time was 11.7 minutes (95% CI = 9.4 to 14). There were 23 true positives, 4 false positives, 45 true negatives, and 0 false negatives. The test characteristics for PLEDVT gave a sensitivity of 100% (95% CI = 82.2 to 100) and a specificity of 91.8% (95% CI = 79.5 to 97.4). CONCLUSION: Emergency medicine residents with limited US experience were able to quickly perform CUS after minimal training for the detection of PLEDVT in a select group of patients.     

Evaluation of an enzyme immunoassay-based stool antigen test to detect Campylobacter jejuni and Campylobacter coli

An enzyme immunoassay-based antigen test (Ridascreen Campylobacter; R-Biopharm, Darmstadt, Germany) was evaluated for the detection of Campylobacter jejuni and Campylobacter coli in 1050 clinical stool samples as compared with culture on selective medium. After routine inoculation for Salmonella, Shigella, Yersinia, Aeromonas, Plesiomonas, and Campylobacter, the same swab specimens were used for the antigen test. The positivity rate for Campylobacter was 9.3% in culture, and the antigen test gave a sensitivity of 69%. Forty-six stool samples culture-negative for Campylobacter grew other enteropathogens; one (positive for Salmonella sp.) was positive in the antigen test. Of all the 952 Campylobacter culture-negative samples, 830 were negative in the antigen test, giving a specificity of 87%. Almost 5% of the samples showed equivocal antigen test results. If the moderate sensitivity of the antigen test was due to a low sensitivity of culture or receiving the stool samples in transportation tubes remains to be studied.