Automated Enzyme Immunoassay Research Articles

Amino Terminal pro B-Type Natriuretic Peptide Levels Are Elevated in the Cord Blood of Neonates With Congenital Heart Defect

This prospective study investigates whether amino terminal proB-type natriuretic peptide (NT-proBNP) levels are elevated in neonates with congenital heart defect (CHD). NT-proBNP levels in the umbilical cord blood of 60 neonates with prenatally diagnosed CHD, in the cord blood of 200 control subjects, and in the plasma of the respective mothers were analyzed using an automated enzyme immunoassay. NT-proBNP concentrations in the cord blood of the CHD group were significantly elevated compared with controls [median (range): 158 pg/mL (378-3631 pg/mL) and 626 pg/mL (153-2518 pg/mL); p value <0.001]. The NT-proBNP concentrations of the newborns and their mothers did correlate neither in the CHD nor in the control group. In 54 patients with CHD, NT-proBNP levels were measured on the median 5th day of life. They showed a significant increase (median 1665 pg/mL and 19222 pg/mL; p < 0.001). NT-proBNP levels in the cord blood of neonates with CHD are significantly elevated and show a marked increase in the first week of life. Furthermore, this study confirms previously published reference intervals of NT-proBNP in the cord blood of healthy neonates as well as the finding that there is no exchange of NT-proBNP across the placenta.

Aminoterminal brain-type natriuretic peptide levels correlate with heart failure in patients with bidirectional Glenn anastomosis and with morbidity after the Fontan operation

The aims of this study were to generate normal values of aminoterminal pro-brain natriuretic peptide in children with a bidirectional Glenn anastomosis without congestive heart failure and to test the hypothesis that plasma levels of aminoterminal pro-brain natriuretic peptide correlate with the clinical severity of congestive heart failure and morbidity after the Fontan operation. Aminoterminal pro-brain natriuretic peptide plasma levels of 78 patients after the bidirectional Glenn operation with a median age of 3.2 years and a median follow-up time of 3 years were measured by using an automated enzyme immunoassay. The severity of heart failure was quantified by using the New York University Pediatric Heart Failure Index. The 97.5th percentile of aminoterminal pro-brain natriuretic peptide level in patients without congestive heart failure was 339 pg/mL. Aminoterminal pro-brain natriuretic peptide levels strongly correlated with the New York University Pediatric Heart Failure Index score (P < .001). In patients with congestive heart failure (31/78), the aminoterminal pro-brain natriuretic peptide levels were significantly higher (median, 670 pg/mL) than in patients without congestive heart failure (median, 171 pg/mL). In 41 patients who underwent the Fontan operation, the time to removal of chest tubes and the length of hospital stay positively correlated with the preoperative value of aminoterminal pro-brain natriuretic peptide. In children with a bidirectional Glenn anastomosis without signs of heart failure, aminoterminal pro-brain natriuretic peptide levels were within the normal range and correlated with the severity of congestive heart failure. Further studies are needed to determine whether aminoterminal pro-brain natriuretic peptide levels can aide clinicians in the early detection of congestive heart failure in this patient group.

Serum autotaxin measurement in haematological malignancies: a promising marker for follicular lymphoma

Autotaxin (ATX) is a tumour cell motility-stimulating factor originally isolated from melanoma cell supernatants. ATX is identical to lysophospholipase D, which produces a bioactive lipid mediator, lysophosphatidic acid (LPA), from lysophosphatidylcholine. ATX is overexpressed in various malignancies, including Hodgkin lymphoma, and ATX may stimulate tumour progression via LPA production. The present study measured the serum ATX antigen levels in patients with haematological malignancies using a recently developed automated enzyme immunoassay. The serum ATX antigen levels in patients with B-cell neoplasms, especially follicular lymphoma (FL), were higher than those in healthy subjects. Serum ATX antigen levels in FL patients were associated with tumour burden and changed in parallel with the patients' clinical courses. The serum ATX antigen levels were little affected by inflammation, unlike the soluble interleukin-2 receptor and beta2-microglobulin levels. As expected, the plasma LPA levels in FL patients were correlated with the serum ATX antigen levels. Given that leukaemic tumour cells from FL patients expressed ATX, the shedding of ATX from lymphoma cells probably leads to the elevation of serum ATX antigen levels. Our results suggest that the serum ATX antigen level may be a promising and novel marker for FL.

Aminoterminal Brain Natriuretic Peptide Levels in Children and Adolescents After Fontan Operation Correlate with Congestive Heart Failure

The aim of this study was to generate normal values of amino-terminal pro-brain natriuretic peptide (NT-pro-BNP) in children and adolescents after Fontan operation without congestive heart failure (CHF) and to test the hypothesis that plasma levels of NT-pro-BNP correlate with the clinical severity of CHF. NT-pro-BNP plasma levels of 59 consecutive patients, with a median age of 8.4 years, after Fontan operation were measured using an automated enzyme immunoassay. The 97.5th percentile of NT-pro-BNP in patients without CHF was 282.3 pg/ml. The severity of heart failure was quantified by a pediatric cardiologist using the New York University Pediatric Heart Failure Index (NYUPHFI). NT-pro-BNP levels correlated with the NYUPHFI (p = 0.001). In patients with CHF (14/59) the NT-pro-BNP levels were significantly higher (median, 399 pg/ml; range, 140-5440 pg/ml) than in patients without CHF (median, 96 pg/ml; range, 11-376 pg/ml). NT-pro-BNP levels of patients with Fontan circulation without CHF are similar to those of healthy children. Plasma NT-pro-BNP concentrations correlate with the severity of CHF in children and adolescents after Fontan operation. Plasma NT-pro-BNP levels can help clinicians in the detection of CHF in pediatric patients with Fontan circulation.

Interleukin 1β Levels around Microscrew Implants during Orthodontic Tooth Movement

To determine whether interleukin 1beta (IL-1beta) levels are elevated around microscrew implants that are used as anchorage for tooth movement. Ten young adults, aged 16.3 +/- 2.5 years and with all four premolars extracted , comprised the study group. Twenty maxillary microscrew implants were placed bilaterally in the alveolar bone between the maxillary second premolars and first molars as anchorage units for distal movement of the maxillary canines. The maxillary canines served as the treatment group, and the microscrew implants were designated as the implant group. The mandibular canines were used as controls. Peri-microscrew implant crevicular fluid (MICF) and gingival crevicular fluid (GCF) were collected at the beginning of tooth movement (2 weeks after implant placement); at 24, 48, and 168 hours later; and on days 14 and 21. An automated enzyme immunoassay was used to measure 1L-1beta in the MICF and the GCF. The mean 1L-1beta level in the treatment group was significantly elevated at 24 hours (P = .003 < .05) and 48 hours (P = .003 < .05), whereas the levels in the control and implant groups did not change significantly during the experimental period. Also, the mean 1L-1beta level of the treatment group was significantly higher that in both the control and implant groups at 24 and 48 hours. The microscrew implants did not demonstrate increased 1L-1beta levels during tooth movement. This supports the concept that microscrew implants might be useful as absolute anchorage devices.

Detection of virus-specific intrathecally synthesised immunoglobulin G with a fully automated enzyme immunoassay system

BackgroundThe determination of virus-specific immunoglobulin G (IgG) antibodies in cerebrospinal fluid (CSF) is useful for the diagnosis of virus associated diseases of the central nervous system (CNS) and for the detection of a polyspecific intrathecal immune response in patients with multiple sclerosis. Quantification of virus-specific IgG in the CSF is frequently performed by calculation of a virus-specific antibody index (AI). Determination of the AI is a demanding and labour-intensive technique and therefore automation is desirable. We evaluated the precision and the diagnostic value of a fully automated enzyme immunoassay for the detection of virus-specific IgG in serum and CSF using the analyser BEP2000 (Dade Behring).MethodsThe AI for measles, rubella, varicella-zoster, and herpes simplex virus IgG was determined from pairs of serum and CSF samples of patients with viral CNS infections, multiple sclerosis and of control patients. CSF and serum samples were tested simultaneously with reference to a standard curve. Starting dilutions were 1:6 and 1:36 for CSF and 1:1386 and 1:8316 for serum samples.ResultsThe interassay coefficient of variation was below 10% for all parameters tested. There was good agreement between AIs obtained with the BEP2000 and AIs derived from the semi-automated reference method.ConclusionDetermination of virus-specific IgG in serum-CSF-pairs for calculation of AI has been successfully automated on the BEP2000. Current limitations of the assay layout imposed by the analyser software should be solved in future versions to offer more convenience in comparison to manual or semi-automated methods.

Clinical evaluation of a microsphere bead-based flow cytometry assay for the simultaneous determination of anti-thyroid peroxidase and anti-thyroglobulin antibodies

Objectives: Multiplexing technologies based on the use of microspheres as the solid phase have opened new possibilities for the analysis of autoantibodies. As an alternative to the traditional immunoassays, it is possible to use these methods in combination with flow cytometry for simultaneous measurement of anti-thyroid peroxidase (anti-TPO) and anti-thyroglobulin (anti-Tg) antibodies. Design and methods: We studied 127 serum samples sent to our laboratory for the quantitation of anti-TPO and anti-Tg antibodies. Clinical information was available for all of the patients studied. The samples were analyzed simultaneously for both antibodies by flow cytometry (FIDIS, BMD, France), and individually for each of the antibodies by an automated enzyme immunoassay (UniCap, Pharmacia Diagnostics, Germany). Results: A significant association between the results was observed. The kappa agreement indices between the methods were 0.859 and 0.832 for anti-TPO and anti-Tg, respectively. Discrepant results between the two techniques were observed with no common cause. Anti-TPO and anti-Tg antibodies exhibited a non-Gaussian distribution. The areas under the ROC curves were similar for both methods used; for anti-TPO, 0.884 (Pharmacia) and 0.853 (BMD), and for anti-Tg, 0.833 (Pharmacia) and 0.837 (BMD). Conclusion: Cytometry multiplex technology offers a true alternative to conventional immunoassays in the analysis of anti-TPO and anti-Tg antibodies.

Determination of opiates and cocaine in hair using automated enzyme immunoassay screening methodologies followed by gas chromatographic–mass spectrometric (GC–MS) confirmation

The objective of this study was to develop a two-step strategy for analysis of opiates and cocaine in hair samples involving an immunological screening procedure followed by confirmation of results using gas chromatography–mass spectrometry (GC–MS). A semi-quantitative automated competitive enzyme-linked immunosorbent assay (ELISA) methodology using Oral Fluid Micro-Plate Enzyme Immunoassays (Orasure Technologies, Inc.) was developed and validated. Applicability was proven by analysis of authentic head hair samples from drug users ( n = 103) and from opiate associated fatalities ( n = 21). The optimum cutoff values for the ELISA tests were 0.1 ng cocaine-equivalents/mg hair and 0.05 ng morphine-equivalents/mg hair using a 50 mg hair sample. Both ELISA tests had a sensitivity of 100%, the specificity was 66% for cocaine-equivalents and 42% for morphine-equivalents. The intraassay precision was 11% for the cocaine and 3% for the opiates ELISA, while interassay precision was 12% for the cocaine and 4% for the opiates ELISA test. The actual analyte concentrations in the hair samples were determined using GC–MS and were between 0.04 and 5.20 ng/mg for heroin (HER), between 0.04 and 30.01 ng/mg for 6-monoacetylmorphine (MAM), between 0.03 and 11.87 ng/mg for morphine (MOR), between 0.02 and 1.84 ng/mg for codeine (COD), between 0.02 and 2.48 ng/mg for acetylcodeine (AC), between 0.01 and 21.37 ng/mg for cocaine (COC), between 0.03 and 10.51 ng/mg for benzoylecgonine (BE) and between 0.05 and 1.26 ng/mg for cocaethylene (CE). The automated ELISA tests were proven to be valid screening procedures for the detection of cocaine and opiates in hair as confirmed by GC–MS. Screening methods provide rapid and inexpensive automated pre-test procedures to detect drugs in hair or other matrices. For forensic purposes screening therefore represents an ideal complement to routinely applied GC–MS procedures.

Anti-cyclic citrullinated peptide autoantibodies measured by an automated enzyme immunoassay: Analytical performance and clinical correlations

Background Autoantibodies against cyclic citrullinated peptide (anti-CCP) are considered to be a sensitive and specific marker for rheumatoid arthritis (RA). This study evaluated the analytical performance and clinical correlation of an automated enzyme immunoassay (DSX, DINEX Technologies), for the detection of anti-CCP autoantibodies (DIASTAT™ anti-CCP, Axis-Shield, DUNDEE UK). Methods Commercial controls and serum pools were used to determine its precision, analytical sensitivity, functional sensitivity and linearity. Sera from 83 patients with established RA and from 140 controls, including patients with various autoimmune diseases, viral infections and cancer, as well as sex- and age-matched healthy subjects, were studied. The rheumatoid factor (RF) was also assayed in each sample, and the results were compared to the anti-CCP findings. Results The total imprecision (CV%) was 4.7–7.2% for concentrations ranging between 1.98 and 71.81 U/mL. The lower detection limit was 0.038 U/mL. At a cut-off of 5 U/mL, the sensitivity and specificity for RA were 67.5% and 99.3%, respectively. The RF had a sensitivity of 66.3% and a lower specificity 82.1% than anti-CCP. When the two antibodies were used together, the specificity was 99.1%. Conclusions The anti-CCP assay we examined on a fully automated system showed a good analytical performance (analytical and functional sensitivity, linearity) and good clinical correlation. We conclude that this system can provide rapid, useful data.

Evaluation of the relationship between cockroach sensitivity and house-dust-mite sensitivity in Turkish asthmatic patients

Exposure to cockroach has been identified as an important source of indoor allergens in patients with asthma and allergic rhinitis. We evaluated the relationship between cockroach sensitivity and other allergens in patients with asthma. A total of 114 patients, defined asthma according to GINA, were enrolled in this study. A questionnaire including age, sex, duration of asthma, history of cockroach presence at home, and total IgE, blood eosinophil count, pulmonary function tests, standard skin prick test additional cockroach and shrimp allergen were performed. There were 84 (73.7%) female and 30 (26.3%) male patients with a mean age of 38.1+10.1 years. The average duration of asthma was 7.7+7.2 years. Sixty five (57%) patients were determined atopic and 49 (43%) nonatopic. Pollen allergen was the most common allergen in 59 (51.8%) patients with asthma, and second common allergen was mite allergen in 43 (37.7%) patients. Cockroach sensitivity were detected in 23 (20.2%) of 114 all asthmatics and 23 (35%) of atopic asthmatics. High rates of house-dust-mite allergy (73.9%) was determined in patients with cockroach sensitivity (P<0.05), while we found no relationship with other allergens. There was no difference for cockroach sensitivity between rural and urban population. Cockroach sensitivity was more common in mild bronchial asthmatics and a female predominance was observed. In addition, there was no association between shrimp and cockroach sensitivity. As a result, a high rate of cockroach sensitivity alone or with mite sensitivity was seen in patients with bronchial asthma in Turkish population. Because of cross-reactivity between mites and cockroach, cockroach sensitivity should be investigated in patients with house-dust-mite allergy. In addition, a high rate of cockroach sensitivity, in terms of IgE sensitization, may be important for the development of new sensitizations.

Improvement of the Lectin-Antibody Enzyme Immunoassay of the Alphafetoprotein Carbohydrate Chain for Automation with the Enzyme Immunoassay Robot

The lectin-antibody enzyme immunoassay of the alphafetoprotein-L3 carbohydrate chain, a tumor marker of liver cancer, has not been automated. We improved the technique of the assay for automation. Consequently, alphafetoprotein-L3 and total alphafetoprotein were detected with two lectins using an automatic paramagnetic bead handling robot. This indicates that the improved method is potentially applicable to the automated enzyme immunoassay robot.

Estudio seroepidemiológico de la hepatitis A en la comunidad de Madrid durante el año 2002

The aim of this study was to investigate the prevalence and risk factors associated with hepatitis A virus (HAV) infection in a representative population sample to determine who can benefit from vaccination strategies and to investigate the age limit at which previous HAV antibody screening is not required. From April to September 2002, we studied a total of 557 patients, 90 children and 467 blood donors, aged 1-65 years. Information on demographic variables (age, gender, place of residence and education level) was recorded. Patients with history of hepatitis or other liver diseases were excluded. Anti-HAV antibodies (IgG) were determined with an automated enzyme immunoassay (AxSYM, Abbott Diagnostics). The chi-square and Mantel-Haenszel tests were used for the statistical analysis. The overall prevalence of HAV infection was 41.5%. There was a significant increase in prevalence with age (chi-square TL:205, P < .0001), with rates from the youngest to oldest groups of 5.5%, 23.5%, 28.1%, 64.2% and 93.2%, respectively. Apart from age, the only other risk factor independently associated with prevalence was the level of education, with higher prevalence at the lower education levels (OR 5 2.7; chi-square = 32.11, P < .0001). The prevalence of anti-HAV antibodies has decreased in recent years in the community of Madrid. Among the population less than 35 years of age, 75% of individuals are susceptible to the infection and could benefit from universal vaccination without previous screening for anti-HAV antibodies.

Comparison of some effects of acetylsalicylic acid and rofecoxib during orthodontic tooth movement

The purpose of this study was to examine the effects of 2 different anti-inflammatory drugs on gingival crevicular fluid (GCF) volume and on prostaglandin E 2 (PGE 2) levels of the GCF during orthodontic tooth movement. A total of 36 extraction patients, aged 17.6 ± 2.5 years (mean ± standard deviation), were divided into 3 groups. Acetylsalicylic acid (aspirin) and rofecoxib (Vioxx, Merck, Whitehouse Station, NJ) were used for pain control in the first and second groups; the third group was used as a control. Gingival crevicular fluid was sampled at the beginning of tooth movement and at 24, 48, and 168 hours. An automated enzyme immunoassay was used to measure PGE 2 in GCF. The intragroup differences were evaluated with the Wilcoxon test, and the differences between the groups were determined with the Mann-Whitney U test. Gingival crevicular fluid volumes of the groups did not change significantly during the experimental period. Depending on the variations of fibroblast activation, PGE 2 levels of all the groups increased at 24 and 48 hours and decreased at 168 hours. When the drugs were compared, it was found that the inhibition effect of aspirin on PGE 2 was more than that of rofecoxib. The results suggest that rofecoxib can be used during orthodontic treatment, but further study is recommended.

Identifying recent HIV infections using the avidity index and an automated enzyme immunoassay.

We evaluated a procedure for identifying recent HIV infections, using sequential serum samples from 47 HIV-positive persons for whom the seroconversion date could be accurately estimated. Each serum sample was divided into two aliquots: one diluted with phosphate-buffered saline and the other diluted with 1 M guanidine. We assayed the aliquots with the automated AxSYM HIV1/2gO test (Abbott Diagnostics Division), without modifying the manufacturer's protocol. We then calculated the avidity index (AI): the ratio of the sample/cutoff value for the guanidine aliquot to that of the phosphate-buffered saline aliquot. We analyzed 216 serum samples: 34 samples were collected within 6 months of seroconversion (recent seroconversions), and 182 were collected after 6 months. The mean AIs, by time from seroconversion, were 0.68 +/- 0.16 (within 6 months) and 0.98 +/- 0.10 (after 6 months) (P < 0.0001). AI of <0.90 correctly identified 88.2% of recent infections but misclassified as recent infections 13.2% of serum samples collected afterward. The probability of an infection being classified as recent and having AI of > or = 0.90 would be 0.7% in a population with 5% recent infections. AI can identify with a certain degree of accuracy recent HIV infections, and being a quantitative index, it provides different levels of sensitivity and specificity, depending on the selected cutoff value. The standard assay procedure is not modified. This test is simple and inexpensive and could be used for surveillance, decision-making in treatment, and prevention.

Developing an Automated Enzyme Immunoassay for High-Throughput Screening of Bovine Serum Antibodies to Neospora Caninum

An enzyme-linked immunosorbent assay (ELISA) for Neospora caninum antibodies was automated with a robotic workstation, the Beckman Coulter Biomek 2000, to screen 200 bovine sera. Comparing these results with manually run ELISA data, a 95.92% agreement (K = 0.9592) between the two assays was obtained. The automated assay was specific and sensitive with excellent positive and negative predictive values. The results were repeatable and reproducible. The automation flexibility was high and the operation complexity was minimal. High-throughput screening (HTS) for bovine antibodies to Neospora caninum was achieved. The assay was developed according to the internationally recognized ISO17025 standard requirements.

Developing an Automated Enzyme Immunoassay for High-Throughput Screening of Bovine Serum Antibodies toNeospora Caninum

An enzyme-linked immunosorbent assay (ELISA) for Neospora caninum antibodies was automated with a robotic workstation, the Beckman Coulter Biomek 2000, to screen 200 bovine sera. Comparing these results with manually run ELISA data, a 95.92% agreement (K = 0.9592) between the two assays was obtained. The automated assay was specific and sensitive with excellent positive and negative predictive values. The results were repeatable and reproducible. The automation flexibility was high and the operation complexity was minimal. High-throughput screening (HTS) for bovine antibodies to Neospora caninum was achieved. The assay was developed according to the internationally recognized ISO17025 standard requirements.

Antibodies to centromere antigens measured by an automated enzyme immunoassay

Background: Anticentromere antibodies (ACA) are frequently observed in patients with Raynaud's phenomenon and in the CREST syndrome, a subclass of systemic sclerosis. Likewise, ACA are also found in other autoimmune and non-autoimmune diseases. The objective of the present study was to evaluate the clinical utility of the measurement of antibodies to the best characterized centromere antigen (CENP-B) protein by an enzyme-linked immunosorbent assay (ELISA) that uses human recombinant CENP-B antigen and compare it with indirect immunofluorescence assay (IFA) on HEp-2 cells. Methods: We have analyzed 128 sera samples from patients with the following diseases: systemic lupus erythematosus (SLE, n=53), mixed connective tissue disease ( n=1), primary Sjögren syndrome ( n=10), primary Raynaud's phenomenon ( n=10), primary systemic sclerosis ( n=7), polymyositis/dermatomyositis ( n=3), rheumatoid arthritis ( n=9), cutaneous lupus ( n=5), primary biliary cirrhosis ( n=9), chronic autoimmune hepatitis ( n=5) and ANA-positive non-autoimmune diseases ( n=16). Results: The ELISA evaluated shows a good concordance with IFA, with the advantage of being an automatable quantitative technique. Conclusions: Measurement of anticentromere antibodies by this ELISA using human recombinant antigen is a useful alternative for the autoimmune laboratory checking for diseases associated with anticentromere antibodies.

Automation of Serological Diagnosis for Herpes B Virus Infections Using Robot-Assisted Integrated Workstations

A robot-assisted automated facility was established within a biosafety level 3 laboratory located on a university campus. A unique integration of a Genesis (TECAN) liquid handler into the SAGIAN Core System (Beckman-Coulter) enabled tube liquid handling and transfer of reagents from tubes to microplates. An automated enzyme immunoassay (ELISA) for detecting herpes B virus antibodies was developed. Repetition experiments of the automated ELISA and comparison to the manual ELISA demonstrated the efficacy and high reproducibility of the automated method. It is anticipated that research and education activities will benefit from the establishment of the automated diagnostic laboratory in an academic environment.

Immunosuppressant effect of gold on IgG subclasses and IgE; evidence for sparing of Th2 responses.

We set out to examine the effect of gold treatment on the Th2-dependent antibodies IgG4 and IgE in relation to other IgG subclasses in patients with rheumatoid arthritis (RA). Eighty-five gold-treated RA patients and 82 RA controls were studied. Serum IgG subclass concentrations were measured by ELISA, IgE was measured by automated enzyme immunoassay. Samples were studied serially in 13 gold-treated patients and in 11 patients with gold-induced adverse events. There was a significant reduction in the concentration of IgG1, IgG2 and IgG3 in gold-treated RA patients compared with RA controls (P 0.004-0.019), whereas IgG4 was less significantly reduced in gold-treated patients (P = 0.044) and there was no difference in IgE. In serial samples there was a significant fall in the concentration of IgG1 (P = 0.001), IgG2 (P = 0.001) and IgG3 (P = 0.026) with time but no change in IgG4 and IgE. The development of gold-induced adverse events was not associated with any change in the concentration of each IgG subclass or IgE. Deficiencies of IgG subclasses were found in 30% of gold-treated RA patients and 8.5% of RA controls, and were associated in gold-treated patients with a longer disease duration (P = 0.003) and with erosive disease (P = 0. 03). IgG2 was affected most frequently and in the majority of these cases subnormal specific IgG2 binding to widespread polysaccharide antigens (Pneumovax II) was found. Gold induces an overall immunosuppressant effect on IgG subclasses, with a deficiency in 21. 5%, adjusted for controls. The effect on the Th2-dependent antibodies IgG4 and IgE is less marked, suggesting a sparing of Th2 responses.

Evaluation of HER-2/neu in serum and tissue of primary and metastatic breast cancer patients using an automated enzyme immunoassay.

Serum HER-2/neu concentrations were evaluated in 172 healthy subjects, 176 primary and 55 metastatic breast cancer patients, employing a new automated assay (Bayer Immuno 1 serum HER-2/neu). Using 13 ng/mL as the cutoff, abnormal HER-2/neu serum levels were found in 8% (14/176) of primary and 50.9% (28/55) of metastatic breast cancer patients. Both in primary and metastatic breast cancer a significant relationship was found with the stage of the disease when serum HER-2/neu was considered as a categorized variable (p=0.0003 and p=0.02, respectively), but not when it was taken as a continuous variable (p=0.247 and p=0.146, respectively). Moreover, we evaluated the correlation between Immuno 1 HER-2/neu and Oncogene Research Products ELISA assay in 53 normal subjects, 46 primary and 34 metastatic breast cancer patients. The correlation was relatively good (p<0.0001), although substantial differences could be found in single cases. The Immuno 1 assay was also evaluated for the first time in breast cancer tissue. The method, which showed good performance both in terms of imprecision and linearity, was used to measure HER-2/neu protein in 140 cytosol samples from primary breast cancer tissue and in homogenates from 40 matched cases. The correlation between the two matrixes was very close (p<0.0001). By contrast, no correlation was found between serum and matched cytosol (p=0.101) or homogenate samples (p=0.511).