PURPOSE: Enzyme immunoassay (EIA) is now a widely used technique. We have described the application of enzyme amplification (sensitive ELISA) in the field of immunoassays of pediatric population. There are two issues with the sensitive ELISA. First is that one can minimize the serum volume, an important concern for pediatricians. The second is the problem of background signal. We demonstrate that it is possible to develop EIAs of high sensitivity and detectability with using very small volume of infant's sera immunized with Hib-PRP vaccine. METHODS: Monoclonal Abs HG11, HP6016, HK2, and KL1 specific for human IgG1, IgG2, C , and C were used. The mAb OAK-1 specific for a subfamily of V I L chains (V Ia), the mAbs KB13 and B12 specific for human V II and V III L chains were also used respectively. Adults were immunized with Hib-CRM vaccine. Immune serum was obtained 4 to 8 wk after immunization. Twenty infants received Hib-CRM vaccine at 2, 4, and 6 month of age and blood samples were obtained at 7 month old. The amount of anti-Hib-PS Ab expressing a V subgroup or V was determined by sandwich type immunoassays using conventional substrate. The amount of the enzyme immobilized to the well was determined with para-nitrophenyl phosphate substrate. A standard ELISA was performed but different substrate (lyophilized NADPH) and amplifier (alcohol dehydrogenase and diaphorase) were used to develop color in final step for enzyme amplification method. RESULTS: We get the dose-response curves obtained using the conventional and amplified detection methods in the anti-PRP Ab assay. The sensitivities of the two assay methods were compared. We can increase the sensitivities four to sixteen folds and minimize the infant's sera volume to perform varing anti-PRP antibody assays. To obtain the advantages of increased sensitivity, any background is minimized by using noncontaminated reagents. CONCLUSIONS: It is possible to develop EIAs of high sensitivity and detectability with using very small volume of infant's sera with using enzyme amplification system (sensitive ELISA).
Adult ; Antibodies* ; Child* ; Enzyme-Linked Immunosorbent Assay* ; Humans ; Immunization ; Immunoassay ; Immunoenzyme Techniques ; Immunoglobulin G ; Indicators and Reagents ; Infant ; Oxidoreductases

Serum IgE level and Clonorchis specific IgE in individuals with Clonorchis sinensis were determined by radioimmunosorbent(RIST) and radioallergosorbent technique(RAST) respectively. Highly significant elevations of serum IgE (P<0.001) and specific IgE antibodies (P<0.01) were observed in area from individuals with clonorchiasis. The mean values of serum IgE in individuals with clonorchiasis and healthy individuals were 2,372 IU/ml and 364 IU/ml respectively and specific IgE antibodies of both groups were 52.0 and 4.4%. A close correlation(r=0.9451) between serum IgE level and specific IgE antibodies were observed and correlation (r=0.6056) between serum IgE and EPG and between specific IgE and EPG(r=0.5693) were also observed.
parasitology-helminth-trematoda ; Clonorchis sinensis ; clonorchiasis ; immunology ; radioimmunosorbent test ; radioallergosorbent test ; IgA ; IgE ; serum

BACKGROUND: Clostridium difficile is one of the most important pathogens responsible for nosocomial diarrhea. The disease is mediated by two toxins, designated as A and B; therefore, identification of the toxins is important for diagnosis. However, culture or cytotoxin assay are not easily done because of tedious procedures. Instead, toxin A immunoassay is widely used. We evaluated two different enzyme immunoassays (EIA) for C. difficile toxin A and compared them with culture and PCR results. METHODS: A total of 65 stool specimens were examined for toxin A using enzyme linked fluorescent immunoassay (ELFA, VIDAS CD II, Bio-Merieux, France) and enzyme linked immunosolvent assay (ELISA, C.DIFFICILE TOX A II, TECHLAB, USA ) and were also cultured for C. difficile using cycloserine cefoxitine fructose agar. We amplified toxin A and B genes using primers NK9-NK 11 and NK104-NK105, respectively, in 23 C. difficile isolates. RESULTS: The concordance rate between ELFA and ELISA was 76.9%. The sensitivity and specificity of the ELFA and ELISA based on the culture and PCR results for toxin A gene were 84.6%/98.1% and 84.6%/67.3%. Positive and negative predictive values were 91%/96.2% in VIDAS and 78.0%/ 94.6% in TECHLAB. The positive rates of toxin B genes were 100%, 83.3% and 50% in toxin A positive, variant and negative strains, respectively. CONCLUSIONS: The sensitivities of the ELFA and ELISA for toxin A were the same, but specificity and positive predictive value of the ELFA were higher than those of the ELISA. PCR or EIA method detecting both toxin A and toxin B is strongly recommended, because the variant strains (toxin A negative and toxin B positive) of C. difficile may be more prevalent than were anticipated in Korea.
Agar ; Cefoxitin ; Clostridium difficile* ; Clostridium* ; Cycloserine ; Diagnosis ; Diarrhea ; Enzyme-Linked Immunosorbent Assay ; Fructose ; Genes, vif ; Immunoassay ; Immunoenzyme Techniques* ; Korea ; Polymerase Chain Reaction ; Sensitivity and Specificity

BACKGROUND: Rheumatoid arthritis (RA) is the most common autoimmune rheumatic disease, but sensitive and specific test for its diagnosis is lack. This study evaluated the analytical performance and diagnostic role of a new automated ELISA anti-cyclic citrullinated peptide (anti-CCP) antibody test. METHODS: Anti-CCP antibody test was done with the enzyme-linked immunosorbent assay (ELISA) in serum samples from 49 RA patients and 104 non-RA patients, and 51 healthy subjects. Serum pools were used to determine its precision and linearity. The optimal cut-off values were determined by the receiver-operator characteristics (ROC) curve. The rheumatoid factor (RF) by turbidimetry was also assayed in every samle and the results were compared to anti-CCP for sensitivity and specificity. RESULTS: The total imprecision (CV%) was 4.8%, 7.6% for serum pools with low (mean concentration: 2.7 U/mL) and high (mean concentration :82.2 U/mL) concentration, respectively. Linearity data were acceptable (R2=0.9907). At each optimal cut-off value, the sensitivity of anti-CCP was higher than that of RF (81.6 % vs 69.4%), but statistical significance was not defined. Specificity of anti-CCP was higher than that of RF (95.5% vs 75.5%, p<0.001). A combination of anti-CCP and RF increased sensitivity and specificity to 87.7%, 98.0%, respectively. Nine of 15 (60.0%) sera from RF negative RA patients were positive for anti-CCP. CONCLUSIONS: Anti-CCP ELISA antibody test, we examined on a fully automated enzyme immunoassay, is easy to assay in routine laboratory, and showed good analytical performance. And anti-CCP antibody test also showed higher diagnostic specificity than RF. So, anti-CCP antibody may be useful serologic marker for diagnosis and monitoring of RA, if performed concomitantly with RF assay.
Antibodies* ; Arthritis, Rheumatoid ; Diagnosis ; Enzyme-Linked Immunosorbent Assay ; Humans ; Immunoenzyme Techniques* ; Nephelometry and Turbidimetry ; Rheumatic Diseases ; Rheumatoid Factor ; Sensitivity and Specificity

BACKGROUND: Hepatitis C virus (HCV) is a major agent of transfusion-associated non-A, non-B hepatitis. The best method for prevention of HCV infection through transfusion is blood donor screening using reliable diagnostic tools. Enzyme immunoassays (EIA) for detection of HCV antibody were developed, but it required equipment and time consuming. In emergency situation such as massive bleeding, HCV screening of blood donor needs more simple, rapid and reliable method. Recently, for rapid detection of HCV antibody, Genedia HCV Rapid assay was developed. To evaluate the usefulness of this assay, comparative studies with third generation anti- HCV EIA and two HCV confirmatory tests (Genedia HCV Confirm 4.0 and Chiron RIBA HCV 3.0 SIA) were performed. METHODS: A total of 156 sera (106 positive and 50 negative), screened by second generation IMx anti-HCV assay (Abbott 2.0; Abbott Laboratories, U.S.A.), were examined with Genedia HCV Rapid (Green Cross, Korea) and Genedia HCV ELISA 3.0 (Green Cross, Korea). The discrepant sera between Genedia HCV Rapid and Genedia HCV ELISA 3.0 were confirmed by both Genedia HCV Confirm 4.0 and Chiron RIBA HCV 3.0 SIA. RESULTS: The concordance rates of Abbott 2.0 vs Genedia HCV Rapid, Abbott 2.0 vs Genedia HCV ELISA 3.0, and Genedia HCV Rapid vs Genedia HCV ELISA 3.0 were 88.4%, 89.7%, and 96.1%, respectively. The sensitivity, specificity, positive predictive value and negative predictive value of Genedia HCV Rapid for Genedia HCV Confirm 4.0 were 97.7%, 100%, 98.9%, and 97.0%, respectively, and for Chiron RIBA HCV 3.0 SIA 97.8%, 98.5%, 98.9%, and 97.0%, respectively. Of discrepant 6 sera between Genedia HCV Rapid and Genedia HCV ELISA 3.0, 2 were positive by Genedia HCV Confirm 4.0, and 3 positive by Chiron RIBA HCV 3.0 SIA. However, 14 negative sera by both Genedia HCV rapid and Genedia HCV ELISA 3.0, which were all positive by Abbott 2.0, were all negative by two confirmatory tests. CONCLUSIONS: These data show that Genedia HCV Rapid could be used in emergency blood donor screening for HCV antibody detection.
Blood Donors ; Emergencies ; Enzyme-Linked Immunosorbent Assay ; Hemorrhage ; Hepacivirus* ; Hepatitis C* ; Hepatitis* ; Humans ; Immunoenzyme Techniques ; Mass Screening ; Sensitivity and Specificity

BACKGROUND: Elecsys 2010 immunoassay system is based on the electrochemiluminescence immunoassay using a ruthenium (II) tris (bipyridyl) label. Since it was the first time to use the system in our laboratory, we would like to evaluate the analytical performances (precision, linearity and recovery rate) and correlation with radioimmunoassay (RIA) and microparticle enzyme immunoassay (MEIA) methods. METHODS: We used precicontrol tumor marker (TM1, TM2) for alpha-fetoprotein (AFP), prostatic specific antigen (PSA) and carcinoembryonic antigen (CEA), Precicontrol universal (Ul, U2) for triiodothyronine (T3) and thyroxine (T4), Precicontrol-TSH for thyrotropin (TSH) and pooled serum for the evaluation of precision and recovery rate. Patients' sera were used for the linearity and comparison study. RESULTS: The coefficients of variatron of Imprecision study were below; 4.0%, 8.7% and 10.2%, respectively in the within-run, within-day and between-day analysis. The recovery rates were 100.5%, 96.1% and 102.5%, respectively in T4, TSH, and AFP. The linearity were y=1.02x-0.182(r=0.99) for T4, y=1.01x+0.12 (r=0.99) for TSH and y=1.01x+0.54(r=1.00) for AFP. T3, T4, TSH, CEA and PSA results showed good correlation with RIA (r>0.90), but AFP showed r=0.88. Also, AFP, CEA and PSA results showed excellent correlation with AxSYM (r>0.99). CONCLUSION: Elecsys 2010 immunoassay system showed excellent precision, recovery rate, clinically acceptable linearity and good correlation with the results obtained by RIA and MEIA methods.
alpha-Fetoproteins ; Carcinoembryonic Antigen ; Immunoassay* ; Immunoenzyme Techniques ; Radioimmunoassay ; Ruthenium ; Thyrotropin ; Thyroxine ; Triiodothyronine

Digoxin plays a part in healing of congestive heart failure in clinic. Its therapeutic dose is very approximate to toxic dose and even they overlap each other sometimes. There are many influencing factors on blood drug level of digoxin. Pharmacodynamics and pharmacokinetics varies with different individuality. It is indispensable to detecting blood drug level in order to treat disease and prevent intoxication. Integrating with the detecting-methods of blood drug level of digoxin home and broad, characteristic of many methods are summarized from sensitivity, linearity range, cross-reaction and precision. These methods include radio immunoassay, enzyme immunoassay, chemiluminescence immunoassay, fluorescence immunoassay and HPLC-MS-MS. These methods are popular for their specialized ascendancy. The cost of radio immunoassay is low. Enzyme immunoassay has good specificity. Sensitivity and stability of chemiluminescence immunoassay is very excellent. Fluorescence polarization immunoassay is sensitive and convenient. HPLC-MS-MS has high resolution and good specificity. One of the development tendencies is to combine two or more methods in detecting the blood drug level of digoxin which contribute to these methods integrated use.
Chemistry Techniques, Analytical ; methods ; Chromatography, High Pressure Liquid ; Digoxin ; blood ; Enzyme-Linked Immunosorbent Assay ; Fluorescence Polarization Immunoassay ; Fluoroimmunoassay ; Humans ; Radioimmunoassay ; Reproducibility of Results ; Tandem Mass Spectrometry

We measured the degree of immunity of 326 Korean children to rubella virus by enzyme-linked immunosorbent assy (ELISA). They were admitted to Pediatrics Unit of Yonsei Medical Center between April and July, 1980 with various illnesses excluding rubella disease. Among the 326 tested, 172 cases gave a positive titer of antibodies (mainly, IgG and IgM antibodies) and 127 had IgG antibody against rubella virus antigen. These represented 52.8% and 39.0%, respectively, of the total number of children tested. There was no significant difference in the rate of positivity between sex, but the positive rate increased as the age increased. The antobody titers of positive individuals to rubella virus were higher among the older children. Results and a brief outline of the ELISA method for serodiagnosis of rubella in clinical use will be discussed.
Adolescent ; Age Factors ; Antibodies, Viral/analysis* ; Child ; Child, Preschool ; Enzyme-Linked Immunosorbent Assay* ; Female ; Human ; Immunoenzyme Techniques* ; Infant ; Infant, Newborn ; Korea ; Male ; Rubella/immunology ; Rubella virus/immunology*

BACKGROUND: The serological diagnosis of herpes simplex virus type 2 (HSV-2) infection has pitfalls, in that most of the antibodies against HSV-2 cross-react with HSV-1 and the prevalence of HSV-1 infection is high, especially in Korea. In this study, we tried to establish the serological diagnostic method, which could detect and measure the specific antibodies against HSV- 2 by competitive immunofluorescent staining method as well as competitive ELISA based on the specific monoclonal antibody, MH2-7. METHODS: Immunofluorescent staining and western blot analysis were used to characterize the antigens recognized by MH2-7. Competitive immunofluorescent staining (IF), competitive enzyme immunoassay (ELISA), and western blot analysis were used to detect specific antibodies against HSV-2 in patients' sera. RESULTS: In western blot analysis, the sera from two of six patients clinically diagnosed as genital herpes showed characteristic band patterns, which have been known to be compatible with HSV-2 infection. In competitive immunofluorescent staining, only the sera from the two patients clinically diagnosed as genital herpes and with characteristic band pattern showed competition with MH2-7 monoclonal antibody. The dilution range of the serum showing specific competition was between 1:10 and 1:80. Competitive ELISA was also performed and evaluated as the diagnostic efficacy as ELISA has been known to be advantageous over IF staining in mass screening. The result showed linear dose-response relationship for the patient's sera in inhibition of the reactivity of MH2-7. CONCLUSION: We suggest that the competitive immunofluorescent staining method and competitive ELISA based on the specific monoclonal antibody MH2-7 is a simple, accurate, and precise method, which can be used in serological diagnosis of HSV-2 infection.
Antibodies ; Blotting, Western ; Diagnosis ; Enzyme-Linked Immunosorbent Assay ; Herpes Genitalis ; Herpesvirus 1, Human ; Herpesvirus 2, Human* ; Humans ; Immunoenzyme Techniques* ; Korea ; Mass Screening ; Prevalence

BACKGROUND: Mutations of the tumor suppressor gene p53 cause subsequent cellular accumulation of p53 gene product and a specific immunologic response. Detection of circulating antibodies against p53 protein has been evaluated for a tumor marker or prognostic factor for several cancers including those of the breast, stomach, ovary and lung. METHODS: Serum samples from 74 colorectal cancer patients were obtained preoperatively and anti-p53 antibody was measured by enzyme immunoassay (Anti-P53 ELISA II: PharmaCell, France). Carcinoembryonic antigen (CEA) and Carbohydrate antigen (CA) 19-9 were measured in parallel. Tissue p53 protein expression was examined by immunohistochemical staining. RESULTS: Anti-p53 antibodies were positive in the serum from 34% (25/74) of patients, but normal controls were all negative. Anti-p53 antibodies were significantly associated with p53 protein overexpression. CEA and CA19-9 were detected in 38% and 11%, respectively. There was no correlation among anti-p53 antibody, CEA and CA19-9. No differences were found between the anti-p53 antibody positive and negative groups in the following parameters: tumor site, histologic grades, Dukes stages, margin invasion, vessel invasion, lymph node and distant metastasis, and survival duration. CONCLUSIONS: The measurement of serum anti-p53 antibody is not suitable for preoperative markers of prognostic significance, but can be used as a simple serological marker for detection of p53 alteration. There should be more studies of the serum p53 antibody as a possible marker for postoperative monitoring in colorectal cancer.
Antibodies* ; Breast ; Carcinoembryonic Antigen ; Colorectal Neoplasms* ; Enzyme-Linked Immunosorbent Assay ; Female ; Genes, p53 ; Genes, Tumor Suppressor ; Humans ; Immunoenzyme Techniques ; Lung ; Lymph Nodes ; Neoplasm Metastasis ; Ovary ; Stomach