Current diagnosis of human immunodeficiency virus (HIV) infection relies on the detection of anti-HIV antibodies by enzyme-linked immunosorbent assay (ELISA). Recently, kits detecting both p24 antigenemia and anti-HIV/anti-HIV2 antibodies have been developed. Thus, it is necessary to compare those kits developed as such. The aim of this study was to evaluate the diagnostic efficiency of a simultaneous detection test of p24 antigen and anti-HIV1/2 antibodies in a low prevalence area. Eight hundred and four randomly selected sera proven negative for HIV infection and 110 sera from 54 patients diagnosed as HIV infected, obtained between 1999 and 2000, were used for this study. One commercial lot of panels composed of consecutive sera obtained from known HIV-infected patient was included. Anti-HIV1/2 antibodies were detected by two different commercial ELISA kits, one from Korean and the other from German manufacturer. P24 antigen test was performed by ELISA. The simultaneous HIV antigen and antibody detection test was carried out. In the meantime, HIV RNA PCR and anti-HIV and anti-HIV2 western blot assays were also performed to confirm the test results in cases the test results didn't agree. The simultaneous detection kit showed 100% sensitivity and 99.6% specificity. Furthermore, the test displayed the possibility of earlier diagnosis than conventional anti-HIV1/2 ELISA with the results obtained from a group of consecutive panel sera infected with HIV. From these results, we concluded that the simultaneous HIV antigen and antibody detection test can be applied as a substitute clinical screening test in the place of conventional anti-HIV1/2 ELISA, and there is the probable benefit of early diagnosis.
Enzyme-Linked Immunosorbent Assay/standards* ; Enzyme-Linked Immunosorbent Assay/instrumentation ; HIV Antibodies/analysis* ; HIV Antigens/analysis* ; Human ; Korea ; Reagent Kits, Diagnostic/standards

Due to the high variation in test results of indirect enzyme-linked immuno sorbent assay (ELISA) and complicated steps involved in the process of standardization, a platform used for standardizing the test results from indirect ELISA was developed. The platform was designed based on 'Improved Standardization Method for Optical Density' (I-STOD). Gauss-Newton iteration was applied to estimate parameters in a standard formula. Programming Language VB was used for developing interface of platform. The results indicated that the validity of experiment could be verified through platform. A well determined scope of standardization could be generated. The sample with concentration within the scope was standardized and the degree of dilution was calculated for those outside the scope. The platform was successfully developed which normalized the process of standardization. The function provides the researchers with an effective and convenient tool for quickly achieving standardization of ELISA test results.
Antibodies ; analysis ; Enzyme-Linked Immunosorbent Assay ; methods ; standards ; Humans ; Optical Phenomena

OBJECTIVETo establish a model for one choosing controls with a suitable concentration for internal quality control (IQC) with qualitative ELISA detection, and a consecutive plotting method on Levey-Jennings control chart when reagent kit lot is changed.

METHODSFirst, a series of control serum with 0.2, 0.5, 1.0, 2.0 and 5.0ng/ml HBsAg respectively were assessed for within-run and between-run precision according to NCCLs EP5 document. Then, a linear regression equation (y=bx + a) with best correlation coefficient (r > 0.99) was established based on S/CO values of the series of control serum. Finally, one could choose controls with S/CO value calculated from the equation (y = bx + a) minus the product of the S/CO value multiplying three-fold between-run CV to be still more than 1.0 for IQC use. For consecutive plotting on Levey-Jennings control chart when ELISA kit lot was changed, the new lot kits were used to detect the same series of HBsAg control serum as above. Then, a new linear regression equation (y2 = b2x2 + a2) with best correlation coefficient was obtained. The old one (y1 =b1x1 + a1) could be obtained based on the mean values from above precision assessment. The S/CO value of a control serum detected by new lot kit could be changed to that detected by old kit lot based on the factor of y2/y1. Therefore, the plotting on primary Levey-Jennings control chart could be continued.

RESULTSThe within-run coefficient of variation CV of the ELISA method for control serum with 0.2, 0.5, 1.0, 2.0 and 5.0ng/ml HBsAg were 11.08%, 9.49%, 9.83%, 9.18% and 7.25%, respectively, and between-run CV were 13.25%, 14.03%, 15.11%, 13.29% and 9.92%. The linear regression equation with best correlation coefficient from a test at random was y = 3.509x + 0.180. The suitable concentration of control serum for IQC could be 0.5ng/ml or 1.0ng/ml. The linear regression equation from the old lot and other two new lots of the ELISA kits were y1 = 3.550(x1) + 0.226, y2 = 3.238(x2) +0.388, and y3 =3.428(x3) + 0.148, respectively. Then, the transferring factors of 0.960 (y2/y1) and 0.908 (y3/y1) were obtained.

CONCLUSIONThe results shows that the model established for IQC control serum concentration selecting and for consecutive plotting on control chart when the reagent lot is changed is effective and practical.

Enzyme-Linked Immunosorbent Assay ; methods ; standards ; Evaluation Studies as Topic ; Hepatitis B Surface Antigens ; blood ; Humans ; Quality Control ; Reagent Kits, Diagnostic ; standards ; Reproducibility of Results

BACKGROUNDTo evaluate homemade and imported HbsAg ELISA kits on screening blood donors.

METHODSSamples for evaluation included 120 HbsAg serum plates for the golden criteria and 400 sets of serum from blood donors in Dongguan. The samples underwent blind screening with homemade and imported ELISA kits respectively.

RESULTSThe sensitivity of homemade (Xinchuang) and imported (Diasorin) HbsAg ELISA kit were 85.71% (72/84) and 100% (84/84), respectively. Their specificity was 100% (436/436) and 96.55% (421/436) respectively. The consistency of two ELISA kits was 100%.

CONCLUSIONThe imported ELISA kit had the highest sensitivity, but its specificity was not as good as that of homemade ELISA kit. The two kinds of ELISA kits had good repetition. The combination of the two reagents may ensure the safety of blood transfusion.

Blood Donors ; China ; Enzyme-Linked Immunosorbent Assay ; instrumentation ; methods ; standards ; Hepatitis B ; blood ; diagnosis ; prevention & control ; Hepatitis B Surface Antigens ; blood ; Humans ; Mass Screening ; methods ; standards ; Reagent Kits, Diagnostic ; standards ; Reference Standards ; Reproducibility of Results ; Sensitivity and Specificity

BACKGROUND: Current status of external quality assessment (EQA) of laboratory tests for syphilis in Korea was analyzed to find out the problems that should be improved in the future. METHODS: Based on the data from the external quality assessment program performed twice a year by the Immunoserology Subcommittee of the Korean Association of Quality Assurance for Clinical Laboratory from the year 2004 to 2006, discordance rates were analyzed according to the test method and commercial kit used. RESULTS: Among the laboratories participating in the EQA program for syphilis test, about 90% of them used non-treponemal tests and about 55% treponemal tests. The non-treponemal tests included RPR (rapid plasma reagin) and VDRL tests used in 88% (363/412) and 11% (45/412), respectively, of the laboratories. The discordance rates were 2.2% for RPR test and 3.6% for VDRL. For the treponemal tests, Treponema pallidum hemagglutination assay (TPHA) was used in 60-76% and Immunochromatography assay (ICA) in about 30% of the laboratories in 2006. A high discordance rate of over 10% was reported in both TPHA and in ICA methods, possibly due to a low titer (1:1 in VDRL) of EQA samples in 2005. Analysis of the accumulated data from year 2004 to 2006 showed that the discordance rates of TPHA, ICA, and FTA-ABS were 4.6%, 3.7%, and 2.7%, respectively. CONCLUSIONS: For syphilis tests, RPR test, TPHA, and ICA are mainly used in Korea. A high discordance rate is still reported in TPHA and ICA, especially when testing samples with a low titer. Further analysis of data and education of laboratory personnel are needed for the improvement of the EQA program.
Enzyme-Linked Immunosorbent Assay ; False Positive Reactions ; Fluorescent Treponemal Antibody-Absorption Test ; Humans ; Korea ; Quality Control ; Reagent Kits, Diagnostic ; Syphilis/*diagnosis ; Syphilis Serodiagnosis/methods/*standards ; Treponema Immobilization Test

Leptospirosis, a zoonotic disease that is caused by many serovars which are more than 200 in the world, is an emerging worldwide disease. Accurate and rapid diagnostic tests for leptospirosis are a critical step to diagnose the disease. There are some commercial kits available for diagnosis of leptospirosis, but the obscurity of a species- or genus-specific antigen of pathogenic Leptospira interrogans causes the reduced sensitivity and specificity. In this study, the polysaccharide derived from lipopolysaccharide (LPS) of nonpathogenic Leptospira biflexa serovar patoc was prepared, and the antigenicity was confirmed by immunoblot and enzyme linked immunosorbent assay (ELISA). The performance of the rapid diagnostic test (RDT) kit using the polysaccharide as a diagnostic antigen was evaluated in Korea, Bulgaria and Argentina. The sensitivity was 93.9%, 100%, and 81.0% and the specificity was 97.9%, 100%, and 95.4% in Korea (which is a rare region occurring with 2 serovars mostly), Bulgaria (epidemic region with 3 serovars chiefly) and Argentina (endemic region with 19 serovars mainly) respectively. These results indicate that this RDT is applicable for global diagnosis of leptospirosis. This rapid and effective diagnosis will be helpful for diagnosis and manage of leptospirosis to use and the polysaccharide of Leptospira may be called as genus specific antigen for diagnosis.
Antigens, Bacterial/*immunology ; Argentina ; Bulgaria ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Leptospira/isolation & purification/metabolism ; Leptospira interrogans/isolation & purification/metabolism ; Leptospirosis/*diagnosis/microbiology ; Male ; Polysaccharides/*immunology ; Reagent Kits, Diagnostic/*standards ; Republic of Korea ; Sensitivity and Specificity

Leptospirosis, a zoonotic disease that is caused by many serovars which are more than 200 in the world, is an emerging worldwide disease. Accurate and rapid diagnostic tests for leptospirosis are a critical step to diagnose the disease. There are some commercial kits available for diagnosis of leptospirosis, but the obscurity of a species- or genus-specific antigen of pathogenic Leptospira interrogans causes the reduced sensitivity and specificity. In this study, the polysaccharide derived from lipopolysaccharide (LPS) of nonpathogenic Leptospira biflexa serovar patoc was prepared, and the antigenicity was confirmed by immunoblot and enzyme linked immunosorbent assay (ELISA). The performance of the rapid diagnostic test (RDT) kit using the polysaccharide as a diagnostic antigen was evaluated in Korea, Bulgaria and Argentina. The sensitivity was 93.9%, 100%, and 81.0% and the specificity was 97.9%, 100%, and 95.4% in Korea (which is a rare region occurring with 2 serovars mostly), Bulgaria (epidemic region with 3 serovars chiefly) and Argentina (endemic region with 19 serovars mainly) respectively. These results indicate that this RDT is applicable for global diagnosis of leptospirosis. This rapid and effective diagnosis will be helpful for diagnosis and manage of leptospirosis to use and the polysaccharide of Leptospira may be called as genus specific antigen for diagnosis.
Antigens, Bacterial/*immunology ; Argentina ; Bulgaria ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Leptospira/isolation & purification/metabolism ; Leptospira interrogans/isolation & purification/metabolism ; Leptospirosis/*diagnosis/microbiology ; Male ; Polysaccharides/*immunology ; Reagent Kits, Diagnostic/*standards ; Republic of Korea ; Sensitivity and Specificity

PURPOSE: Japanese hop (Humulus spp.) and mugwort (Artemisia spp.) are notable causes of autumn pollinosis in East Asia. However, Japanese hop and mugwort pollen extracts, which are widely used for the diagnosis, have not been standardized. This study was performed to standardize Japanese hop and mugwort pollen extracts. MATERIALS AND METHODS: Allergen extracts were prepared in a standardized way using locally collected Humulus japonicus and purchased Artemisia vulgaris pollens. The immunoglobulin E (IgE) reactivities of prepared extracts were compared with commercial extracts via IgE immunoblotting and inhibition analyses. Intradermal skin tests were performed to determine the bioequivalent allergy unit (BAU). RESULTS: The IgE reactive components of the extracts via IgE immunoblotting were similar to those of commercial extracts. A 11-kDa allergen showed the strongest IgE reactivity in Japanese hop, as did a 28-kDa allergen in mugwort pollen extracts. Allergenic potencies of the investigatory Japanese hop and mugwort extracts were essentially indistinguishable from the commercial ones. Sums of erythema of 50 mm by the intradermal skin test (SigmaED50) were calculated to be 14.4th and 13.6th three-fold dilutions for Japanese hop and mugwort extracts, respectively. Therefore, the allergenic activity of the prepared extracts was 90827.4 BAU/mg for Japanese hop and 34412 BAU/mg for mugwort. CONCLUSION: We produced Japanese hop and mugwort pollen extracts using a standardized method. Standardized Japanese hop and mugwort pollen extracts will facilitate the production of improved diagnostic and immunotherapeutic reagents.
Allergens/*analysis/*immunology ; Antibody Specificity ; *Artemisia ; Bronchial Hyperreactivity/blood/immunology ; Cross Reactions ; Enzyme-Linked Immunosorbent Assay ; Humans ; Immunoblotting ; Immunoglobulin E/blood/*immunology ; Pollen/*chemistry/*immunology ; Reference Standards ; Republic of Korea ; Rhinitis, Allergic, Seasonal

OBJECTIVETo compare the 4 test kits on severe acute respiratory syndrome coronavirus (SARS-CoV) gene, antigen and antibody for early diagnose of SARS patients.

METHODSThree enzyme linked immunosorbent assay (ELISA) kits were used to detect SARS-CoV IgG, IgM and N protein and fluorescent polymerase chain reaction (F-PCR) kit was used to detect SARS-CoV RNA.

RESULTSIn 162 serum samples, 90.2% (55/61) became N protein positive in 1 - 5 days and 92.8% (13/14) became positive IgM and IgG in 15 - 18 days after the onset of disease, respectively. On 82 gorgling samples, the positive rates of F-PCR were 56.3% (14/24) in 1 - 5 days and 71.4% (10/14) in 6 - 9 days after the onset.

CONCLUSIONOther than F-PCR, N protein had good effect in the early detection on dubious patients which could lead to effective prevention and control of the epidemic.

Antibodies, Viral ; blood ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Male ; Nucleocapsid Proteins ; blood ; Polymerase Chain Reaction ; RNA, Viral ; blood ; Reagent Kits, Diagnostic ; standards ; SARS Virus ; isolation & purification ; Severe Acute Respiratory Syndrome ; diagnosis ; virology

OBJECTIVETo study the pharmacokinetics of a novel recombinant human granulocyte colony-stimulating factor (rhG-CSFa) in rats and to determine the proteolytic rates of rhG-CSFa in the whole blood and serum of rats in vitro.

METHODSThe pharmacokinetics of rhG-CSFa and conventional (wild type, WT) granulocyte colony-stimulating factor (G-CSF) were investigated in Sprague-Dawley rats which received either intravenous or subcutaneous injection of rhG-CSFa or WT G-CSF at three different doses (20, 50, or 100 microg/kg). The blood samples of rats were collected at multiple time points (from 0.08 to 12 h) and the concentrations of rhG-CSFa and WT G-CSF in serum were determined with a sandwich enzyme-linked immunosorbent assay (ELISA). For the study of proteolytic rates in vitro, the concentrations of rhG-CSFa or WT G-CSF were determined at 3-minute intervals after addition of the respective drug to rat's whole blood or serum.

RESULTSPharmacokinetic analysis of serum rhG-CSFa or WT G-CSF levels indicated that, at each dose tested, for either route of drug administration, the area under concentration-time curve values and the maximum serum concentration of rhG-CSFa were higher than those of WT G-CSF, and the serum half life of rhG-CSFa was longer than that of WT G-CSF. Subsequent in vitro whole blood and serum stability study showed that the rates of drug degradation in WT G-CSF were 1.8 folds and 1.5 folds higher than those in rhG-CSFa, respectively.

CONCLUSIONrhG-CSFa has better serum and whole blood stability in vitro and higher bioavailability in vivo as compared to WT G-CSF.

Animals ; Cell Line ; Enzyme-Linked Immunosorbent Assay ; methods ; standards ; Female ; Granulocyte Colony-Stimulating Factor ; blood ; genetics ; pharmacokinetics ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Rats ; Rats, Sprague-Dawley ; Recombinant Proteins ; Sensitivity and Specificity