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PubMed2003

The commercially available ELISA for pancreatic elastase 1 based on polyclonal antibodies does measure an as yet unknown antigen different from purified elastase 1. Binding studies and clinical use in patients with exocrine pancreatic insufficiency.

The measurement of fecal elastase 1 concentrations by means of an ELISA based on monoclonal antibodies (mABs) highly specific for human elastase 1 (ELISA 1) has become an accepted indirect test of the exocrine pancreatic function during the last years. Its use has been demonstrated in many clinical studies including comparison with direct function tests and ERCP morphology. Recently, a new ELISA, also named "elastase 1" based on polyclonal antibodies (pABs; ELISA 2) became available. In the present investigation we performed binding studies with purified elastase 1 as well as studies on patients with exocrine insufficiency with both ELISAs. Surprisingly, the pABs on the solid phase (catcher antibodies) of ELISA 2 did not bind purified elastase 1. These antibodies seem to react with an as yet unknown antigen associated with elastase 1. Measurement of samples from patients suspected to suffer from exocrine insufficiency showed a weak correlation of both assays but higher levels in ELISA 2, resulting in false normal results even in some patients with pancreatic steatorrhea. Since the reference range used in both assays has been established using ELISA 1, ELISA 2 must be re-evaluated in comparison to direct function tests. ELISA 2 should be renamed, since obviously it does react with an antigen (antigens) different from elastase 1.

Chronic Disease↗
PubMed1990

Malaria sporozoite detection by dissection and ELISA to assess infectivity of afrotropical Anopheles (Diptera: Culicidae).

Malaria infection rates determined by dissection and Plasmodium falciparum enzyme-linked immunosorbent assay (ELISA) were compared for 26,935 Anopheles gambiae Giles sensu lato and 17,739 Anopheles funestus Giles collected during 20 mo in western Kenya. ELISA infection rates were about 43% higher than dissection sporozoite rates. In dissection-negative Anopheles, circumsporozoite (CS) protein was detected by ELISA in 5.2% of 10,017 salivary gland samples and in 12.2% of 237 thorax samples. The accuracy of dissection and ELISA techniques was compared by the following tests on a group of 352 field-collected Anopheles (held 10 d to ensure sporogonic development): salivary gland dissection, examination of Giemsa-stained dissection slides, ELISA tests on salivary gland and thorax body parts, and microscopic techniques for determining sporozoite loads. Respective infection rates were 9.9%, 10.8%, and 15.6% for dissection, stained slides, and ELISA. Sporozoite loads were associated significantly with ELISA absorbance values (r = 0.76). Compared with Giemsa-stained dissection slide results, the sensitivity of sporozoite detection was 92.1% for dissection compared with 78.9% for ELISA; specificity was 100.0% for dissection versus 92.0% for ELISA. Immunological detection of CS protein in head-thorax samples of Afrotropical vectors overestimated the proportion of infective Anopheles because the comparison of techniques indicated that 45.4% of the ELISA positive Anopheles did not contain salivary gland sporozoites.

Animals↗
PubMed2002

Comparison of an ELISA and an LC/MS/MS method for measuring tacrolimus concentrations and making dosage decisions in transplant recipients.

This study compared an enzyme-linked immunosorbent assay (ELISA) to a liquid chromatography-tandem mass spectrometry (LC/MS/MS) technique for measurement of tacrolimus concentrations in adult kidney and liver transplant recipients, and investigated how assay choice influenced pharmacokinetic parameter estimates and drug dosage decisions. Tacrolimus concentrations measured by both ELISA and LC/MS/MS from 29 kidney (n = 98 samples) and 27 liver (n = 97 samples) transplant recipients were used to evaluate the performance of these methods in the clinical setting. Tacrolimus concentrations measured by the two techniques were compared via regression analysis. Population pharmacokinetic models were developed independently using ELISA and LC/MS/MS data from 76 kidney recipients. Derived kinetic parameters were used to formulate "typical dosing" regimens for concentration targeting. Dosage recommendations for the two assays were compared. The relation between LC/MS/MS and ELISA measurements was best described by the regression equation ELISA = 1.02. (LC/MS/MS) + 0.14 in kidney recipients, and ELISA = 1.12. (LC/MS/MS) - 0.87 in liver recipients. ELISA displayed less accuracy than LC/MS/MS at lower tacrolimus concentrations. Population pharmacokinetic models based on ELISA and LC/MS/MS data were similar with residual random errors of 4.1 ng/mL and 3.7 ng/mL, respectively. Assay choice gave rise to dosage prediction differences ranging from 0% to 30%. ELISA measurements of tacrolimus are not automatically interchangeable with LC/MS/MS values. Assay differences were greatest in adult liver recipients, probably reflecting periods of liver dysfunction and impaired biliary secretion of metabolites. While the majority of data collected in this study suggested assay differences in adult kidney recipients were minimal, findings of ELISA dosage underpredictions of up to 25% in the long term must be investigated further.

Chromatography, Liquid↗
PubMed1995

Soluble HLA antigens and ELISA--a new technology for crossmatch testing.

A soluble HLA ELISA for the detection of donor specific anti-HLA class I IgG antibodies was developed and compared with microlymphocytotoxicity. Donor sHLA was prepared from donor blood or purified blood lymphocytes and captured onto monoclonal antibody coated ELISA plates. After incubation of captured HLA with test serum, bound IgG antibodies were detected using a peroxidase-conjugated anti-human IgG antibody. Serum samples from patients on waiting lists to receive kidney transplants were tested by lymphocytotoxicity (AHG protocol) and/or sHLA ELISA in four different laboratories using HLA preparations from eight organ donors. Concordant crossmatch results were obtained for 854 (99%) of 864 ELISA crossmatches. In contrast, concordant results were obtained for 234 (91%) of 256 lymphocytotoxicity crossmatches. Interlaboratory reproducibility of ELISA results was 99%. In contrast, interlaboratory reproducibility of lymphocytotoxicity assay results was 78%. Endpoint titrations of serum specimens containing anti-HLA antibodies demonstrated equivalent sensitivity of ELISA and AHG lymphocytotoxicity crossmatch and similar sensitivity of ELISA and flow cytometry crossmatch. Specimens tested positive by lymphocytotoxicity without DTT treatment but negative with DTT treatment were tested negative by ELISA. Comparison of lymphocytotoxicity and ELISA crossmatch results showed an agreement of 94%. This demonstrates that detection of anti-donor HLA class I antibodies by ELISA is a reliable alternative to microlymphocytotoxicity testing.

Enzyme-Linked Immunosorbent Assay↗
PubMed2000

Detection of HLA class I and II antibodies by ELISA and complement-dependent cytotoxicity before and after transplantation.

BACKGROUND: Anti-class I IgG can be detected by complement-dependent cytotoxicity (CDC) and by ELISA. We compared ELISA and CDC for both class I and class II antibodies on method agreement and relation to rejection-free and graft survival. METHODS: Peak, current, and posttransplant sera (n=429) of 143 renal allograft patients were tested by National Institutes of Health technique (NIHT), two-color fluorescence (TCF), and ELISA. Method agreement was assessed by intraclass correlation coefficient (ICC). Rejection and graft survival were analyzed by uni- and multivariate techniques. The screening results for each serum were compared, as was the change in result of current to posttransplant serum. RESULTS: The ICC of ELISA and NIHT was insufficient; it was lower for TCF than NIHT. Graft survival was not related to the result of any assay. Rejection-free survival was related to ELISA and NIHT in current and posttransplant serum. With the NIHT, the change in percent panel-reactive antibody (%PRA) correlated better with rejection than it did with ELISA. The combined antibody status of current and posttransplant serum was a risk factor for rejection in all assays, and for TCF also in multivariate analysis. The rejection rate was higher if the posttransplant serum was ELISA-negative/CDC-positive, rather than ELISA-positive/CDC-negative. For ELISA, class I specificities (and not %PRA) in peak and current sera were related to rejection, even if the antibodies were not donor-directed. In the case of the National Institutes of Health technique (NIHT), %PRA and not specificity was related to rejection. Class II antibodies were never related to rejection. CONCLUSIONS: ELISA and NEIT are complementary screening techniques in this patient population. They are of equal predictive value for rejection. The optimal strategy in combining these techniques must be determined.

Adolescent↗
PubMed1988

Evaluation of a monoclonal antibody blocking ELISA for the detection of group-specific antibodies to bluetongue virus in experimental and field sera.

In order to overcome serological cross-reactions among orbivirus serogroups, which can hinder the accurate diagnosis of bluetongue virus (BTV) infection of livestock, a blocking ELISA (B-ELISA) incorporating a monoclonal antibody (20E9B7G2) with specificity for the BTV serogroup was developed. Experimental antisera raised to South African BTV serotypes 1 to 19 were tested in the B-ELISA and all blocked the binding of 20E9B7G2 to BTV antigen. The sensitivity and specificity of the assay was evaluated with a range of experimental and field sera and compared to a sensitive indirect ELISA (I-ELISA) for the detection of BTV-specific antibodies. The specificity of the B-ELISA was absolute for antibodies to BTV, showing no cross-reaction with experimental antisera to serotypes of the closely related orbivirus causing epizootic haemorrhagic disease of deer. The sensitivity of the B-ELISA exceeded that of the I-ELISA. In particular, the B-ELISA detected a BTV-specific antibody response much earlier after infection that the I-ELISA, while still exhibiting full sensitivity to BTV antibody titres several months after infection.

Animals↗
PubMed1992

Early antihepatitis C virus response with second-generation C200/C22 ELISA.

Detection of early antibody to hepatitis C virus (HCV) by a new second-generation C200/C22 anti-HCV enzyme-linked immunosorbent assay (ELISA) and a four-antigen recombinant immunoblot assay (4-RIBA) was compared with the first-generation anti-HCV C100 ELISA using sequential serum samples of 9 recipients who were infected with HCV, as detected by polymerase chain reaction after transfusion of blood products. Within 26 weeks after transfusion, 9/9 (100%) recipients seroconverted with C200/22 ELISA, and 6/9 (67%) seroconverted with C100 ELISA. Compared with C100 ELISA, C200/C22 ELISA seroconversion occurred simultaneously in 3 cases, 5-6 weeks earlier in 3 other cases, and 20 weeks earlier in 1 case. Seven of 9 (78%) recipients became positive, and 2/9 (22%) became indeterminate with 4-RIBA. In 8 cases with clinical posttransfusion hepatitis non-A, non-B (PTH-NANB), anti-HCV C200/C22 ELISA seroconversion occurred 2-17 (mean 6) weeks after the onset of hepatitis. In 6 cases of PTH-NANB, anti-HCV C100 ELISA seroconversion occurred 2-26 (mean 9) weeks after the onset of hepatitis. It is concluded that the second-generation C200/C22 ELISA is more sensitive than the C100 ELISA for the detection of antibody during early HCV infection. Indeterminate 4-RIBA results are found in the early phase of HCV infection.

Enzyme-Linked Immunosorbent Assay↗
PubMed1995

Improved detection of anti-HCV in post-transfusion hepatitis by a third-generation ELISA.

The sensitivity of ORTHO HCV 3.0 ELISA Test System (ELISA 3) for the detection of anti-HCV was compared with the second-generation ELISA, OR-THO HCV 2.0 ELISA Test System (ELISA 2). ELISA 3 differs from ELISA 2 in that it incorporates the HCV recombinant antigen NS5, in addition to recombinant antigens derived from the NS3, NS4 and core regions of the HCV genome. Specimens tested consisted of serial bleeds obtained from 21 individuals undergoing seroconversion following acquisition of post-transfusion HCV infection. ELISA 3 demonstrated significantly greater sensitivity than ELISA 2, detecting seroconversion earlier in 24% (5/21) of cases. Although one of these cases appeared to represent early seroconversion to NS5, most of the improved sensitivity of ELISA 3 appeared to derive from increased detectability of anti-c33c.

Enzyme-Linked Immunosorbent Assay↗
PubMed1994

An ELISA based on whole virus for the detection of antibodies to small-ruminant lentiviruses.

A new ELISA kit was developed, based on highly purified whole-virus antigen derived from the Swiss maedi-visna virus strain OLV. The sensitivity, specificity and accuracy of this assay were compared with that of an established ELISA based on recombinant GAG (group-specific antigens)-GST (glutathione S-transferase) fusion protein expressed in E. coli (GAG-GST ELISA). The whole-virus ELISA exhibits at least comparable specificity (99.3%) but higher sensitivity (98.6 versus 86.3%) and agreement with the 'true' status beyond chance in the detection of antiviral antibodies in serum from goats. Antibodies in milk samples are detected with higher specificity (98.9 versus 97.8%) but lower sensitivity (91.4 versus 98.2%) than in GAG-GST ELISA. The specificity of the new ELISA in the detection of antibodies in serum might be superior, since a set of 40 samples falsely rated positive in GAG-GST ELISA in routine diagnostic work was negative in the new ELISA. In both assays, milk samples can be tested instead of serum, although with slightly reduced sensitivity in the new ELISA. The major advantage of the new test kit is the low number of equivocal samples needing confirmation in a supplementary test. Results obtained with sheep sera indicate that the new ELISA kit is also suitable for the detection of antibodies to maedi-visna virus.

Animals↗
PubMed2003

Audit of the clinical usefulness of a rapid qualitative ELISA screen for antimyeloperoxidase and antiproteinase 3 antibodies in the assessment of patients with suspected vasculitis.

BACKGROUND: The need for urgent antineutrophil cytoplasmic antibody (ANCA) results when assessing patients with acute renal failure, pulmonary renal syndrome, or mononeuritis multiplex has led to the development of a rapid qualitative ELISA screening assay for antibodies to myeloperoxidase (MPO) and proteinase 3 (PR3). AIMS: To report the use of a rapid qualitative ELISA screen for PR3-ANCA and MPO-ANCA in a regional immunology laboratory and its correlation with standard indirect immunofluorescence (IIF) and quantitative ELISA for PR3-ANCA and MPO-ANCA. METHODS: Over 12 months, 103 samples requiring urgent ANCA testing were screened by a rapid qualitative ELISA and the results compared with IIF and quantitative ELISA assays for PR3-ANCA and MPO-ANCA. RESULTS: There was an excellent correlation between the rapid qualitative ELISA and standard ANCA IIF and a routine ELISA for MPO/PR3-ANCA, with sensitivities ranging from 82% to 100%. There were two false negatives, which gave weak to moderately positive values as determined by routine ELISA. However, the clinical relevance of these two cases is doubtful. CONCLUSIONS: The rapid ELISA for anti-MPO and anti-PR3 correlates well with quantitative ELISA and IIF ANCA, and urgent management decisions in patients with suspected small vessel vasculitis can be based with confidence on this test.

Antibodies, Antineutrophil Cytoplasmic↗
PubMed1995

Evaluation of an O-antigen ELISA for screening cattle herds for Salmonella typhimurium.

A total of 2585 serum samples from 62 dairy herds located in four different regions of Denmark were tested in an O-antigen (0:1,4,5,12)-based ELISA for the detection of antibodies against Salmonella typhimurium. Ten closed herds from an island with no reported occurrence of salmonellosis for several years, and 12 herds from a salmonella enzootic area which had had clinical outbreaks of S typhimurium were used to define a herd ELISA cut-off value. When herds with at least 5 per cent of the serum samples having an optical density of > 0.5 were considered ELISA-positive, all 10 herds from the salmonellosis-free island were ELISA-negative, and all but one of the 12 S typhimurium-infected herds were ELISA-positive, which resulted in a herd test sensitivity of 0.92 and herd test specificity of 1.0. Eleven of the 12 S typhimurium-infected herds were negative in a blocking ELISA based on a monoclonal antibody to the 0:9 antigen of the serogroup D salmonellas, indicating the possibility of rapid serogroup-specific screening of herds by means of these two tests. Ten other randomly selected herds with clinical outbreaks of S dublin were all, to a large extent, positive in the 0:1,4,5,12-ELISA, whereas a S dublin (0:1,9,12)-ELISA described previously appeared to be more serogroup D-specific. Thus, the 0:1,4,5,12-ELISA appears to be useful for detecting herd infections with S typhimurium, and positive reactions may be further discriminated by the serogroup D-specific ELISA.

Animals↗
PubMed2005

Evaluating concentration estimation errors in ELISA microarray experiments.

BACKGROUND: Enzyme-linked immunosorbent assay (ELISA) is a standard immunoassay to estimate a protein's concentration in a sample. Deploying ELISA in a microarray format permits simultaneous estimation of the concentrations of numerous proteins in a small sample. These estimates, however, are uncertain due to processing error and biological variability. Evaluating estimation error is critical to interpreting biological significance and improving the ELISA microarray process. Estimation error evaluation must be automated to realize a reliable high-throughput ELISA microarray system. In this paper, we present a statistical method based on propagation of error to evaluate concentration estimation errors in the ELISA microarray process. Although propagation of error is central to this method and the focus of this paper, it is most effective only when comparable data are available. Therefore, we briefly discuss the roles of experimental design, data screening, normalization, and statistical diagnostics when evaluating ELISA microarray concentration estimation errors. RESULTS: We use an ELISA microarray investigation of breast cancer biomarkers to illustrate the evaluation of concentration estimation errors. The illustration begins with a description of the design and resulting data, followed by a brief discussion of data screening and normalization. In our illustration, we fit a standard curve to the screened and normalized data, review the modeling diagnostics, and apply propagation of error. We summarize the results with a simple, three-panel diagnostic visualization featuring a scatterplot of the standard data with logistic standard curve and 95% confidence intervals, an annotated histogram of sample measurements, and a plot of the 95% concentration coefficient of variation, or relative error, as a function of concentration. CONCLUSIONS: This statistical method should be of value in the rapid evaluation and quality control of high-throughput ELISA microarray analyses. Applying propagation of error to a variety of ELISA microarray concentration estimation models is straightforward. Displaying the results in the three-panel layout succinctly summarizes both the standard and sample data while providing an informative critique of applicability of the fitted model, the uncertainty in concentration estimates, and the quality of both the experiment and the ELISA microarray process.

Algorithms↗
PubMed2006

Development of ELISA using recombinant antigens for specific detection of mouse parvovirus infection.

Nucleotide sequences of mouse parvovirus (MPV) isolate, named MPV/UT, and mouse minute virus (MMV) were analyzed and used for expressing recombinant proteins in E. coli. ELISA tests using recombinant major capsid protein (rVP2) and recombinant major non-structural protein (rNS1) as antigens were developed and their performance in serologic detection of rodent parvovirus infection was assessed. MPV-rVP2 and MMV-rVP2 ELISAs reacted specifically with anti-MPV and anti-MMV mouse sera, respectively. MMV-rNS1 antigen had a wide reaction range with antisera to rodent parvoviruses including MPV, MMV, Kilham rat virus (KRV) and H-1 virus. All mice oronasally infected with MPV were seropositive at 4 weeks post-infection in screening by ELISAs using MPV-rVP2 and MMV-rNS1 antigens, but were negative by conventional ELISA using whole MMV antigen. A contact transmission experiment revealed that transmission of MPV occurred up to 4 weeks post-infection, and all cage mates were seropositive in screening with MPV-rVP2 and MMV-rNS1 ELISAs. These results indicate that MPV-rVP2 and MMV-rVP2 are specific ELISA antigens which distinguish between MPV and MVM infection, while MMV-rNS1 antigen can be used in generic ELISA for a variety of rodent parvoviruses. The higher sensitivity of MPV-rVP2 ELISA than conventional ELISA for detecting seroconversion to MPV in oronasally infected mice as well as in cage mates suggests the usefulness of MPV-rVP2 ELISA in quarantine and microbiological monitoring of MPV infection in laboratory mice.

Animals↗
PubMed2004

[Inter-laboratory evaluation studies for establishment of notified ELISA methods for allergic substances (buckwheat)].

Inter-laboratory evaluation studies were conducted for the ELISA methods for allergic substances (buckwheat). Extracts of snack, bun and udon spiked with buckwheat standard protein at a level of 5-20 ng/mL as sample solutions were analyzed in replicate at 10 laboratories. Coefficients of variation (CVs) of the ELISA methods using a Buckwheat Protein ELISA Kit (Buckwheat kit) and a FASTKIT Buckwheat ELISA kit (Buckwheat ELISA kit) were mostly below 10%. Mean recoveries of the buckwheat standard protein from the food extracts were over 40% in the two ELISA methods. Repeatability relative standard deviations of buckwheat standard protein in three food extracts were in the ranges of 6.8-78.5% and 5.0-33.9% for the Buckwheat kit and the Buckwheat ELISA kit, respectively. Reproducibility relative standard deviations of buckwheat standard protein in three food extracts were 11.9-69.5% and 16.5-34.1% for the Buckwheat kit and the Buckwheat ELISA kit, respectively. The detection limits of both ELISA methods were 1 ng/mL in sample solutions. These results suggest that the notified ELISA methods are reliable and reproducible for the inspection of buckwheat protein levels in extracts of snack, bun and udon.

Allergens↗
PubMed2004

[Inter-laboratory evaluation studies for establishment of notified ELISA methods for allergic substances (peanuts)].

Inter-laboratory evaluation studies were conducted for ELISA methods for allergic substances (peanuts). Extracts of biscuit, sauce, chocolate and butter spiked with peanut standard protein at a level of 5-20 ng/mL as sample solutions were analyzed in replicate in 10 laboratories. Coefficients of variation (CVs) of the ELISA methods using a Peanut Protein ELISA Kit (Peanut kit) and a FASTKIT Peanut ELISA kit (Peanut ELISA kit) were mostly below 10%. Mean recoveries of the peanut standard protein from the food extracts were over 40% in the two ELISA methods. Repeatability relative standard deviations of peanut standard protein in four food extracts were in the ranges of 15.2-49.7% and 3.0-28.3% for the Peanut kit and the Peanut ELISA kit, respectively. Reproducibility relative standard deviations of peanut standard protein in four food extracts were 23.5-44.4%, 9.6-28.4% for the Peanut kit and the Peanut ELISA kit, respectively. The detection limits of both ELISA methods were 2-2.5 ng/mL in sample solutions. These results suggested that the notified ELISA methods are reliable and reproducible for the inspection of peanut protein levels in extracts of biscuit, sauce, chocolate and butter.

Allergens↗
PubMed1999

Use of ELISA to measure antinuclear antibodies in children with juvenile rheumatoid arthritis.

OBJECTIVE: To compare a series of commercial ELISA tests with an indirect immunofluorescent antibody (IFA) test for the detection of antinuclear antibodies (ANA) in children with juvenile rheumatoid arthritis (JRA). METHODS: Sera from 178 patients with JRA (88 pauciarticular, 68 polyarticular, 22 systemic) were compared with 26 healthy pediatric subjects. Twenty-one samples from patients with systemic lupus erythematosus (SLE) were also tested. All samples were analyzed by IFA and by 3 commercial ELISA methods. Concordance of ELISA results with IFA results (selected standard) were used as a measure of performance. Sensitivity and specificity were calculated for each test and likelihood ratios (LR) were established for IFA and ELISA in pauciarticular and polyarticular JRA sera. The increment in pretest probability was then obtained for each test as an additional measure of test performance. RESULTS: IFA rendered positive results on 18-77% of the JRA sera depending upon the subset, 100% of SLE sera, and 15% of normal patient sera. Using IFA as the standard, correspondence with positive results among patients with JRA ranged from 0 to 74% for the 3 ELISA tests, while it ranged from 5 to 73% in IFA negative sera. IFA tests showed intermediate range likelihood ratios (0.3, 0.5, 3.5, and 5) and increments in pretest probability ranging from 25 to 45%. While one of the ELISA tests attained 50% of increment in pretest probability for the positive test, it showed 0% increment as a negative test. The other 2 ELISA tests incremented the pretest probability from 0 to 25%. CONCLUSION: Our findings indicate that in JRA, the lack of correspondence with the historic standard IFA precludes the use of ELISA tests for detection of ANA. In addition, IFA out-performs ELISA by a substantial degree when "clinical utility" analysis of test performance is utilized. Detection of ANA in children with JRA should either continue to rely on IFA or be based on a different set of antigens if an ELISA format is chosen.

Antibodies, Antinuclear↗
PubMed1979

Quantification of total IgM and IgG and specific IgM and IgG to a thymus-independent (LPS) and a thymus-dependent (tetanus toxoid) antigen in the rat by enzyme-linked immunosorbent assay (ELISA).

Rat IgM and IgG was determined by mechanized "sandwich" enzyme-linked immunosorbent assay (ELISA) using peroxidase labeled anti-rat-IgM and -IgG. Linear ranges in standard curves of a reference rat serum had a slope similar to the slopes found with sera of 25 rats of various age. IgM and IgG measurements by ELISA in these sera correlated well with results obtained by single radial immuno-diffusion (SRID). In addition, the precision of the enzyme immunoassay was the same as obtained with the SRID. Compared with SRID, ELISA is less time consuming and the amount of antiserum used in the macro-ELISA is one order of magnitude lower; and again 10 times lower in the mechanized micro-ELISA that is currently being developed. In conclusion, the ELISA is a specific, reliable, sensitive, and economic method for routine measurement of rat serum IgG and IgM e.g. in toxicity studies. In the second part of this study, ELISA and the passive hemagglutination test were compared to determine the primary and secondary antibody response to E. coli lipopolysaccharide (LPS) and tetanus toxoid in rats. In the ELISA, the antigens were bound to the wells of polystyrene microplates. Tetanus toxoid was coated directly, LPS after complexing with methylated bovine serum albumin. After incubation with dilutions of the rat sera, the amount of antibody bound to the solid phase was quantified by means of peroxidase-labeled antiimmunoglobulin. The specificity of the enzyme immunoassay was tested by absorption of the sera with the respective antigens. ELISA proved to be more sensitive than the hemagglution reaction, except when titers were determined during the secondary response to tetanus toxoid. Besides its specificity and sensitivity, ELISA is a convenient method for measuring both IgM and IgG antibodies. Finally, evidence is presented that in the rat, the humoral immune response to LPS is a thymus-independent phenomenon. Thus, by using the antibody response to LPS and tetanus toxoid in function studies of the immune system of the rat, insight can be obtained in the thymus-independent and thymus-dependent humoral immune response.

Age Factors↗
PubMedDate not supplied

Comparison of ELISA level and immunohistochemical status for thymidine phosphorylase (TP) in invasive breast carcinoma.

BACKGROUND: The correlation between ELISA level and immunohistochemical status for thymidine phosphorylase (TP) in invasive breast carcinoma was examined. MATERIALS AND METHODS: Specimens were obtained from 84 patients with invasive breast carcinoma and both the ELISA level and immunohistochemical status for TP of breast carcinoma tissue were determined. The results of ELISA for the 84 cases were categorized into quartiles (E-scores 1 to 4) with every category including an equal number of patients. In addition, both the staining intensity of carcinoma cells and relative number of stained stromal cells identified by immunohistochemistry were classified into 4 degrees (I-scores 1 to 4). We also divided the patients into two groups: a negative group (I-scores 1 and 2) and a positive group (I-scores 3 and 4). Furthermore, sums of scores for carcinoma cells and stromal cells (S-scores 2 to 8) were divided into two groups, a low group (S-scores 2 to 5) and a high group (S-scores 6 to 8). The correlation between the ELISA level and immunohistochemical status for TP was evaluated and the means of the ELISA level of each group were compared. RESULTS: Scores of ELISA (E-scores) were significantly positively-correlated with scores for immunohistochemical status (I-scores) of only carcinoma cells (lambda=0.158, p<0.05). The means of TP levels determined by ELISA were significantly higher in the carcinoma cell-positive group (p<0.01) and summed-score high group (p<0.0002). CONCLUSION: A positive correlation between the results of the ELISA and immunohistochemical status for TP was found only for carcinoma cells and comparison of means of the ELISA level indicated that they reflected total immunohistochemical TP status of carcinoma cells and stromal cells. Immunohistochemical examinations should be performed to clarify the in situ localization of TP in carcinoma tissue and the results obtained from the immunohistochemistry, as well as the ELISA of TP, may be useful in selection of patients for doxifluridine and capecitabine therapy.

Adult↗