Time-resolved immunofluorometric assay for the ovarian carcinoma-associated antigenic determinant CA 125 in serum.

1987 ◽  
Vol 33 (12) ◽  
pp. 2191-2194 ◽  
Author(s):  
O C Boerman ◽  
C M Thomas ◽  
M F Segers ◽  
P Kenemans ◽  
T Lövgren ◽  
...  
Keyword(s):  
Ovarian Carcinoma ◽  
Antigenic Determinant ◽  
Fluorescence Enhancement ◽  
Ca 125 ◽  
Serum Samples ◽  
Measuring Range ◽  
Time Resolved ◽  
Europium Chelate ◽  
Analytical Range ◽  
Automated Procedures

Abstract A time-resolved immunofluorometric assay (IFMA) is described for quantifying the ovarian carcinoma-associated antigenic determinant CA 125 in human serum. Monoclonal antibody to CA 125 is immobilized onto a microtiter well, and the same antibody labeled with a europium chelate is used as a tracer. After the immunoreaction the bound portion of the labeled antibody is quantified by dissociating the Eu3+ in a fluorescence-enhancement solution and measuring its fluorescence with a time-resolved fluorometer. The detection limit of the IFMA is 1.5 arb. units/mL, being about the same as that of a commercially available immunoradiometric assay (IRMA) for CA 125 (1.4 arb. units/mL). The analytical range of the IFMA extends to 2000 arb. units/mL, whereas the range of the IRMA is 500 arb. units/mL. For 29 serum samples from ovarian-cancer patients measured simultaneously in the IFMA and IRMA, orthogonal regression analysis gave the equation CA 125 (IFMA) = 0.9937 CA 125 (IRMA) - 1.211 arb. units/mL (Syx = 6.8681, r = 0.9932). Apparently, the IFMA for CA 125 is a convenient alternative to the IRMA for CA 125 because of short counting times, the use of nonradioactive, stable reagents, and the much-extended measuring range. Additionally, the microtiter format should lend itself to more fully automated procedures in laboratories doing many such analyses.

scholarly journals Streptavidin-Polyvinylamine Conjugates Labeled with a Europium Chelate: Applications in Immunoassay, Immunohistochemistry, and Microarrays

2000 ◽  
Vol 46 (9) ◽  
pp. 1450-1455 ◽  
Author(s):  
Andreas Scorilas ◽  
Anders Bjartell ◽  
Hans Lilja ◽  
Christina Moller ◽  
Eleftherios P. Diamandis
Keyword(s):  
Solid Phase ◽  
Specific Antigen ◽  
Microarray Experiment ◽  
Immunohistochemical Localization ◽  
Macromolecular Complex ◽  
Serum Samples ◽  
Time Resolved ◽  
Europium Chelate ◽  
Highly Correlated ◽  
Detection Reagent

Abstract Background: The favorable properties of lanthanide chelates compared with conventional fluorescent probes have attracted considerable interest. A Eu3+ chelator, 4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA), has been synthesized previously. Methods: We here describe immunoassay, immunohistochemistry, and microarray applications of a new streptavidin-based universal polyvinylamine (PVA) detection reagent that is multiply labeled with the europium chelate of BCPDA. Solid-phase time-resolved immunofluorometric assays for biotinylated mouse IgG and prostate-specific antigen (PSA) were developed using the new conjugate as a detection reagent. The new conjugate was also used for the immunohistochemical localization of PSA expression in paraffin-embedded prostatic tissues. A model microarray with spotted biotinylated antibody as target was also performed. Results: Approximately 50–100 BCPDA moieties were covalently bound to PVA, which was then linked to streptavidin via biotin interaction. The macromolecular complex successfully recognized and bound biotinylated detection reagents, e.g., antibodies. The new reagent enabled measurement of solid phase-immobilized biotinylated mouse IgG with a detection limit of ∼1 pg/assay and demonstrated excellent linearity. In an ELISA-type sandwich PSA assay that included two PSA monoclonal antibodies using the new conjugate as detection reagent, we detected 0.001 μg/L PSA (∼100 fg or ∼3 amol/assay). Serum samples analyzed for PSA by this method and a commercial assay gave highly correlated results. The new reagent enabled excellent immunohistochemical localization of PSA expression in prostate tissues. Using the new reagent in a model microarray experiment with biotinylated mouse IgG as target, we demonstrated excellent spatial resolution of 5- to 10-nL microspots. Conclusions: The new detection reagent may find important applications in biotechnology.

Two-site time-resolved immunofluorometric assay of human insulin.

1986 ◽  
Vol 32 (4) ◽  
pp. 637-640 ◽  
Author(s):  
E Toivonen ◽  
I Hemmilä ◽  
J Marniemi ◽  
P N Jørgensen ◽  
J Zeuthen ◽  
...  
Keyword(s):  
Human Insulin ◽  
Fluorescence Enhancement ◽  
Microtiter Plate ◽  
The Other ◽  
Plasma Samples ◽  
Serum Samples ◽  
Time Resolved ◽  
Standard Curve ◽  
Sandwich Type ◽  
Plate Strip

Abstract We describe a two-site "sandwich"-type time-resolved immunofluorometric assay for human insulin, based on use of two monoclonal antibodies with different specificities. The first antibody is immobilized on the surface of microtiter plate strip wells, the other is labeled with Eu3+. Serum samples can be assayed with one incubation step; two incubation steps are required when plasma samples are assayed. After the immunoreactions are complete, the bound fraction of Eu3+-label is quantified by dissociating it in a fluorescence-enhancement solution and measuring its fluorescence with a fluorometer with time-resolution. The sensitivity of the assay is 0.24 micro-int. units/mL. The standard curve is linear from 0.24 to 2400 micro-int. units/mL.

Immunofluorometric demonstration and quantification of placental protein 5 in the absence of pregnancy.

1988 ◽  
Vol 34 (8) ◽  
pp. 1591-1593 ◽  
Author(s):  
R Bützow ◽  
H Alfthan ◽  
U H Stenman ◽  
A M Suikkari ◽  
H Bohn ◽  
...  
Keyword(s):  
Detection Limit ◽  
Solid Phase ◽  
Polyclonal Antiserum ◽  
Serum Samples ◽  
Measuring Range ◽  
Time Resolved ◽  
Healthy Men ◽  
Placental Protein ◽  
The Mean ◽  
Mean Concentration

Abstract This time-resolved immunofluorometric assay (IFMA) developed for measurement of placental protein 5 (PP5) involves two antibodies: a monoclonal anti-PP5 antibody attached to a solid phase and an europium(III) chelate-labeled polyclonal anti-PP5 antibody as a tracer. The measuring range is 0.05-100 micrograms/L and the detection limit is 20 times lower than that of a PP5 radioimmunoassay (RIA) performed with the same polyclonal antiserum. By IFMA, PP5 could be detected and quantified in all plasma and serum samples of nonpregnant and pregnant individuals, whereas PP5 was undetectable by RIA in serum of healthy men and nonpregnant women. The mean concentration of PP5 in sera from men was 0.43 micrograms/L (SD 0.13, range 0.19-0.75, n = 47) and in sera from nonpregnant women 0.49 micrograms/L (SD 0.19, range 0.20-0.90, n = 41). PP5 concentrations in serum showed no systematic variation during the menstrual cycle. In serum samples from 60 pregnant women the results obtained by IFMA and RIA correlated well (r = 0.97).

Time-resolved immunofluorometry and other frequently used immunoassay types for follicle-stimulating hormone compared by using identical monoclonal antibodies

1993 ◽  
Vol 39 (7) ◽  
pp. 1435-1439 ◽  
Author(s):  
S Madersbacher ◽  
T Shu-Chen ◽  
S Schwarz ◽  
S Dirnhofer ◽  
G Wick ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Signal To Noise Ratio ◽  
Follicle Stimulating Hormone ◽  
Serum Samples ◽  
Measuring Range ◽  
Time Resolved ◽  
Assay Performance ◽  
Reference Preparation ◽  
Stimulating Hormone

Abstract The influence of assay design and quantification system on assay performance was investigated by developing, optimizing, and comparing a time-resolved immunofluorometric assay (IFMA), an immunoenzymometric assay (IEMA), an immunoradiometric assay (IRMA), and a competitive radioimmunoassay (RIA), all performed with the same monoclonal antibodies (MCA) directed against human follicle-stimulating hormone (hFSH). The lowest detection limit (2 ng/L for hFSH-I-3, corresponding to 2.5 mIU of 1st International Reference Preparation of hFSH 78/549 per liter), the widest measuring range (2-160,000 ng/L), and the greatest signal-to-noise ratio (13,000:1 at 160,000 ng/L) were obtained in the IFMA. For analysis of serum samples from 101 male (ages 2-91 years) and 99 female (ages 2-90 years) individuals at a single dilution, 100% of samples were within the measuring range of the IFMA, whereas only 87%, 55%, 32%, and 8% of the sera were for the IRMA, the IEMA evaluated with double-wavelength measurement, the conventional IEMA, and the competitive MCA-based RIA, respectively. These studies demonstrate clear advantages of the IFMA in sensitivity and assay range, which allows reliable and cost- and time-effective determination of hFSH in individuals from infancy to senescence.

Comparison of five immunoassay procedures for the ovarian carcinoma-associated antigenic determinant CA 125 in serum

1992 ◽  
Vol 47 (3) ◽  
pp. 245-252 ◽  
Author(s):  
Kees A. Yedema ◽  
Chris M.G. Thomas ◽  
Martin F.G. Segers ◽  
Wim H. Doesburg ◽  
Peter Kenemans
Keyword(s):  
Ovarian Carcinoma ◽  
Antigenic Determinant ◽  
Ca 125

Double monoclonal time-resolved immunofluorometric assay of beta 2-microglobulin in serum

1990 ◽  
Vol 36 (11) ◽  
pp. 1961-1964 ◽  
Author(s):  
A Tienhaara ◽  
J U Eskola ◽  
V Näntö
Keyword(s):  
Serum Samples ◽  
Time Resolved ◽  
Assay Procedure ◽  
Analytical Recovery ◽  
Sandwich Type ◽  
Capture Antibody ◽  
Polyclonal Antisera ◽  
Europium Chelate ◽  
Beta 2 ◽  
Beta 2 Microglobulin

Abstract Previously reported immunochemical assays of beta 2-microglobulin (beta 2m) have usually been based on polyclonal antisera. We have developed a "sandwich"-type time-resolved immunofluorometric assay (TR-IFMA) for beta 2m in serum, based on two monoclonal antibodies against human beta 2m. Microtiter wells are coated with the capture antibody, and the tracer antibody is labeled with a europium chelate. In a simple and fast assay procedure, prediluted serum samples are incubated with the tracer for 1 h in the microtiter wells, after which the wells are washed and the fluorescence of Eu is measured. The mean analytical recovery was 101.8% and results by TR-IFMA showed a good linear correlation with those by an established radioimmunoassay. The analytical range of TR-IFMA is large and well suited for clinical purposes.

TO DETERMINE THE ROLE OF CA125 IN THE DIAGNOSIS OF ACUTE APPENDICITIS

2020 ◽  
Vol 4 (7) ◽  
Author(s):  
Vinod Kumar ◽  
Bhupen Songra ◽  
Richa Jain ◽  
Deeksha Mehta
Keyword(s):  
Acute Appendicitis ◽  
Clinical Diagnosis ◽  
Tertiary Care Hospital ◽  
Tertiary Care ◽  
Ca 125 ◽  
Care Hospital ◽  
Serum Samples ◽  
Roc Curve Analysis ◽  
Surgery Department

Background: the present study was under taken to determine the role of CA-125 in the diagnosis of acute appendicitis (AA), to prevent its complications and also in preventing negative appendicectomies in tertiary care hospital. Methods: The study was conducted at a tertiary care and research center between 01/03/2018 to 30/06/2019. Patients admitted to the surgery department with diagnosis of AA were considered for the study. After informed consent, a, standardized history was obtained as a case Performa. Serum samples from all the cases with clinical diagnosis of AA were obtained and stored. Only the cases with histopathologically approved AA were included in the study. Cases operated for clinical diagnosis of AA, but not histopathologically proven AA was not included in the study. CA125 levels in cases with definitive diagnosis of AA were measured. Results: In present study, ROC curve analysis revealed the sensitivity of 87.27 % and specificity of 90.91 % when the CA 125 cut-off value of > 16.8 was taken to diagnose acute appendicitis. AUC was 0.911 with a standard error of 0.0292. Conclusion: In this study we have observed that CA125 showed a positive correlation with acute appendicitis, that was statistically not significant (P>0.05). We didn’t evaluate the correlation with the disease severity. We consider that CA125 can be used as a marker in acute appendicitis cases although further research is still needed. Keywords: CA125, Acute Appendicitis, Surgery.

scholarly journals Automated Assay of a Four-Protein Biomarker Panel for Improved Detection of Ovarian Cancer

Cancers ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 325
Author(s):  
Christopher Walker ◽  
Tuan-Minh Nguyen ◽  
Shlomit Jessel ◽  
Ayesha B. Alvero ◽  
Dan-Arin Silasi ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Early Detection ◽  
Predictive Value ◽  
Early Stage ◽  
Ca 125 ◽  
Healthy Controls ◽  
Classification Models ◽  
Serum Samples ◽  
Protein Biomarker ◽  
Biomarker Panel

Background: Mortality from ovarian cancer remains high due to the lack of methods for early detection. The difficulty lies in the low prevalence of the disease necessitating a significantly high specificity and positive-predictive value (PPV) to avoid unneeded and invasive intervention. Currently, cancer antigen- 125 (CA-125) is the most commonly used biomarker for the early detection of ovarian cancer. In this study we determine the value of combining macrophage migration inhibitory factor (MIF), osteopontin (OPN), and prolactin (PROL) with CA-125 in the detection of ovarian cancer serum samples from healthy controls. Materials and Methods: A total of 432 serum samples were included in this study. 153 samples were from ovarian cancer patients and 279 samples were from age-matched healthy controls. The four proteins were quantified using a fully automated, multi-analyte immunoassay. The serum samples were divided into training and testing datasets and analyzed using four classification models to calculate accuracy, sensitivity, specificity, PPV, negative predictive value (NPV), and area under the receiver operating characteristic curve (AUC). Results: The four-protein biomarker panel yielded an average accuracy of 91% compared to 85% using CA-125 alone across four classification models (p = 3.224 × 10−9). Further, in our cohort, the four-protein biomarker panel demonstrated a higher sensitivity (median of 76%), specificity (median of 98%), PPV (median of 91.5%), and NPV (median of 92%), compared to CA-125 alone. The performance of the four-protein biomarker remained better than CA-125 alone even in experiments comparing early stage (Stage I and Stage II) ovarian cancer to healthy controls. Conclusions: Combining MIF, OPN, PROL, and CA-125 can better differentiate ovarian cancer from healthy controls compared to CA-125 alone.

scholarly journals Role of human epididymis protein 4 for detection of ovarian carcinoma in adnexal masses: A pilot study

2016 ◽  
Author(s):  
Nidhi Bansal ◽  
A. Suneja ◽  
K. Guleria ◽  
N. B. Vaid ◽  
K. Mishra ◽  
...  
Keyword(s):  
Diagnostic Accuracy ◽  
Cross Sectional Study ◽  
Ca 125 ◽  
Serum Samples ◽  
Cross Sectional ◽  
Adnexal Masses ◽  
Good Discriminator ◽  
Risk Of Malignancy ◽  
Risk Of Malignancy Index ◽  
Benign Tumours

Introduction: HE4 is a novel tumour biomarker used for early diagnosis of ovarian cancer. This study evaluated the diagnostic accuracy of HE4 alone and in combination with CA125, risk of malignancy index (RMI), risk of malignancy algorithm (ROMA). Methods: It was a cross sectional study conducted recruiting 88 women with adnexal masses who were planned for surgery. After baseline work up and ultrasound examination, serum samples were collected for estimation of CA 125 and HE4 levels. Serum HE4 levels were estimated using ELISA kit. RMI and ROMA score were calculated and diagnostic accuracy of HE4, CA 125, RMI, ROMA and their combination were compared. Cut off for HE4 and ROMA score were calculated using ROC curve. Results: Of 88 subjects, 66 were analyzed with 19 malignant (including 5 LMP) and 47 benign cases. The median value of HE4 among malignant cases was found to be significantly higher than among the benign cases. PPV and NPV of HE4 at a cut off 130.8 pMol/ml was 85.7% and 77.9% respectively. Highest PPV (88.9%) with acceptable NPV (80.7%) was found with ROMA followed by HE4 (PPV 85.7%; NPV 77.97%), RMI (PPV 76.92%; NPV 83%) and CA125 (PPV 52%; NPV 80.85%). Conclusion: HE4 levels were lower in Indian population both in malignant and benign tumours as compared to other studies. HE4 is a good discriminator and gives best accuracy when it is combined with CA125 in a logistic algorithm, ROMA.

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