scholarly journals A robust enzyme-linked immunosorbent assay to measure serum ramucirumab concentrations

Bioanalysis ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 565-574
Author(s):  
Céline Desvignes ◽  
David Ternant ◽  
Thierry Lecomte ◽  
Astrid Lièvre ◽  
Marc Ohresser ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Monoclonal Antibody ◽  
Cross Reactivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Pharmacokinetic Studies ◽  
Serum Concentrations ◽  
Response Relationship ◽  
Binding Assays ◽  
High Quality ◽  
Calibration Standards

Aim: Ramucirumab, an anti-VEGFR2 monoclonal antibody, has been approved for the treatment of metastatic gastric and colorectal cancer. An assay measuring ramucirumab serum concentrations was needed to investigate its pharmacokinetics and concentration–response relationship. Results: An ELISA was developed and validated according to the international guidelines for ligand-binding assays. Ramucirumab calibration standards ranged from 0.125 to 40 mg/l. Low, middle and high quality controls were spiked at 0.2, 4 and 8 mg/l, respectively. The limits of quantification were established to be 0.125 and 10 mg/l for LLOQ and ULOQ, respectively. No cross-reactivity with anti-VEGF or anti-EGFR was detected. Conclusion: This in-house-developed ELISA is sensitive, accurate, reproducible and suitable for pharmacokinetic studies of ramucirumab.

Generation of Human Monoclonal Anti-idiotypic Antibodies with Specificity to the Murine Monoclonal Anti-CA 125 Antibody B43.13

1996 ◽  
Vol 11 (4) ◽  
pp. 211-215
Author(s):  
J.B. Oltrogge ◽  
B. Donnerstag ◽  
R.P. Baum ◽  
A.A. Noujaim ◽  
L. Träger
Keyword(s):  
Ovarian Cancer ◽  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Peripheral Blood Lymphocytes ◽  
Cross Reactivity ◽  
Tumor Burden ◽  
Ca 125 ◽  
Enzyme Linked Immunosorbent Assay ◽  
Human Monoclonal Antibodies ◽  
Ebv Transformation

Two human monoclonal antibodies, HID-7E7 and ROB-6F2, were produced by EBV transformation of peripheral blood lymphocytes (PBL). PBL were obtained from a patient with ovarian cancer who had been exposed several times to a Tc-99m labeled murine monoclonal anti-CA 125 antibody (B43.13, Biomira, Edmonton) for immunoscintigraphy. The HID-7E7 and ROB-6F2 producing B-cells were cloned with a limiting dilution technique and have shown stable immunoglobulin secretion within a period of three years. The human monoclonal antibodies HID-7E7 and ROB-6F2 are of the IgG isotype, and bind with significant affinity to the murine monoclonal antibody B43.13, which was used for immunoscintigraphy. Binding affinity of ROB-6F2 to other murine antibodies could not be detected. Cross reactivity of HID-7E7 to a murine anti-CEA monoclonal antibody was observed. In order to verify the anti-idiotypic character of the generated human antibodies, the ability of HID-7E7 and ROB-6F2, respectively, to inhibit the formation of the CA125/B43.13 complex is demonstrated via an enzyme-linked immunosorbent assay. These human anti-idiotypic antibodies are possible candidates for immunotherapy of ovarian cancer in patients with a small tumor burden following surgery and/or chemotherapy.

scholarly journals Two-site immunofluorometric assay of intact salmon calcitonin with improved sensitivity

1997 ◽  
Vol 43 (1) ◽  
pp. 71-75 ◽  
Author(s):  
Haiqin Rong ◽  
Leonard J Deftos ◽  
Hong Ji ◽  
Elisabet Bucht
Keyword(s):  
Monoclonal Antibody ◽  
Detection Limit ◽  
Polyclonal Antibody ◽  
Salmon Calcitonin ◽  
Polyclonal Antibodies ◽  
Pharmacokinetic Studies ◽  
Human Calcitonin ◽  
Plasma Samples ◽  
Serum Concentrations ◽  
Capture Antibody

Abstract We recently developed a two-site immunofluorometric assay (IFMA) of salmon calcitonin (SCT) by DELFIA (dissociation enhancement lanthanide fluoroimmunoassay) technique using the same polyclonal antibodies both for “catching” the antigen and for signaling. In the present study we used a monoclonal antibody to SCT 1–11 as the capture antibody. This antibody was biotinylated before use in streptavidin-coated microtitration plates. The polyclonal antibody labeled with Eu chelate was used as a signaling marker. This combination of antibodies resulted in an assay that was three to four times more sensitive than the previous IFMA, with a detection limit of 0.3 pmol/L serum. Intact SCT 1–32 was detected by the assay (recoveries 94–96%), but not the fragments SCT 1–11 and SCT 10–32 or human calcitonin. Dilutions of plasma samples containing SCT were parallel to the calibration curve of SCT 1–32. Pharmacokinetic studies of SCT, 100 IU administered intramuscularly to 10 men, indicated peak serum concentrations of 32–128 pmol/L within 10–20 min with apparent half-life of 56 ± 18 min (mean ± SD). This new assay will allow study of the pharmacokinetics of new calcitonin preparations that do not require injection.

Preparation and Identification of Monoclonal Antibody against BPA

2012 ◽  
Vol 550-553 ◽  
pp. 1438-1442
Author(s):  
Lei Zhang ◽  
Su Qing Zhao ◽  
Hong Huang
Keyword(s):  
Monoclonal Antibody ◽  
Detection Limit ◽  
Linear Regression ◽  
Regression Line ◽  
High Specificity ◽  
Cross Reactivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Hydroxybenzoic Acid ◽  
Coating Antigen ◽  
Line Equation

Firstly, BPA structure was modified, then coupling BPA with BSA or OVA to prepare immunogen and coating antigen. Five Balb/C mice were immunized with BPA-BSA. Finally an antibody was prepared and the indirect competitive enzyme-linked immunosorbent assay was founded. Results:(1) The monoclonal antibody belongs to IgG1 subtype and К light chain.(2) The antibody titer is 1：256000, the most suitable concentration of coating antigen is 2μg/mL, and the optimal dilution of antibody and HRP are 1:16000 and 1:10000 respectively. (3)The linear regression line equation is y = 0.1139x + 0.1046, correlation coefficient is R2=0.97, the detection limit is 0.911ng/mL and IC50 is 2.454×103ng/mL. (4)The monoclonal antibody has high specificity for the cross reactivity with phenol, hydroquinone, and tert-butyl hydroquinone being lower than 0.01%, except ortho-hydroxybenzoic acid 2.1%. (5)The recovery range is 93%～116% and 89%～112% when adding BPA into black samples.（6）When the method was used in real materials to detect BPA residual, the results were proximate to the dates by HPLC.

scholarly journals Design of Novel Haptens and Development of Monoclonal Antibody-Based Immunoassays for the Simultaneous Detection of Tylosin and Tilmicosin in Milk and Water Samples

Biomolecules ◽  
2019 ◽  
Vol 9 (12) ◽  
pp. 770 ◽  
Author(s):  
Jian-Xin Huang ◽  
Chan-Yuan Yao ◽  
Jin-Yi Yang ◽  
Zhen-Feng Li ◽  
Fan He ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Water Samples ◽  
Simultaneous Detection ◽  
Aldehyde Group ◽  
Cross Reactivity ◽  
Side Chain ◽  
Enzyme Linked Immunosorbent Assay ◽  
Inhibition Concentration ◽  
Ic50 Values ◽  
Optimized Conditions

In this work, a monoclonal antibody-based indirect competitive enzyme-linked immunosorbent assay (icELISA) was established to detect tylosin and tilmicosin in milk and water samples. A sensitive and specific monoclonal antibody was prepared by rational designed hapten, which was achieved by directly oxidizing the aldehyde group on the side chain of tylosin to the carboxyl group. Under the optimized conditions, the linear range of icELISA for tylosin and tilmicosin were 1.3 to 17.7 ng/mL and 2.0 to 47.4 ng/mL, with half-maximal inhibition concentration (IC50) values of 4.7 and 9.6 ng/mL, respectively. The cross-reactivity with other analogues of icELISA was less than 0.1%. The average recoveries of icELISA for tylosin and tilmicosin ranged from 76.4% to 109.5% in milk and water samples. Besides, the detection results of icELISA showed good correlations with HPLC-MS/MS. The proposed icELISA was satisfied for rapid and specific screening of tylosin and tilmicosin residues in milk and water samples.

Serological assessment of type ΧVΙ collagen in patients with colorectal cancer and ulcerative colitis.

2018 ◽  
Vol 36 (4_suppl) ◽  
pp. 698-698
Author(s):  
Christina Jensen ◽  
Signe Holm Nielsen ◽  
Joachim Høg Mortensen ◽  
Jens Kjeldsen ◽  
Lone Gabriels Klinge ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Ulcerative Colitis ◽  
Monoclonal Antibody ◽  
Cell Invasion ◽  
Odds Ratio ◽  
Enzyme Linked Immunosorbent Assay ◽  
Healthy Controls ◽  
Ecm Remodeling ◽  
Future Studies ◽  
Ecm Architecture

698 Background: Altered extracellular matrix (ECM) remodeling is an important part of the pathology of gastrointestinal (GI) disorders. In the intestine, type XVI collagen (col-16) plays a role in pathogenesis by affecting ECM architecture and induce cell invasion. Measuring col-16 in serum may therefore have biomarker potential in GI disorders such as colorectal cancer (CRC) and ulcerative colitis (UC). The aim of this study was to determine whether col-16 can serve as a biomarker for altered ECM remodeling in patients with CRC and UC. Methods: A monoclonal antibody was raised against the C-terminal end of col-16 (C16-C) and a competitive enzyme-linked immunosorbent assay (ELISA) was developed and technically validated. Levels of C16-C were measured in serum from patients with CRC (before (n = 50) and three months after (n = 23) tumor resections), UC (n = 39) and healthy controls (n = 50). Results: The C16-C ELISA was specific towards the C-terminal of col-16. C16-C was significantly elevated both in serum from patients with CRC (p = 0.0026) and UC (p < 0.0001) compared to controls. No difference was detected in levels of C16-C between patients with CRC at baseline and three months after tumor resections (p > 0.999). Levels of C16-C identified patients with a GI disorder with a positive predictive value of 0.9 and an odds ratio of 12 (95%CI = 4.5-29.5, p < 0.0001). Conclusions: The newly developed assay detected significantly elevated levels of C16-C in serum from patients with GI disorders compared to controls suggesting its potential as a biomarker in this setting. Future studies are needed to validate these findings.

Sensitive Measurement of Thrombopoietin by a Monoclonal Antibody Based Sandwich Enzyme-linked Immunosorbent Assay

1997 ◽  
Vol 78 (04) ◽  
pp. 1262-1267 ◽  
Author(s):  
Claudia C Folman ◽  
Albert E G K von dem Borne ◽  
Irma H J A M Rensink ◽  
Winald Gerritsen ◽  
C Ellen van der Schoot ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Immunosorbent Assay ◽  
Platelet Transfusion ◽  
Large Scale ◽  
Limit Of Detection ◽  
Cross Reactivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Reliable Tool

SummaryIn this report a sensitive enzyme-linked immunosorbent assay (ELISA) for the measurement of plasma thrombopoietin (Tpo) is described that is solely based on monoclonal antibodies (MoAbs).The assay has an intra and inter-assay variance of 5-7% and 7-13%, respectively. Native and recombinant human Tpo (rhTpo) were recognized equally well, no cross reactivity with other cytokines was found and rhTpo added to plasma and serum was completely recovered. With the ELISA, Tpo concentrations in EDTA-anticoagulated plasma of all controls (n = 193) could be determined, since the limit of detection (2 ± 0.8 A.U./ml, mean ± sd) was lower than the concentration found in controls (11 ± 8 A.U./ml, mean ± sd; 2.5th-97.5th percentile: 4-32 A.U./ml). Tpo levels in serum were on average 3.4 times higher than in plasma.We showed in vivo that Tpo is bound by platelets, as in thrombocytopenic patients (n = 5) a platelet transfusion immediately led to a drop in plasma Tpo level, whereas in patients receiving chemotherapy the induced thrombocytopenia was followed by a rise in plasma Tpo levels.In summary, these results indicate that this ELISA is a reliable tool for Tpo measurements and is applicable for large scale studies.

Crystal adsorption and growth slowing by nephrocalcin, albumin, and Tamm-Horsfall protein

1988 ◽  
Vol 255 (6) ◽  
pp. F1197-F1205 ◽  
Author(s):  
E. M. Worcester ◽  
Y. Nakagawa ◽  
C. L. Wabner ◽  
S. Kumar ◽  
F. L. Coe
Keyword(s):  
Monoclonal Antibody ◽  
Crystal Growth ◽  
Calcium Binding ◽  
Calcium Oxalate Monohydrate ◽  
Cross Reactivity ◽  
Competitive Elisa ◽  
Enzyme Linked Immunosorbent Assay ◽  
Renal Cells ◽  
Carboxyglutamic Acid ◽  
Albumin Adsorption

Urine inhibition of calcium oxalate monohydrate (COM) crystal growth (CG) seems due to a glycoprotein that contains gamma-carboxyglutamic acid and has been named nephrocalcin (NC); however, Tamm-Horsfall protein (THP) and albumin resemble NC and make its measurement and role uncertain. NC in urine is aggregated to molecular mass 64 kDa and higher, similar to albumin (64 kDa) and THP (87 kDa). Albumin and THP are calcium binding, albumin adsorbs to COM crystals, and THP has been described as an inhibitor of COM growth. Antisera to NC have cross-reacted with THP even though the NC was isolated from cultured renal cells. Here we have compared highly purified NC, THP, and albumin adsorption with COM crystals and CG inhibition; also we compared their patterns of cross-reactivities with a new antiserum against NC and a monoclonal antibody to THP. NC adsorbs to COM crystals, THP does not. Albumin and THP do not inhibit CG. Cross-reactivity of albumin and THP to the antiserum is slight by direct enzyme-linked immunosorbent assay and nonexistent by competitive ELISA; reaction of NC to the anti-THP monoclonal antibody is absent.

scholarly journals New Sensitive Competitive Enzyme-Linked Immunosorbent Assay Using a Monoclonal Antibody against Nonstructural Protein 1 of West Nile Virus NY99

2011 ◽  
Vol 19 (2) ◽  
pp. 277-283 ◽  
Author(s):  
Jiro Hirota ◽  
Yoshihiro Shimoji ◽  
Shinya Shimizu
Keyword(s):  
Monoclonal Antibody ◽  
West Nile Virus ◽  
Neutralization Test ◽  
Nonstructural Protein ◽  
Cross Reactivity ◽  
Encephalitis Virus ◽  
The Other ◽  
Enzyme Linked Immunosorbent Assay ◽  
West Nile ◽  
Nonstructural Protein 1

ABSTRACTAn anti-West Nile virus (anti-WNV) monoclonal antibody, SHW-7A11, was developed for competitive enzyme-linked immunosorbent assays (c-ELISAs). SHW-7A11 reacted with nonstructural protein 1 in Western blot analysis. SHW-7A11 was relatively specific for the WNV strain NY99 and recognized Kunjin and Eg101 strains in indirect ELISAs. Two c-ELISAs were developed for sera diluted 10 and 100 times and named c-ELISA10 and c-ELISA100, respectively. Both c-ELISAs detected antibodies against WNV NY99 and Kunjin strains. Little cross-reactivity was observed for antibodies against Japanese encephalitis virus and St. Louis encephalitis virus in these assays. Using the cutoff point for the St. Louis encephalitis virus, all WNV-infected chickens were found to be positive on day 21 after infection in both c-ELISAs. On the other hand, all infected chickens were found to be positive on day 35 after infection in a virus neutralization test. Our newly developed SHW-7A11-based c-ELISA can detect WNV infection with sera diluted 10 to 100 times. Therefore, this c-ELISA can be used for WNV serosurveillance of chickens and wild birds.

scholarly journals Detection of Pork in Heat-Processed Meat Products by Monoclonal Antibody-Based ELISA

2000 ◽  
Vol 83 (1) ◽  
pp. 79-85 ◽  
Author(s):  
Fur-Chi Chen ◽  
Y-H Peggy Hsieh
Keyword(s):  
Monoclonal Antibody ◽  
Muscle Protein ◽  
Meat Products ◽  
Antibody Test ◽  
Cross Reactivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Processed Meat ◽  
Coefficients Of Variation ◽  
Test Kit ◽  
Meat Samples

Abstract An enzyme-linked immunosorbent assay (ELISA) using a monoclonal antibody to a porcine thermal-stable muscle protein was developed for detection of pork in cooked meat products. The assay specifically detects porcine skeletal muscle, but not cardiac muscle, smooth muscle, blood, and nonmuscle organs. No cross-reactivity was observed with common food proteins. Validity of the assay was evaluated with laboratory formulated and commercial meat samples. The detection limit was determined as 0.5% (w/w) pork in heterologous meat mixtures. Overall, intra- and inter-assay coefficients of variation were 5.8 and 7.9%, respectively. The accuracy in analyzing market samples was 100% as verified by product labeling and confirmed by a commercial polycolonal antibody test kit.

scholarly journals Differentiation of Two Bovine Lentiviruses by a Monoclonal Antibody on the Basis of Epitope Specificity

2001 ◽  
Vol 8 (2) ◽  
pp. 283-287 ◽  
Author(s):  
Ling Zheng ◽  
Shucheng Zhang ◽  
Charles Wood ◽  
Sanjay Kapil ◽  
Graham E. Wilcox ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Fusion Protein ◽  
Capsid Protein ◽  
Recombinant Fusion Protein ◽  
Bovine Immunodeficiency Virus ◽  
Antigenic Determinants ◽  
Enzyme Linked Immunosorbent Assay ◽  
Binding Assays ◽  
Immunodeficiency Virus ◽  
Competitive Binding Assays

ABSTRACT Bovine immunodeficiency virus (BIV) and Jembrana disease virus (JDV) are bovine lentiviruses that are closely related genetically. A recombinant fusion protein containing the capsid protein of BIV expressed in Escherichia coli was used to immunize mice and produce monoclonal antibodies. Six hybridomas specific for BIV capsid protein were identified, and one antibody, designated 10H1, was characterized further. Competitive binding assays were performed to analyze the topography of antigenic determinants by enzyme-linked immunosorbent assay and demonstrated the existence of at least three distinct antigenic determinants on capsid protein. The monoclonal antibody reacted specifically with both BIV capsid and the recombinant fusion protein in Western immunoblot analyses. However, it did not react with the recombinant capsid fusion protein of JDV, indicating that BIV contains at least one unique epitope in the capsid protein that is absent in JDV. Further mapping of the epitope by chemical cleavage analysis identified that the epitope is located at the 6.4-kDa N terminus of the 29-kDa capsid protein. This monoclonal antibody assay will be valuable for distinguishing the two closely related lentiviruses by Western blotting.

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