Monoclonal antibodies evaluated for use in a screening assay for conjugates of human serum albumin and pyridoxal 5'-phosphate.

1989 ◽  
Vol 35 (8) ◽  
pp. 1756-1760 ◽  
Author(s):  
B B Miller ◽  
W E Turner
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Human Serum Albumin ◽  
Human Serum ◽  
Limit Of Detection ◽  
Screening Method ◽  
Serum Samples ◽  
Screening Assay ◽  
Antibody Avidity ◽  
Negative Results

Abstract This enzyme immunoassay (EIA) was developed to measure pyridoxal 5'-phosphate bound to albumin (PLP-HSA) in human serum. The monoclonal antibody titer was 1:2000 and a sequential saturation analysis curve, prepared with samples containing from 10 to 1000 nmol/L, showed a 50% inhibition of antibody at 50 nmol of the conjugate per liter. The lower limit of detection for PLP-HSA was 10 nmol/L, a sensitivity 1000-fold greater than that for any potential interferent. When serum samples gave negative results in the assay, we compared the antigenicity of the principal sites for PLP binding on HSA. It was apparent that the preferred physiological site was not antigenic; however, three additional sites for PLP binding on HSA elicited comparable antibody avidity. This EIA is potentially quite sensitive and specific for PLP-HSA, but considerable additional effort is required to convert serum PLP to an HSA-bound form detectable in the assay, which limits its application as a screening method.

scholarly journals Development of Aflatoxin B1-Lysine Adduct Monoclonal Antibody for Human Exposure Studies

2001 ◽  
Vol 67 (6) ◽  
pp. 2712-2717 ◽  
Author(s):  
Jia-Sheng Wang ◽  
Salahaddin Abubaker ◽  
Xia He ◽  
Guiju Sun ◽  
Paul T. Strickland ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
High Risk ◽  
Bovine Serum Albumin ◽  
Dna Adducts ◽  
Human Exposure ◽  
Limit Of Detection ◽  
Serum Samples ◽  
Bovine Serum Albumin Conjugate ◽  
Aflatoxin B

ABSTRACT Mouse monoclonal antibodies were developed against a synthetic aflatoxin B1 (AFB)-lysine–cationized bovine serum albumin conjugate. The isotype of one of these antibodies, IIA4B3, has been classified as immunoglobulin G1(λ). The affinity and specificity of IIA4B3 were further characterized by a competitive radioimmunoassay. The affinities of IIA4B3 for AFB and its associated adducts and metabolites are ranked as follows: AFB-lysine > 8,9-dihydro-8-(2,6-diamino-4-oxo-3,4-dihydropyrimid-5-yl formamido)-9-hydroxy-AFB > AFB = 8,9-dihydro-8-(N 7-guanyl)-9-hydroxy-AFB > aflatoxin M1 > aflatoxin Q1. IIA4B3 had about a 10-fold higher affinity for binding to AFB-lysine adduct than to AFB when 3H-AFB–lysine was used as the tracer. The concentration for 50% inhibition for AFB-lysine was 0.610 pmol; that for AFB was 6.85 pmol. IIA4B3 had affinities at least sevenfold and twofold higher than those of 2B11, a previously developed antibody against parent AFB, for the major aflatoxin-DNA adducts 8,9-dihydro-8-(N 7-guanyl)-9-hydroxy-AFB and 8,9-dihydro-8-(2,6-diamino-4-oxo-3,4-dihydropyrimid-5-yl formamido)-9-hydroxy-AFB, respectively. An analytical method based on a competitive radioimmunoassay with IIA4B3 and3H-AFB–lysine was validated with a limit of detection of 10 fmol of AFB-lysine adduct. The method has been applied to the measurement of AFB-albumin adduct levels in human serum samples collected from the residents of areas at high risk for liver cancer.

scholarly journals Gradients of a Sandwich-structured Immunosensor Controlled Using Magnetized Masks for Determination of Human Serum Albumin Using Electrochemiluminescence

2021 ◽  
Author(s):  
Chun-Yao Huang ◽  
Chi-Jung Chang ◽  
Bohr-Ran Huang ◽  
Chien-Hsing Lu ◽  
jemkun chen
Keyword(s):  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
Magnetic Beads ◽  
Limit Of Detection ◽  
High Reliability ◽  
Water Evaporation ◽  
Serum Samples ◽  
Dry Process ◽  
Physical Features

Abstract Background: Separation of macromolecules or particles from a colloid system to from gradient structure on the surface has been employed for biosensing systems, suggesting an enhancement of the chemical and physical features of particles. Performance of an electrochemiluminescence (ECL) immunosensor was employed to improve with a particle gradient.Results: Magnetic beads with silicon dioxide coating were adopted as nanocarriers for gradient manipulation and immobilized with the primary antibody. Cadmium telluride quantum dots (CdTe QDs) were coated with a layer of protein G for conjugation and orientation of secondary antibody as signal labels. ECL immunosensor gradients upon the electrode were formed by magnetolithography with magnetized nickel masks of column and stripe arrays at various scales. The immunosensor generally aggregated as island on the substrate through a dry process of water evaporation leading to a decrease of efficiency in the characteristic signals. Stripe arrays of magnetized nickel were designed to generate cylindrical magnetic flux on the substrate to improve the particle manipulation with the gradient. Various gradients of the sandwich-structured immunosensor substantially affected the electrochemical performance. Compared to the gradient-free immunosensor, the gradient of the immunosensor generated using the 3-μm-stripe array mask of magnetolithography enhanced the ECL intensity ~2.2 times. Conclusions: The results of quantification of human serum albumin (HSA) with the gradient immunosensor showed a broad linear range (15–420 ng mL−1), a low limit of detection (8 ng mL−1) and high reliability for HSA-spiked serum samples, indicating that the immunosensor gradient substantially enhances the performance of the ECL assay.

Competitive Direct Enzyme-Linked Immunosorbent Screening Assay for the Detection of Sulfamethazine Contamination of Animal Feeds

1991 ◽  
Vol 74 (5) ◽  
pp. 784-789 ◽  
Author(s):  
Dixon E Holland Deborah ◽  
Stanley E Katz
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Polyclonal Antibodies ◽  
Screening Method ◽  
Screening Assay ◽  
Animal Feeds ◽  
Feed Samples

Abstract A sensitive screening method has been developed for detecting sulfamethazine (SMZ) contamination of feeds by using either polyclonal or monoclonal antibodies and a direct competitive enzyme-linked immunosorbent screening assay (ELISA). Feed samples of 25.0 g are extracted with 0.5N HCI and centrifuged. The extract is adjusted to pH 7.0 with 3.0N NaOH and recentrifuged. This pH-adjusted extract is used in the EUSA. Levels as low as 0.004 μg SMZ/g feed were detected In supplemented extracts by polyclonal antibodies; levels of 0.4 μg SMZ/g feed were detected by a monoclonal antibody.

Study of the antigenic structure of human serum albumin with monoclonal antibodies

1985 ◽  
Vol 22 (1) ◽  
pp. 1-10 ◽  
Author(s):  
Noëlle Doyen ◽  
Claude Lapresle ◽  
Pierre Lafaye ◽  
Jean-Claude Mazie
Keyword(s):  
Monoclonal Antibodies ◽  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
Antigenic Structure

Drug/Antibody Conjugates. In Vitro Cytotoxicity of a Human Serum Albumin-Mediated Conjugate of Methotrexate with Anti-MM46 Monoclonal Antibody

1987 ◽  
Vol 29 (4) ◽  
pp. 566-568
Author(s):  
Noriaki Endo ◽  
Yoshinori Kato ◽  
Yumiko Takeda ◽  
Masahiko Saito ◽  
Naoji Umemoto ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
In Vitro Cytotoxicity ◽  
Antibody Conjugates

scholarly journals Development of a Sensitive Bead-Based Assay for Enhanced Monoclonal Antibody Detection

2011 ◽  
Vol 1346 ◽  
Author(s):  
Manuel E. Ruidíaz ◽  
Natalie Mendez ◽  
Ana B. Sanchez ◽  
Bradley T. Messmer ◽  
Andrew C. Kummel
Keyword(s):  
Monoclonal Antibody ◽  
Limit Of Detection ◽  
Solid Phase ◽  
Solid Phase Peptide Synthesis ◽  
Fluorescent Labeling ◽  
Antibody Detection ◽  
Serum Samples ◽  
Mimetic Peptide ◽  
Mimetic Peptides ◽  
Zero Response

ABSTRACTMonoclonal antibodies are increasingly used in the treatment of cancer due to their enhanced targeting and immune system stimulation properties. Dosage guidelines typically do not account for personal cancer load or metabolism, thereby possibly affecting treatment outcome or causing unwanted side effects. The requirement for an assay that can quickly and precisely measure the concentration of the monoclonal antibody in a serum sample of a patient during therapy is unmet. A bead-based assay with peptide antigen mimetics has been developed to rapidly determine the concentration of antibody drug present in serum specimens with high sensitivity. Alemtuzumab (anti-CD52) and rituximab (anti-CD20) antigen mimetic peptides, as discovered by phage display, were synthesized on 10 um TentaGel resin beads using conventional solid phase peptide synthesis techniques. The beads were modified to allow for multiplexing and microfluidic handling via fluorescent labeling and magnetic functionalization. The antigen-displaying fluoromagnetic particles were incubated with spiked serum samples which allowed free antibody to be captured. Primary antibody detection was performed on alemtuzumab while rituximab detection was used to compensate for non-specific serum binding to the beads. After washing, the beads were incubated with a fluorescently tagged secondary label for detection by flow cytometry. (Results) A fast, low cost, specific assay has been developed with several key techniques which allows detection at low concentration (0.1ug/ml) of spiked samples. Primary to achieving this detection limit was the implementation of a compensation scheme where two antigen mimetic peptides behave linearly (R2=0.996) which enables the calculation of the zero response of the antigen mimetic peptide of interest (alemtuzumab antigen mimetic) while measuring the zero response of the compensatory antigen mimetic peptide (rituximab antigen mimetic) during primary assay measurement. This reduces fluorescence response variation due to variations present due to sample preparation, storage and different patients because of the equivalent interactions these effects have on the compensatory beads. The developed assay is therefore robust against serum variation and enables a lower limit of detection.

scholarly journals Development of Vasoinhibin-Specific Monoclonal Antibodies

2021 ◽  
Vol 12 ◽  
Author(s):  
Nils Müller ◽  
Juan Pablo Robles ◽  
Magdalena Zamora ◽  
Johannes Ebnet ◽  
Hülya Markl-Hahn ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Serum Proteins ◽  
Limit Of Detection ◽  
Sandwich Elisa ◽  
Vascular Diseases ◽  
High Specificity ◽  
Cross Reactivity ◽  
Dimensional Structure ◽  
Serum Samples ◽  
Limit Of Quantitation

Vasoinhibin is a protein hormone with antiangiogenic, antivasodilatatory, and antivasopermeability effects generated by the proteolytic cleavage of prolactin. The discovery of its role in diabetic retinopathy and peripartum cardiomyopathy led to the evaluation of new pharmacological treatments in clinical interventional trials. However, the quantitative evaluation of vasoinhibin in biological samples from patients has not been possible due to the lack of vasoinhibin-specific antibodies. Recently, loop 1 of vasoinhibin was identified to have a different three-dimensional structure compared to PRL, and thus to contain vasoinhibin-specific epitopes. Here, we report the development of two sets of vasoinhibin-specific monoclonal antibodies against two neighboring regions of the vasoinhibin loop 1. An experimental sandwich ELISA with two monoclonal anti-vasoinhibin antibodies was developed, which had no cross-reactivity to recombinant human full-length prolactin. The ELISA had a quantitation limit of 100 ng/ml, and intra-assay- and inter-assay coefficients of variation of 12.5% and 14%, respectively. The evaluation of 15 human serum samples demonstrated concentrations of below limit of detection (n=3), below limit of quantitation (n=1) and between 0.23 µg/ml (230 ng/ml) to 605 µg/ml (n=12) in the quantifiable range. Despite the high specificity of the monoclonal-monoclonal antibody sandwiches which discriminate vasoinhibin from PRL, there might be cross-reactivities by serum proteins other than vasoinhibin. A fully established vasoinhibin ELISA may support diagnostic and therapeutic measures in vascular diseases.

scholarly journals Reusable, Noninvasive, and Sensitive Fluorescence Enhanced ZnO-Nanorod-Based Microarrays for Quantitative Detection of AFP in Human Serum

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Saima Rafique ◽  
Farukh Kiyani ◽  
Sumbal Jawaid ◽  
Rubina Nasir ◽  
Mahmoosh Ahmad ◽  
...  
Keyword(s):  
Human Serum ◽  
Dynamic Range ◽  
Limit Of Detection ◽  
Zno Nanorods ◽  
Protein Microarrays ◽  
Great Promise ◽  
Quantitative Detection ◽  
Zno Nanorod ◽  
Serum Samples ◽  
Protein Assay

The fabrication of sensitive protein microarrays such as PCR used in DNA microarray is challenging due to lack of signal amplification. The development of microarrays is utilized to improve the sensitivity and limitations of detection towards primal cancer detection. The sensitivity is enhanced by the use of ZnO-nanorods and is investigated as a substrate which enhance the florescent signal to diagnose the hepatocellular carcinoma (HCC) at early stages. The substrate for deposition of ZnO-nanorods is prepared by the conventional chemical bath deposition method. The resultant highly dense ZnO-nanorods enhance the fluorescent signal 7.2 times as compared to the substrate without ZnO-nanorods. The microarray showed sensitivity of 1504.7 ng ml-1 and limit of detection of 0.1 pg ml-1 in wide dynamic range of 0.05 pg-10 μg ml-1 for alpha fetoprotein (AFP) detection in 10% human serum. This immunoassay was successfully applied for human serum samples to detect tumor marker with good recoveries. The ZnO-nanorod substrate is a simple protein microarray which showed a great promise for developing a low-cost, sensitive, and high-throughput protein assay platform for several applications in both fundamental research and clinical diagnosis.

scholarly journals A Fluorescence Inner-Filter Effect Based Sensing Platform for Turn-On Detection of Glutathione in Human Serum

Sensors ◽  
2019 ◽  
Vol 19 (2) ◽  
pp. 228 ◽  
Author(s):  
Shurong Tang ◽  
Xiuhua You ◽  
Quanhui Fang ◽  
Xin Li ◽  
Guangwen Li ◽  
...  
Keyword(s):  
Human Serum ◽  
Limit Of Detection ◽  
Cost Effective ◽  
Inner Filter Effect ◽  
Disease Diagnosis ◽  
Fluorescent Properties ◽  
Serum Samples ◽  
Turn On ◽  
Filter Effect ◽  
Sensing Platform

A novel turn-on fluorescence assay was developed for the rapid detection of glutathione (GSH) based on the inner-filter effect (IFE) and redox reaction. Molybdenum disulfide quantum dots (MoS2 QDs), which have stable fluorescent properties, were synthesized with hydrothermal method. Manganese dioxide nanosheets (MnO2 NSs) were prepared by exfoliating the bulk δ-MnO2 material in bovine serum albumin (BSA) aqueous solution. The morphology structures of the prepared nanoparticles were characterized by transmission electron microscope (TEM). Studies have shown that the fluorescence of MoS2 QDs could be quenched in the presence of MnO2 NSs as a result of the IFE, and is recovered after the addition of GSH to dissolve the MnO2 NSs. The fluorescence intensity showed a good linear relationship with the GSH concentration in the range 20–2500 μM, the limit of detection was 1.0 μM. The detection method was applied to the analysis of GSH in human serum samples. This simple, rapid, and cost-effective method has great potential in analyzing GSH and in disease diagnosis.

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