scholarly journals Rapid and simultaneous purification of aflatoxin B1, zearalenone and deoxynivalenol using their monoclonal antibodies and magnetic nanoparticles

2021 ◽  
Author(s):  
Hyuk-Mi Lee ◽  
Hwan-Goo Kang
Keyword(s):  
Monoclonal Antibodies ◽  
Magnetic Nanoparticles ◽  
Aflatoxin B1 ◽  
Animal Feed ◽  
Immunoaffinity Chromatography ◽  
The Novel ◽  
Buffer Solution ◽  
Purification Method ◽  
Recovery Rates ◽  
Single Spike

AbstractTo develop a new simple and simultaneous purification method for mycotoxins in feeds and grains, magnetic nanoparticles (MNPs) conjugated with monoclonal antibodies (mAbs) against mycotoxins were used to separate aflatoxin B1 (AFB1), zearalenone (ZEA) and deoxynivalenol (DON). For a single spike of each mycotoxin into the buffer solution (16% MeOH in PBS), mean recoveries were 93.1–95.0% for AFB1 (5–20 ng/mL spiked), 87.2–96.0% for ZEA (125–500 ng/mL spiked) and 75.2–96.9% for DON (250–1,000 ng/mL spiked) by HPLC and ELISA. Recoveries of AFB1 (20 ng/mL) and ZEA (500 ng/mL) simultaneously spiked into the buffer solution were 87.0 and 99.8%, respectively. Recovery rates of AFB1/DON and DON/ZEA spiked simultaneously were 86.2%/76.6% and 92.0%/86.7%, respectively, at concentrations of 20 ng/mL AFB1, 500 ng/mL ZEA, and 1,000 ng/mL DON. Recoveries using the novel mAb–MNP conjugated system in a buffer solution simultaneously spiked with AFB1, ZEA and DON were 82.5, 94.6 and 73.4%, respectively. Recoveries of DON in animal feed were 107.7–132.5% at concentrations of 250–1,000 ng/g spiked in feed. The immunoaffinity chromatography (IAC) clean-up method was compared with the purification method using novel mAb–MNP. After fortification of animal feed with AFB1 (5, 10 and 20 ng/g feed) and ZEA (125, 250 and 500 ng/g feed), AFB1 and ZEA were purified using both the methods. In the case of the novel mAb-MNP conjugated system, mean recoveries for AFB1 were 89.4, 73.1 and 88.3% at concentrations of 5, 10 and 20 ng/g feed, respectively. For ZEA, mean recoveries were 86.7, 85.9 and 79.1% at concentrations of 125, 250 and 500 ng/g, respectively. For IAC purification, recoveries were 42.9–45.1% for AFB1 and 96.8–103.2% for ZEA. In conclusion, the present purification method using monoclonal antibodies conjugated to MNPs can be used for simple and simultaneous purification of mycotoxins from feed and maize.

Use of immunoaffinity chromatography for purification of 125I-labeled human prolactin.

1983 ◽  
Vol 29 (2) ◽  
pp. 241-245 ◽  
Author(s):  
M C Stuart ◽  
L M Boscato ◽  
P A Underwood
Keyword(s):  
Monoclonal Antibodies ◽  
Affinity Chromatography ◽  
Gel Filtration ◽  
Immunoaffinity Chromatography ◽  
Purification Method ◽  
Assay Sensitivity ◽  
Simple Method ◽  
Antigen B ◽  
Human Prolactin ◽  
Chloramine T

Abstract We assessed a simple method for purifying 125I-labeled human prolactin, taking advantage of the abundant supplies of monoclonal antibodies available. 125I-Labeled human prolactin purified by immunoaffinity chromatography is compared with that purified by gel filtration on Sephadex G-100. We used monoclonal antibodies to prolactin to prepare the affinity chromatography columns. Prolactin was radiolabeled by the Chloramine T method, purified by each of the above procedures, and the binding and displacement characteristics were studied in radioimmunoassays in which either monoclonal antibodies or a rabbit anti-prolactin serum was the first antibody. A nonimmune fraction of 125I-labeled prolactin that co-eluted with the immunoreactive hormone from Sephadex G-100 was removed by affinity chromatography, which increased the antibody binding of 125I-labeled prolactin in the radioimmunoassay in the absence of unlabeled antigen (B/T0, in percent) twofold or more, increased the assay sensitivity, and increased the slope of antigen displacement measured by the 50% intercept. Several advantages make this the purification method of choice.

Study on the Novel Label Free Impedimetric Immunosensor for Aflatoxin B1 Based on Poly(o-phenylenediamine) Film Modified Gold Electrode

2011 ◽  
Vol 322 ◽  
pp. 385-389 ◽  
Author(s):  
Ning Ning Wu ◽  
Li Xin Cao ◽  
Pei Sheng Yan ◽  
Ming Hao Wang
Keyword(s):  
Aflatoxin B1 ◽  
Gold Electrode ◽  
Biomolecular Recognition ◽  
The Novel ◽  
Label Free ◽  
Buffer Solution ◽  
Solution Ph ◽  
Linear Detection ◽  
Impedimetric Immunosensor

A label free impedimetric immunosensor for the determination of aflatoxin B1 (AFB1) was fabricated by immobilizing anti-AFB1 onto Poly o-phenylenediamine (PoPD) electropolymerized film modified gold electrode by glutaraldehyde (GA) cross-linking. An electrochemical interfacial modeling of biomolecular recognition was recommended and reasonably interpreted. EIS technology was employed for the quantitative determination. The linear detection concentration ranges of AFB1 were 0.03~0.1 μg/mL and 0.1~0.7 μg/mL. The detection limit was 2.8×10-8μg/mL. The immunosensor could be reused more than 10 times when renewed by HCl-Glycine buffer solution (pH 2.6).

scholarly journals Novel idiotypic and antigen-binding characteristics in two anti-dinitrophenyl monoclonal antibodies.

1977 ◽  
Vol 146 (1) ◽  
pp. 282-286 ◽  
Author(s):  
N H Sigal
Keyword(s):  
Monoclonal Antibodies ◽  
Germ Line ◽  
Hypervariable Region ◽  
Weight Basis ◽  
Antigen Binding ◽  
The Novel ◽  
Maximum Frequency ◽  
Somatic Variant ◽  
Binding Characteristics ◽  
Cell Pool

Monoclonal anti-dinitrophenyl antibodies generated in the splenic focus system from B cells of adult BALB/c mice were studied for the presence or absence of murine anti-T15 (M anti-T15) reactivity and for their ability to bind phosphorylcholine (PC). Two foci of the 680 clones analyzed bound PC, and one of these antibodies reacted with M anti-T15 and anti-Fab on a 1:1 weight basis. The discovery of a clonotype reactive with M anti-15 but not with rabbit anti-T15 (R anti-T15) serum, the converse of the R anti-T15+, M anti-T15- clonotype identified in the PC-specific repertoire, points to the novel idiotypic relationships which may be found among homogeneous antibodies binding diverse antigens. The R anti-T15-, M anti-T15+ clonotype may represent a distinct set of hypervariable region sequences inserted into the T15 framework or may be a somatic variant of the T15 germ-line sequence. In addition, the maximum frequency with which this clonotype occurs within the B-cell pool is estimated.

scholarly journals Preparation of Hybridomas Producing Monoclonal Antibodies against Aflatoxin B1 as a Tool to Control Hepatocellular Carcinoma

2019 ◽  
Vol 13 ◽  
pp. 1-12
Author(s):  
Manal M.E. Ahmed ◽  
Rafik Soliman ◽  
Jakeen Eljakee ◽  
Ahmed El-Sanousi ◽  
Haitham Amer ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Aflatoxin B1 ◽  
Quantitative Measurement ◽  
Myeloma Cell ◽  
Selective Medium ◽  
Immunization Schedule ◽  
Spleen Cells ◽  
Myeloma Cell Line ◽  
High Affinity ◽  
Multiple Uses

Hybridomas that secreted antibodies against aflatoxin B1 for multiple uses were prepared using a unique immunization schedule. Aflatoxin B1-BSA conjugate was used for immunization of Balb/c mice. Spleen cells were harvested from the hyper immunized mice to be fused with myeloma cell line (P3NS1) using polyethylene glycol 3000, 50% concentration as a fusogenic agent. The produced hybridomas were selected using HAT selective medium that was replaced by complete HT medium. From the 10thday after fusion, wells that contain colonies of hybridomas covering 30% or greater of the wells surface were screened for production of monoclonal antibodies against aflatoxin B1 using ELISA. 21 hybridomas were found to be reactive to aflatoxin B1. All were found to belong to IgG2aisotype except one was found to belong to IgM isotype. The prepared monoclonal antibodies and their application to immunoassays represents a useful and rapid quantitative measurement with high affinity and low detection limits in order to purify environmentally occurring levels of this carcinogen specially in areas at high risk for liver cancer.

scholarly journals The Immune Response to Tumors as a Tool toward Immunotherapy

2011 ◽  
Vol 2011 ◽  
pp. 1-12 ◽  
Author(s):  
F. Pandolfi ◽  
R. Cianci ◽  
D. Pagliari ◽  
F. Casciano ◽  
C. Bagalà ◽  
...  
Keyword(s):  
Immune Response ◽  
Monoclonal Antibodies ◽  
Cancer Colorectal ◽  
Immune Cell ◽  
Cytotoxic T Cells ◽  
Tumor Infiltrating Lymphocytes ◽  
Hematologic Malignancies ◽  
The Novel ◽  
Infiltrating Lymphocytes

Until recently cancer medical therapy was limited to chemotherapy that could not differentiate cancer cells from normal cells. More recently with the remarkable mushroom of immunology, newer tools became available, resulting in the novel possibility to attack cancer with the specificity of the immune system. Herein we will review some of the recent achievement of immunotherapy in such aggressive cancers as melanoma, prostatic cancer, colorectal carcinoma, and hematologic malignancies. Immunotherapy of tumors has developed several techniques: immune cell transfer, vaccines, immunobiological molecules such as monoclonal antibodies that improve the immune responses to tumors. This can be achieved by blocking pathways limiting the immune response, such as CTLA-4 or Tregs. Immunotherapy may also use cytokines especially proinflammatory cytokines to enhance the activity of cytotoxic T cells (CTLs) derived from tumor infiltrating lymphocytes (TILs). The role of newly discovered cytokines remains to be investigated. Alternatively, an other mechanism consists in enhancing the expression of TAAs on tumor cells. Finally, monoclonal antibodies may be used to target oncogenes.

Magnetic nanoparticles-based immunoassay for aflatoxin B1 using porous g-C3N4 nanosheets as fluorescence probes

2019 ◽  
Vol 278 ◽  
pp. 147-152 ◽  
Author(s):  
Huizhi Xie ◽  
Jing Dong ◽  
Junling Duan ◽  
Juying Hou ◽  
Shiyun Ai ◽  
...  
Keyword(s):  
Magnetic Nanoparticles ◽  
Aflatoxin B1 ◽  
Fluorescence Probes

Biochemical and Biophysical characterization of the main protease, 3-chymotrypsin-like protease (3CLpro), from the novel coronavirus disease 19 (COVID-19)

2020 ◽  
Author(s):  
Juliana C. Ferreira ◽  
Wael M. Rabeh
Keyword(s):  
Antiviral Drug ◽  
Analytical Techniques ◽  
The Novel ◽  
Buffer Solution ◽  
Biophysical Characterization ◽  
Main Protease ◽  
Wide Ph Range ◽  
The Stability ◽  
Ph Range ◽  
Novel Coronavirus

Abstract Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) is responsible for the novel coronavirus disease 2019 (COVID-19). An appealing antiviral drug target is the coronavirus 3C-like protease (3CLpro) that is responsible for the processing of the viral polyproteins and liberation of functional proteins essential for the maturation and infectivity of the virus. In this study, multiple thermal analytical techniques have been implemented to acquire the thermodynamic parameters of 3CLpro at different buffer conditions. 3CLpro exhibited relatively high thermodynamic stabilities over a wide pH range; however, the protease was found to be less stable in the presence of salts. Divalent metal cations reduced the thermodynamic stability of 3CLpro more than monovalent cations; however, altering the ionic strength of the buffer solution did not alter the stability of 3CLpro. Furthermore, the most stable thermal kinetic stability of 3CLpro was recorded at pH 7.5, with the highest enthalpy of activation calculated from the slope of Eyring plot. The biochemical and biophysical properties of 3CLpro explored here will improve the solubility and stability of 3CLpro for optimum conditions for the setup of an enzymatic assay for the screening of inhibitors to be used as lead candidates in the drug discovery and antiviral design for therapeutics against COVID-19.

scholarly journals Carbon-Based Magnetic Nanocarrier for Controlled Drug Release: A Green Synthesis Approach

2018 ◽  
Vol 5 (1) ◽  
pp. 1 ◽  
Author(s):  
Jessica Oliveira ◽  
Raquel Rodrigues ◽  
Lillian Barros ◽  
Isabel Ferreira ◽  
Luís Marchesi ◽  
...  
Keyword(s):  
Drug Delivery ◽  
Magnetic Nanoparticles ◽  
Drug Release ◽  
Phosphate Buffer ◽  
Colloidal Stability ◽  
Ph Dependence ◽  
Phosphate Buffer Solution ◽  
Controlled Drug Release ◽  
Buffer Solution ◽  
Carbon Based

In this study, hydrophilic magnetic nanoparticles were synthesized by green routes using a methanolic extract of Rubus ulmifolius Schott flowers. The prepared magnetic nanoparticles were coated with carbon-based shell for drug delivery application. The nanocomposites were further chemically functionalized with nitric acid and, sequentially, with Pluronic® F68 (CMNPs-plur) to enhance their colloidal stability. The resulting material was dispersed in phosphate buffer solution at pH 7.4 to study the Doxorubicin loading. After shaking for 48 h, 99.13% of the drug was loaded by the nanocomposites. Subsequently, the drug release was studied in different working phosphate buffer solutions (i.e., PB pH 4.5, pH 6.0 and pH 7.4) to determine the efficiency of the synthesized material for drug delivery as pH-dependent drug nanocarrier. The results have shown a drug release quantity 18% higher in mimicking tumor environment than in the physiological one. Therefore, this study demonstrates the ability of CMNPs-plur to release a drug with pH dependence, which could be used in the future for the treatment of cancer "in situ" by means of controlled drug release.

scholarly journals Determination of Morphine in Human Urine by the Novel Competitive Fluorescence Immunoassay

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Jie Cao ◽  
Xiao-Ying Chen ◽  
Wu-Rong Zhao
Keyword(s):  
Monoclonal Antibodies ◽  
Human Urine ◽  
Screening Tool ◽  
Fluorescein Isothiocyanate ◽  
Cross Reactivity ◽  
The Novel ◽  
Fluorescence Immunoassay ◽  
Coated Plate ◽  
Efficient Screening

A competitive fluorescence immunoassay for the identification and quantification of morphine has been developed on the basis of hapten-coated plate format. Hapten was prepared through covalent conjugating a morphine derivative with albumin bovine. In the immunoassay, the hapten was inoculated on a 96-well plate and then bound with monoclonal antibodies labeled with a signal indicating dye, fluorescein isothiocyanate (FITC). Unbound FITC-antibodies were rinsed off from the plate. The fluorescein intensity decreases in the presence of morphine molecules due to the competitively binding to antibodies against hapten. The intensity is inversely correlated with the concentration of morphine. In quantitative analysis for urine samples, we obtained a linearity range of 0.2 μg/mL∼2.5 μg/mL, along with a detection limit of c.a. 1 ng/mL. The fluorescence immunoassay shows low cross-reactivity (below 10%) to 6-acetylmorphine, 3-acetylmorphine, and heroine. The developed method produced comparable results to the standard GC-MS/MS method. In conclusion, a rapid and efficient screening tool for morphine in clinical human urine has been established.

A 96-well Microfiltration Assay for Measurement of Total Clottable Fibrinogen

1999 ◽  
Vol 81 (02) ◽  
pp. 264-267 ◽  
Author(s):  
A. Keller ◽  
S. Argirion ◽  
D. L. Heene ◽  
C. E. Dempfle
Keyword(s):  
Clinical Trials ◽  
Protein Concentration ◽  
The Novel ◽  
Plasma Samples ◽  
Clotting Time ◽  
Buffer Solution ◽  
Clinical Routine ◽  
Vacuum Suction ◽  
Time Determination ◽  
Method R

SummaryIn clinical routine use, fibrinogen is measured by clotting-time methods, or by clot turbidity in photometric prothrombin time determination. For calibration of these assays measurement of total thrombinclottable protein has been recommended. We have now developed a microfiltration assay for total thrombin-clottable protein. Plasma samples were mixed with thrombin in a 96-well microfiltration device. After clot formation, the fluid was extracted by vacuum suction, and fibrin adherent to the filter membranes washed with buffer. Membrane segments with adherent fibrin were recovered from the 96-well manifold with a punch and transferred to tubes containing denaturing buffer solution. After dissolution of fibrin, protein concentration was determined by optical absorption at 280 nm. The microfiltration assay displayed a high correlation with the total clottable protein method (R = 0.95), and fibrinogen antigen (r = 0.96). Correlation with clotting time assays, and PT-derived fibrinogen in 150 clinical plasma samples was in the range of r = 0.84 to r = 0.97. Intraassay and day-to-day variability of the assay was comparable to the conventional total clottable fibrinogen assay. The novel microfiltration assay appears to be well suited for measurement of large series of samples for calibration, screening purposes, and clinical trials.

Sign in / Sign up

Export Citation Format

Share Document