scholarly journals Cytotoxin antibody-based colourimetric sensor for field-level differential detection of elapid among big four snake venom

2021 ◽  
Vol 15 (10) ◽  
pp. e0009841
Author(s):  
Sherin Kaul ◽  
L. Sai Keerthana ◽  
Pankaj Kumar ◽  
Komal Birader ◽  
Yathirajarao Tammineni ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Differential Detection ◽  
Serum Samples ◽  
Naja Naja ◽  
Detection Tool ◽  
Limit Of Quantitation ◽  
Immunological Reactions ◽  
Medical Guidelines ◽  
Big Four ◽  
Spiked Serum

Development of a rapid, on-site detection tool for snakebite is highly sought after, owing to its clinically and forensically relevant medicolegal significance. Polyvalent antivenom therapy in the management of such envenomation cases is finite due to its poor venom neutralization capabilities as well as diagnostic ramifications manifested as untoward immunological reactions. For precise molecular diagnosis of elapid venoms of the big four snakes, we have developed a lateral flow kit using a monoclonal antibody (AB1; IgG1 – κ chain; Kd: 31 nM) generated against recombinant cytotoxin-7 (rCTX-7; 7.7 kDa) protein of the elapid venom. The monoclonal antibody specifically detected the venoms of Naja naja (p < 0.0001) and Bungarus caeruleus (p<0.0001), without showing any immunoreactivity against the viperidae snakes in big four venomous snakes. The kit developed attained the limit of quantitation of 170 pg/μL and 2.1 ng/μL in spiked buffer samples and 28.7 ng/μL and 110 ng/μL in spiked serum samples for detection of N. naja and B. caeruleus venoms, respectively. This kit holds enormous potential in identification of elapid venom of the big four snakes for effective prognosis of an envenomation; as per the existing medical guidelines.

A Simple and Rapid Light-Initiated Chemiluminescence Assay for Quantitation of Artemisia-Specific Immunoglobulin E

2021 ◽  
pp. 1-8
Author(s):  
Dandan Liu ◽  
Bei Zhang ◽  
Lina Zhu ◽  
Lisheng Zheng ◽  
Shaoshen Li ◽  
...  
Keyword(s):  
Food Allergen ◽  
Clinical Guideline ◽  
Immunoglobulin E ◽  
Limit Of Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Chemiluminescence Assay ◽  
Limit Of Quantitation ◽  
Specific Immunoglobulin E ◽  
Specific Immunoglobulin

<b><i>Background:</i></b> Light-initiated chemiluminescence assay (LICA) is a homogeneous assay that has been successfully used for the quantitation of food allergen-specific immunoglobulin E (sIgE), but not inhaled allergen-sIgE. Simultaneously, current assays used to detect allergen-sIgE are serum consuming and/or time consuming. Hence, we established a method for the quantitation of <i>Artemisia</i>-sIgE based on LICA and verified its performance according to the clinical guideline documents, laying a foundation for the quantitation of inhaled and food allergen-sIgE in parallel on LICA. <b><i>Methods:</i></b> The assay was established after optimizing the first incubation time and the dilutions of <i>Artemisia</i>-coated chemibeads, biotinylated goat anti-human IgE, and serum. In order to quantitate <i>Artemisia</i>-sIgE, the calibration curve was established with a high positive serum of known concentration. The assay performance was confirmed per the clinical guideline documents. In addition, the correlation between the results of LICA and capture enzyme-linked immunosorbent assay was evaluated. <b><i>Results:</i></b> The developed LICA’s coefficients of variation of repeatability and intermediate precision were 3.20%, 2.14%, and 3.85% and 4.30%, 4.00%, and 4.40%, respectively. The limit of detection was 0.10 kU<sub>A</sub>/L, and the limit of quantitation was 0.11 kU<sub>A</sub>/L. The range of linearity was from 0.27 kU<sub>A</sub>/L to 97.53 kU<sub>A</sub>/L (<i>r</i> = 0.9968). The correlation coefficient (<i>r</i>) for the correlation analysis between results of LICA and capture ELISA was 0.9087. This assay was successfully applied in 64 human serum samples, showing good sensitivity (82.20%) and specificity (100%). <b><i>Conclusion:</i></b> An <i>Artemisia</i>-sIgE quantitation assay based on LICA was successfully established. Its performance satisfied the clinical requirements and could be widely used in clinical laboratories.

scholarly journals Novel multiplex real-time quantitative PCR detecting system approach for direct detection ofCandida aurisand its relatives in spiked serum samples

2019 ◽  
Vol 14 (1) ◽  
pp. 33-45 ◽  
Author(s):  
Amir Arastehfar ◽  
Wenjie Fang ◽  
Farnaz Daneshnia ◽  
Abdullah MS Al-Hatmi ◽  
Wanqing Liao ◽  
...  
Keyword(s):  
Real Time ◽  
Quantitative Pcr ◽  
System Approach ◽  
Direct Detection ◽  
Serum Samples ◽  
Real Time Quantitative Pcr ◽  
Spiked Serum

Snakebite Mitigation Project of the Madras Crocodile Bank/Centre for Herpetology, India: background and a brief summary of activities

2018 ◽  
Vol 113 (12) ◽  
pp. 818-819
Author(s):  
Romulus Whitaker
Keyword(s):  
Public Awareness ◽  
Rural India ◽  
Awareness Campaign ◽  
Naja Naja ◽  
Life Saving ◽  
Mitigation Project ◽  
Treatment Centre ◽  
Echis Carinatus ◽  
Agricultural Areas ◽  
Big Four

Abstract Snakebite is a serious problem in rural India where several highly venomous species are commonly found in and around agricultural areas where prey such as rodents and amphibians are abundant. Four snake species, referred to as the Big Four, are responsible for the most serious and fatal bites: spectacled cobra (Naja naja), Russell’s viper (Daboia russelii), common krait (Bungarus caeruleus) and saw-scaled viper (Echis carinatus). A polyvalent antivenom is made to treat these bites but public awareness and distribution of this life-saving drug is inadequate. The Madras Crocodile Bank and its partners are conducting a snakebite project which includes venom sampling and research, snake and snakebite treatment centre mapping, and a nationwide awareness campaign for snakebite mitigation.

Evaluation of two point of care technologies for measuring monoclonal antibody therapeutic-A concentrations in blood

Bioanalysis ◽  
2020 ◽  
Vol 12 (20) ◽  
pp. 1449-1458
Author(s):  
Saloumeh K Fischer ◽  
Kathi Williams ◽  
Ian Harmon ◽  
Bryan Bothwell ◽  
Hua Xu ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Real Time ◽  
Point Of Care ◽  
Turnaround Time ◽  
Monitoring Methods ◽  
Sample Collection ◽  
Time Data ◽  
Serum Samples ◽  
Quick Turnaround Time

Aim: Current blood monitoring methods require sample collection and testing at a central lab, which can take days. Point of care (POC) devices with quick turnaround time can provide an alternative with faster results, allowing for real-time data leading to better treatment decisions for patients. Results/Methodology: An assay to measure monoclonal antibody therapeutic-A was developed on two POC devices. Data generated using 75 serum samples (65 clinical & ten spiked samples) show correlative results to the data generated using Gyrolab technology. Conclusion: This case study uses a monoclonal antibody therapeutic-A concentration assay as an example to demonstrate the potential of POC technologies as a viable alternative to central lab testing with quick results allowing for real-time decision-making.

scholarly journals Development of Recombinant Nucleoprotein-Based Diagnostic Systems for Lassa Fever

2007 ◽  
Vol 14 (9) ◽  
pp. 1182-1189 ◽  
Author(s):  
Masayuki Saijo ◽  
Marie-Claude Georges-Courbot ◽  
Philippe Marianneau ◽  
Victor Romanowski ◽  
Shuetsu Fukushi ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Hela Cells ◽  
Recombinant Baculovirus ◽  
Lassa Fever ◽  
Rabbit Serum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Lassa Virus ◽  
Diagnostic Systems ◽  
Serum Samples ◽  
Capture Elisa

ABSTRACT Diagnostic systems for Lassa fever (LF), a viral hemorrhagic fever caused by Lassa virus (LASV), such as enzyme immunoassays for the detection of LASV antibodies and LASV antigens, were developed using the recombinant nucleoprotein (rNP) of LASV (LASV-rNP). The LASV-rNP was expressed in a recombinant baculovirus system. LASV-rNP was used as an antigen in the detection of LASV-antibodies and as an immunogen for the production of monoclonal antibodies. The LASV-rNP was also expressed in HeLa cells by transfection with the expression vector encoding cDNA of the LASV-NP gene. An immunoglobulin G enzyme-linked immunosorbent assay (ELISA) using LASV-rNP and an indirect immunofluorescence assay using LASV-rNP-expressing HeLa cells were confirmed to have high sensitivity and specificity in the detection of LASV-antibodies. A novel monoclonal antibody to LASV-rNP, monoclonal antibody 4A5, was established. A sandwich antigen capture (Ag-capture) ELISA using the monoclonal antibody and an anti-LASV-rNP rabbit serum as capture and detection antibodies, respectively, was then developed. Authentic LASV nucleoprotein in serum samples collected from hamsters experimentally infected with LASV was detected by the Ag-capture ELISA. The Ag-capture ELISA specifically detected LASV-rNP but not the rNPs of lymphocytic choriomeningitis virus or Junin virus. The sensitivity of the Ag-capture ELISA in detecting LASV antigens was comparable to that of reverse transcription-PCR in detecting LASV RNA. These LASV rNP-based diagnostics were confirmed to be useful in the diagnosis of LF even in institutes without a high containment laboratory, since the antigens can be prepared without manipulation of the infectious viruses.

Sandwich enzyme immunoassay of osteocalcin in serum with use of an antibody against human osteocalcin

1993 ◽  
Vol 39 (6) ◽  
pp. 942-947 ◽  
Author(s):  
D A Monaghan ◽  
M J Power ◽  
P F Fottrell
Keyword(s):  
Monoclonal Antibody ◽  
Enzyme Immunoassay ◽  
Solid Phase ◽  
Sandwich Elisa ◽  
Normal Subjects ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Standard Curve ◽  
Microtiter Plates ◽  
Human Osteocalcin

Abstract We have developed and thoroughly validated a solid-phase sandwich enzyme-linked immunosorbent assay (ELISA) on microtiter plates for osteocalcin in human serum with use of an antibody raised against human osteocalcin. We used a monoclonal antibody against bovine osteocalcin as the capture antibody; the second antibody was a polyclonal antibody against human osteocalcin. The amount of bound second antibody was determined with use of swine anti-rabbit antibody labeled with horseradish peroxidase. We demonstrated independence of volume and determined the recovery of added standard and within- and between-assay precision. The minimal detection limit for osteocalcin was between 1.0 and 1.5 micrograms/L and the midpoint of the standard curve ranged from 14 to 17 micrograms/L. The intraassay CV was &lt; or = 8% in the range 2.7-52 micrograms/L; the interassay CV was usually &lt; or = 15% in the same range. Analytical recovery of human osteocalcin standard added to serum samples was consistently &gt; 90%. Values for osteocalcin measured in serum from 44 normal subjects were similar to those obtained with a competitive enzyme immunoassay (EIA) that used a monoclonal antibody against bovine osteocalcin. There was a good correlation between the two assays [r2 = 0.877, slope and intercept (+/- SE) = 0.88(+/- 0.051) and 0.316(+/- 0.523), respectively]. The range and mean (+/- SD) for the sandwich ELISA and the competitive EIA were 1.7-18.1 micrograms/L [8.7(+/- 4.4) micrograms/L] and 1.9-22.8 micrograms/L [9.1(+/- 4.4) micrograms/L], respectively.

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.

Immunoradiometric assay of human intact proinsulin applied to patients with type 2 diabetes, impaired glucose tolerance, and hyperandrogenism

1994 ◽  
Vol 40 (5) ◽  
pp. 754-757 ◽  
Author(s):  
D Chevenne ◽  
J Ruiz ◽  
L Lohmann ◽  
A Laudat ◽  
H Leblanc ◽  
...  
Keyword(s):  
Type 2 Diabetes ◽  
Monoclonal Antibody ◽  
Detection Limit ◽  
Glucose Tolerance ◽  
Impaired Glucose Tolerance ◽  
Immunoradiometric Assay ◽  
Serum Samples ◽  
The Mean

Abstract We describe an immunoradiometric assay for human intact proinsulin in serum. In this method, one monoclonal antibody, coated onto polyacrylamide beads, cross-reacts with proinsulins and insulin. A sandwich is formed with intact proinsulin, split (65-66) proinsulin, and des (64-65) proinsulin binding with an 125I-labeled monoclonal antibody specific for an epitope at the intact B-C junction of proinsulin. Because split (65-66) and des (64-65) proinsulin concentrations are very low in serum, this assay essentially measures intact proinsulin. When we used 1-mL serum samples, the mean detection limit was 0.4 pmol/L. Mean proinsulin concentrations (pmol/L) were 3.4 (range 1-9.1) in healthy fasting subjects, 28.5 (9.7-101) in patients with type 2 diabetes (treated with metformin and sulfonylureas), 5.0 (1.6-9.3) in women with hyperandrogenism and normal insulinemia, 10.3 (2.6-36) in women with hyperandrogenism and hyperinsulinemia, and 8.5 (4.8-21.3) in patients with impaired glucose tolerance.

A novel monoclonal antibody specific for the 402His variant of complement factor H enables identification of the Tyr402His polymorphic status in serum samples

2007 ◽  
Vol 44 (16) ◽  
pp. 3951
Author(s):  
Svetlana Hakobyan ◽  
Claire L. Harris ◽  
Agustin Tortojada ◽  
Elena Giocochea de Jorge ◽  
Santiago Rodriguez de Cordoba ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Factor H ◽  
Complement Factor H ◽  
Complement Factor ◽  
Serum Samples

Validation of galectin-7 using backscattering interferometry.

2012 ◽  
Vol 30 (30_suppl) ◽  
pp. 15-15
Author(s):  
Ian Roys Olmsted ◽  
Pierre P. Massion ◽  
Darryl J. Bornhop ◽  
Mohamed Hassanein ◽  
Megan Hoeksema
Keyword(s):  
Lung Cancer ◽  
Dose Response ◽  
Response Curve ◽  
Limit Of Detection ◽  
Detection Limits ◽  
Small Sample ◽  
Dose Response Curve ◽  
Label Free ◽  
Serum Samples ◽  
Spiked Serum

15 Background: More than 60% of lung cancer patients are diagnosed with advanced disease due to a lack of early diagnosis tools. These patients are ineligible for surgical resection and have a poor prognosis. This situation could be avoided if we could quantify lung cancer specific biomarkers at sufficiently low concentrations, providing an indication of disease threat. Backscattering Inerferometry (BSI) is a label-free sensor with a simple optical train that is used to quantify biomarkers in complex matrices at picomolar to femtomolar levels. Here we demonstrate that BSI can enable lung cancer biomarker validation in complex, volume constrained samples and at detection limits significantly better than ELISA. Methods: A BSI dose response curve for Galectin-7 was constructed by incubating increasing concentrations (0-20 ng/mL) of recombinant galectin-7 with 100 ng/mL of polyclonal antibody. The samples were mixed at 300 RPM for 2 before being injected into the BSI instrument and measured in triplicate each day. All reagents used were obtained from a commercial ELISA kit. An ELISA dose response curve was similarly constructed using spiked serum and spiked plasma according to the manufacturer’s recommended procedure. Finally, 9 patient serum samples were quantified using BSI and ELISA for direct comparison of the two technologies. Results: BSI was used to quantify galectin-7 in spiked serum and patients samples. The lower limit of detection with standards was determined to be 0.5 ng/mL for ELISA in serum, 10 ng/mL for ELISA in plasma, and 0.04 ng/mL for BSI in serum. In the analysis of 9 patient serum samples, all were quantifiable using BSI, which enjoys low pg/mL detection limits, yet with ELISA only 5 samples contained a galectin-7 concentration high enough to be measured. In this small sample set there was a good correlation between disease state and galectin-7 concentration as measured by BSI. Conclusions: This study demonstrates that BSI is well-suited for biomarker detection and validation, having a greater dynamic operating range and much lower limits of detection than standard commercially available ELISA kits. Future work will focus on detecting other NSCLC biomarkers that are undetectable with currently available technology.

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