scholarly journals Modified ELISA for Ultrasensitive Diagnosis

2021 ◽  
Vol 10 (21) ◽  
pp. 5197
Author(s):  
Naoko Tsurusawa ◽  
Jyunhao Chang ◽  
Mayuri Namba ◽  
Daiki Makioka ◽  
Sou Yamura ◽  
...  
Keyword(s):  
Infectious Diseases ◽  
Present Article ◽  
Immunosorbent Assay ◽  
Current Literature ◽  
Quantitative Measurement ◽  
Review Article ◽  
Detection Sensitivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Ultrasensitive Detection ◽  
Lifestyle Related Diseases

An enzyme-linked immunosorbent assay (ELISA) can be used for quantitative measurement of proteins, and improving the detection sensitivity to the ultrasensitive level would facilitate the diagnosis of various diseases. In the present review article, we first define the term ‘ultrasensitive’. We follow this with a survey and discussion of the current literature regarding modified ELISA methods with ultrasensitive detection and their application for diagnosis. Finally, we introduce our own newly devised system for ultrasensitive ELISA combined with thionicotinamide adenine dinucleotide cycling and its application for the diagnosis of infectious diseases and lifestyle-related diseases. The aim of the present article is to expand the application of ultrasensitive ELISAs in the medical and biological fields.

scholarly journals An Antigen Capture Enzyme-Linked Immunosorbent Assay Reveals High Levels of the Dengue Virus Protein NS1 in the Sera of Infected Patients

2000 ◽  
Vol 38 (3) ◽  
pp. 1053-1057 ◽  
Author(s):  
Paul R. Young ◽  
Paige A. Hilditch ◽  
Cheryl Bletchly ◽  
Wendy Halloran
Keyword(s):  
Monoclonal Antibodies ◽  
Dengue Virus ◽  
Acute Phase ◽  
Immunosorbent Assay ◽  
Nonstructural Protein ◽  
Surrogate Marker ◽  
Detection Sensitivity ◽  
Severe Dengue ◽  
Clinical Samples ◽  
Enzyme Linked Immunosorbent Assay

We describe the development of a capture enzyme-linked immunosorbent assay for the detection of the dengue virus nonstructural protein NS1. The assay employs rabbit polyclonal and monoclonal antibodies as the capture and detection antibodies, respectively. Immunoaffinity-purified NS1 derived from dengue 2 virus-infected cells was used as a standard to establish a detection sensitivity of approximately 4 ng/ml for an assay employing monoclonal antibodies recognizing a dengue 2 serotype-specific epitope. A number of serotype cross-reactive monoclonal antibodies were also shown to be suitable probes for the detection of NS1 expressed by the remaining three dengue virus serotypes. Examination of clinical samples demonstrated that the assay was able to detect NS1 with minimal interference from serum components at the test dilutions routinely used, suggesting that it could form the basis of a useful additional diagnostic test for dengue virus infection. Furthermore, quantitation of NS1 levels in patient sera may prove to be a valuable surrogate marker for viremia. Surprisingly high levels of NS1, as much as 15 μg/ml, were found in acute-phase sera taken from some of the patients experiencing serologically confirmed dengue 2 virus secondary infections but was not detected in the convalescent sera of these patients. In contrast, NS1 could not be detected in either acute-phase or convalescent serum samples taken from patients with serologically confirmed primary infection. The presence of high levels of secreted NS1 in the sera of patients experiencing secondary dengue virus infections, and in the context of an anamnestic antibody response, suggests that NS1 may contribute significantly to the formation of the circulating immune complexes that are suspected to play an important role in the pathogenesis of severe dengue disease.

Evaluation of a Sensitive Reverse Transcription PCR–Enzyme-Linked Immunosorbent Assay for Detection of Hepatitis A Virus in Oysters (Saccostrea glomerata) on the East Coast of the Gulf of Thailand

2014 ◽  
Vol 77 (5) ◽  
pp. 859-863 ◽  
Author(s):  
URAIWAN INTAMASO ◽  
SITTHISAK KETKHUNTHOD
Keyword(s):  
Immunosorbent Assay ◽  
Reverse Transcription ◽  
Hepatitis A ◽  
East Coast ◽  
Hepatitis A Virus ◽  
Detection Sensitivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Gulf Of Thailand ◽  
Rt Pcr ◽  
The Gulf Of Thailand

Hepatitis A virus (HAV) contamination in food can lead to major health problems. We developed a combination reverse transcription (RT) PCR method plus enzyme-linked immunosorbent assay (ELISA) to detect HAV in fresh oysters harvested along the east coast of the Gulf of Thailand. Viral nucleic acid was extracted via the glycine–arginine–polyethylene glycol method followed by RT-PCR amplification with specifically designed primers against HAV and an ELISA to detect the digoxigenin-labeled RT-PCR products. The ELISA in concert with the RT-PCR protocol further increased the detection sensitivity by 100-fold for the HAV genome and 10-fold in artificially contaminated oysters. The overall sensitivity of the RT-PCR in combination with the ELISA was 31.88 pg and 16 PFU/g, respectively. The ELISA increases the specificity of the RT-PCR assay for detecting naturally occurring HAV in oysters. This combined RT-PCR-ELISA approach is a practical and sensitive method for HAV detection and can be utilized in routine screening for HAV in shellfish.

scholarly journals An enzyme-linked immunosorbent assay for erythropoietin using monoclonal antibodies, tetrameric immune complexes, and substrate amplification

Blood ◽  
1989 ◽  
Vol 74 (2) ◽  
pp. 622-628 ◽  
Author(s):  
AW Wognum ◽  
PM Lansdorp ◽  
AC Eaves ◽  
G Krystal
Keyword(s):  
Monoclonal Antibodies ◽  
Immunosorbent Assay ◽  
Biological Fluids ◽  
Normal Serum ◽  
Detection Sensitivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Response Curves ◽  
Human Erythropoietin ◽  
Healthy Donors ◽  
Noncovalent Labeling

We recently reported the development of several monoclonal antibodies (MoAbs) to native human erythropoietin (Ep). In the present study we have used the two antibodies with highest affinity to develop a two- sided or sandwich enzyme-linked immunosorbent assay (ELISA) to measure Ep in human serum. In this assay Ep is incubated in microtiter wells precoated with the first (IgE) anti-Ep antibody. Assay wells are then incubated with the second (IgG1) anti-Ep antibody, which is labeled noncovalently with the enzyme alkaline phosphatase (AP) by means of bispecific tetrameric antibody complexes consisting of IgG1 anti-Ep cross-linked to IgG1 anti-AP using rat MoAbs specific for mouse IgG1. Application of this noncovalent labeling procedure, in combination with substrate amplification, results in a detection sensitivity of 0.5 to 1.0 mU/sample (5 to 10 mU/mL), which makes this assay suitable for measuring normal serum Ep levels. The validity of this ELISA for quantitating Ep in biological fluids was demonstrated by the parallelism obtained between pure recombinant Ep dose-response curves and those obtained with plasma and serum from healthy donors and patients with various hematologic disorders. Normal plasma Ep levels detected with this ELISA ranged from 9 to 101 mU/mL with a mean of 32 +/- 23 (SD) mU/mL. Ep levels in sera from patients with polycythemia vera were in the low to normal range, whereas Ep levels in sera from patients with secondary polycythemia and patients with aplastic anemia were moderately to strongly elevated. These results demonstrate that the Ep-ELISA is a sensitive, reliable, and nonradioactive immunologic method for quantitating Ep levels and should prove useful in a variety of clinical and laboratory settings.

scholarly journals A rapid and highly sensitive biomarker detection platform based on a temperature-responsive liposome-linked immunosorbent assay

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Runkai Hu ◽  
Keitaro Sou ◽  
Shinji Takeoka
Keyword(s):  
Immunosorbent Assay ◽  
Limit Of Detection ◽  
Specific Antigen ◽  
Fluorescence Emission ◽  
Detection Sensitivity ◽  
Sensitive Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Temperature Responsive ◽  
Highly Sensitive ◽  
Highly Sensitive Detection

Abstract The enzyme-linked immunosorbent assay (ELISA) is widely used in various fields to detect specific biomarkers. However, ELISA tests have limited detection sensitivity (≥ 1 pM), which is insufficiently sensitive for the detection of small amounts of biomarkers in the early stages of disease or infection. Herein, a method for the rapid and highly sensitive detection of specific antigens, using temperature-responsive liposomes (TLip) containing a squaraine dye that exhibits fluorescence at the phase transition temperature of the liposomes, was developed. A proof-of-concept study using biotinylated TLip and a streptavidin-immobilized microwell plate showed that the TLip bound to the plate via specific molecular recognition could be distinguished from unbound TLip within 1 min because of the difference in the heating time required for the fluorescence emission of TLip. This system could be used to detect prostate specific antigen (PSA) based on a sandwich immunosorbent assay using detection and capture antibodies, in which the limit of detection was as low as 27.6 ag/mL in a 100-μL PSA solution, 0.97 aM in terms of molar concentration. The present temperature-responsive liposome-linked immunosorbent assay provides an advanced platform for the rapid and highly sensitive detection of biomarkers for use in diagnosis and biological inspections.

scholarly journals Detection of Type A, B, E, and F Clostridium botulinum Neurotoxins in Foods by Using an Amplified Enzyme-Linked Immunosorbent Assay with Digoxigenin-Labeled Antibodies

2006 ◽  
Vol 72 (2) ◽  
pp. 1231-1238 ◽  
Author(s):  
Shashi K. Sharma ◽  
Joseph. L. Ferreira ◽  
Brian S. Eblen ◽  
Richard C. Whiting
Keyword(s):  
Immunosorbent Assay ◽  
Clostridium Botulinum ◽  
Botulinum Neurotoxin ◽  
Large Scale ◽  
Lethal Dose ◽  
Meat Products ◽  
Type A ◽  
Detection Sensitivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Botulinum Neurotoxins

ABSTRACT An amplified enzyme-linked immunosorbent assay (ELISA) for the detection of Clostridium botulinum complex neurotoxins was evaluated for its ability to detect these toxins in food. The assay was found to be suitable for detecting type A, B, E, and F botulinum neurotoxins in a variety of food matrices representing liquids, solid, and semisolid food. Specific foods included broccoli, orange juice, bottled water, cola soft drinks, vanilla extract, oregano, potato salad, apple juice, meat products, and dairy foods. The detection sensitivity of the test for these botulinum complex serotypes was found to be 60 pg/ml (1.9 50% lethal dose [LD50]) for botulinum neurotoxin type A (BoNT/A), 176 pg/ml (1.58 LD50) for BoNT/B, 163 pg/ml for BoNT/E (4.5 LD50), and 117 pg/ml for BoNT/F (less than 1 LD50) in casein buffer. The test could also readily detect 2 ng/ml of neurotoxins type A, B, E, and F in a variety of food samples. For specificity studies, the assay was also used to test a large panel of type A C. botulinum, a smaller panel of proteolytic and nonproteolytic type B, E, and F neurotoxin-producing Clostridia, and nontoxigenic organisms using an overnight incubation of toxin production medium. The assay appears to be an effective tool for large-scale screening of the food supply in the event of a botulinum neurotoxin contamination event.

Ultrasensitive optofluidic enzyme-linked immunosorbent assay by on-chip integrated polymer whispering-gallery-mode microlaser sensors

Lab on a Chip ◽  
2020 ◽  
Vol 20 (14) ◽  
pp. 2438-2446 ◽  
Author(s):  
Xia Ouyang ◽  
Tong Liu ◽  
Yangxi Zhang ◽  
Jijun He ◽  
Zijian He ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Microfluidic Chip ◽  
Whispering Gallery Mode ◽  
Enzyme Linked Immunosorbent Assay ◽  
Ultrasensitive Detection ◽  
Whispering Gallery ◽  
On Chip ◽  
Detection Of Biomarkers

Polymer whispering-gallery-mode microlaser sensors are optically 3D μ-printed and then integrated within a microfluidic chip for ultrasensitive detection of biomarkers.

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