Solid-Phase Amplification and Detection: A Single-Tube Diagnostic Assay for Infectious Agents

2001 ◽  
Vol 6 (2) ◽  
pp. 131-136 ◽  
Author(s):  
MARIA SOMODEVILLA-TORRES ◽  
PETER TIMMS ◽  
RAY HARRIS ◽  
C. PHILLIP MORRIS ◽  
ANGELA VAN DAAL
Keyword(s):  
Solid Phase ◽  
Infectious Agents ◽  
Diagnostic Assay ◽  
Single Tube

Neonatal thyroxine screening by use of a single-tube solid-phase radioimmunoassay.

1978 ◽  
Vol 24 (10) ◽  
pp. 1755-1758 ◽  
Author(s):  
A B Bell ◽  
L H Coleman
Keyword(s):  
Mass Screening ◽  
Filter Paper ◽  
Coefficient Of Variation ◽  
Binding Proteins ◽  
Solid Phase ◽  
Serum Samples ◽  
Single Tube ◽  
Screening Programs ◽  
Quantitative Results ◽  
Solid Phase Radioimmunoassay

Abstract Filter paper discs saturated with dried blood can be used in the Immunotube solid-phase thyroxine radioimmunoassay. This assay utilizes polypropylene tubes to which antibody to thyroxine is covalently bound. The filter paper standards and samples are placed in the tubes, followed by an assay buffer that contains I125-labeled thyroxine and compounds to displace thyroxine from its binding proteins. After incubation, bound and free thyroxine are separated by aspirating or decanting the disc and buffer from the tube. The test can be used with 0.32 cm (1/8 inch) or 0.64 cm(1/4 inch) discs, and gives quantitative results that correlate well with those for serum samples. The intra-assay coefficient of variation is less than 10%. The assay may readily be mechanized with existing disc-punching equipment, and results of its use in mass screening programs are described.

Immunoenzymometric determination of trypsin/alpha 1-protease inhibitor complex in plasma of rats with experimental pancreatitis.

1986 ◽  
Vol 32 (9) ◽  
pp. 1738-1741 ◽  
Author(s):  
T S Fletcher ◽  
H Tsukamoto ◽  
C Largman
Keyword(s):  
Protease Inhibitor ◽  
Solid Phase ◽  
Gel Filtration ◽  
Experimental Pancreatitis ◽  
Size Fractionation ◽  
Single Tube ◽  
Inhibitor Complex ◽  
Cationic Trypsin ◽  
High Concentrations

Abstract The currently available radioimmunoassays of trypsin measure total immunoreactive trypsin (EC 3.1.1.7), which includes both trypsinogen and alpha 1-protease inhibitor-bound trypsin. Hitherto, the only way to differentiate these two forms of trypsin has been to fractionate them on a gel-filtration column. We describe here a solid-phase immunoenzymometric assay that rapidly measures the amount of cationic trypsin bound to alpha 1-protease inhibitor in the plasma of rats with experimental pancreatitis. The assay specifically measures this complex within the range from 0.2 to 5.0 ng without interference by high concentrations of free alpha 1-protease inhibitor. The high correlation (r = 0.985) of the values obtained by size fractionation and by this assay demonstrates the accuracy of the assay, which is the first single-tube method for determining this form of activated cationic trypsin in plasma.

scholarly journals Single-Tube Method for Nucleic Acid Extraction, Amplification, Purification, and Sequencing

2004 ◽  
Vol 50 (10) ◽  
pp. 1755-1761 ◽  
Author(s):  
Rebecca L Margraf ◽  
Sam Page ◽  
Maria Erali ◽  
Carl T Wittwer
Keyword(s):  
Nucleic Acid ◽  
Reverse Transcription ◽  
Solid Phase ◽  
Sequence Data ◽  
Hcv Rna ◽  
Acid Extraction ◽  
Nucleic Acid Extraction ◽  
Single Tube ◽  
Reverse Transcription Pcr ◽  
Conventional Methods

Abstract Background: The hepatitis C virus (HCV) genotype determines patient prognosis and duration of treatment, but sequencing of the gene is lengthy and labor-intensive. We used a commercially available nucleic acid extraction system to develop a single-tube extraction-to-sequencing (STETS) method for HCV genotyping. Methods: HCV RNA was purified and amplified in tubes coated with a solid-phase matrix that irreversibly bound nucleic acid during the extraction step. After reverse transcription-PCR, the amplicon was adsorbed to the original extraction matrix for purification and use in the subsequent sequencing reactions. Results: The STETS method generated genotyping-quality sequence for a range of HCV titers from 500 to 6 000 000 IU/mL. If a viral sample was detected during real-time reverse transcription-PCR, it could be sequenced and genotyped. Read lengths >600 bases were observed with the STETS method. Mixed infections were detected and genotyped if at least 15% of the minor species was present. Combining the STETS method with consecutive sequencing provided a means of performing both forward and reverse sequencing in a single tube. Conclusions: A single-tube nucleic acid extraction-to-sequencing method, which requires less time and labor than conventional methods, generates HCV sequence data that are equivalent to conventional methods and can be used to genotype HCV.

Pathology of Bone Cells in Pigs with Experimental Turbinate Osteoporosis (Atrophic Rhinitis)

1971 ◽  
Vol 29 ◽  
pp. 304-305
Author(s):  
A. W. Fetter ◽  
C. C. Capen
Keyword(s):  
Mucous Membrane ◽  
Bone Cells ◽  
Atrophic Rhinitis ◽  
Infectious Agents ◽  
Metabolic Disturbances ◽  
Progressive Reduction ◽  
Naturally Occurring ◽  
Fundamental Cause ◽  
Osteocytic Osteolysis ◽  
Uncertain Etiology

Atrophic rhinitis in swine is a disease of uncertain etiology in which infectious agents, hereditary predisposition, and metabolic disturbances have been reported to be of primary etiologic importance. It shares many similarities, both clinically and pathologically, with ozena in man. The disease is characterized by deformity and reduction in volume of the nasal turbinates. The fundamental cause for the localized lesion of bone in the nasal turbinates has not been established. Reduced osteogenesis, increased resorption related to inflammation of the nasal mucous membrane, and excessive resorption due to osteocytic osteolysis stimulated by hyperparathyroidism have been suggested as possible pathogenetic mechanisms.The objectives of this investigation were to evaluate ultrastructurally bone cells in the nasal turbinates of pigs with experimentally induced atrophic rhinitis, and to compare these findings to those in control pigs of the same age and pigs with the naturally occurring disease, in order to define the fundamental lesion responsible for the progressive reduction in volume of the osseous core.

Solid-phase immunoelectron microscopy for rapid diagnosis of enteroviruses

1984 ◽  
Vol 42 ◽  
pp. 226-227 ◽  
Author(s):  
K. Pegg-Feige ◽  
F. W. Doane
Keyword(s):  
Electron Microscopy ◽  
Cell Culture ◽  
Immunoelectron Microscopy ◽  
Detection Method ◽  
Solid Phase ◽  
Protein A ◽  
Plant Viruses ◽  
Rapid Diagnosis ◽  
Virus Diagnosis ◽  
Antiviral Antibody

Immunoelectron microscopy (IEM) applied to rapid virus diagnosis offers a more sensitive detection method than direct electron microscopy (DEM), and can also be used to serotype viruses. One of several IEM techniques is that introduced by Derrick in 1972, in which antiviral antibody is attached to the support film of an EM specimen grid. Originally developed for plant viruses, it has recently been applied to several animal viruses, especially rotaviruses. We have investigated the use of this solid phase IEM technique (SPIEM) in detecting and identifying enteroviruses (in the form of crude cell culture isolates), and have compared it with a modified “SPIEM-SPA” method in which grids are coated with protein A from Staphylococcus aureus prior to exposure to antiserum.

Application of solid-phase immune electron microscopy to study physical and stability characteristics of enterically transmitted non-a, non-b hepatitis virus

1988 ◽  
Vol 46 ◽  
pp. 80-81
Author(s):  
Charles D. Humphrey ◽  
E. H. Cook ◽  
Karen A. McCaustland ◽  
Daniel W. Bradley
Keyword(s):  
Electron Microscopy ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Solid Phase ◽  
Etiologic Agent ◽  
World Health ◽  
Health Concern ◽  
Morphological Identification ◽  
Immune Electron Microscopy

Enterically transmitted non-A, non-B hepatitis (ET-NANBH) is a type of hepatitis which is increasingly becoming a significant world health concern. As with hepatitis A virus (HAV), spread is by the fecal-oral mode of transmission. Until recently, the etiologic agent had not been isolated and identified. We have succeeded in the isolation and preliminary characterization of this virus and demonstrating that this agent can cause hepatic disease and seroconversion in experimental primates. Our characterization of this virus was facilitated by immune (IEM) and solid phase immune electron microscopic (SPIEM) methodologies.Many immune electron microscopy methodologies have been used for morphological identification and characterization of viruses. We have previously reported a highly effective solid phase immune electron microscopy procedure which facilitated identification of hepatitis A virus (HAV) in crude cell culture extracts. More recently we have reported utilization of the method for identification of an etiologic agent responsible for (ET-NANBH).

Identification of hepatitis a virus in cell cultures by solid phase immune electron microscopy

1986 ◽  
Vol 44 ◽  
pp. 238-239
Author(s):  
C.D. Humphrey ◽  
T.L. Cromeans ◽  
E.H. Cook ◽  
D.W. Bradley
Keyword(s):  
Electron Microscopy ◽  
Liquid Phase ◽  
Cell Cultures ◽  
Rapid Detection ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Solid Phase ◽  
Immune Electron Microscopy ◽  
Detection And Identification

There is a variety of methods available for the rapid detection and identification of viruses by electron microscopy as described in several reviews. The predominant techniques are classified as direct electron microscopy (DEM), immune electron microscopy (IEM), liquid phase immune electron microscopy (LPIEM) and solid phase immune electron microscopy (SPIEM). Each technique has inherent strengths and weaknesses. However, in recent years, the most progress for identifying viruses has been realized by the utilization of SPIEM.

Dynamic studies of silicide-mediated crystallization of amorphous silicon

1992 ◽  
Vol 50 (2) ◽  
pp. 1352-1353
Author(s):  
C. Hayzelden ◽  
J. L. Batstone
Keyword(s):  
Ion Implantation ◽  
Solid Phase ◽  
Metallic Phase ◽  
Single Crystal Substrate ◽  
Free Electrons ◽  
Melt Temperature ◽  
Transmission Electron ◽  
Crystal Substrate ◽  
High Resolution Tem

Epitaxial reordering of amorphous Si(a-Si) on an underlying single-crystal substrate occurs well below the melt temperature by the process of solid phase epitaxial growth (SPEG). Growth of crystalline Si(c-Si) is known to be enhanced by the presence of small amounts of a metallic phase, presumably due to an interaction of the free electrons of the metal with the covalent Si bonds near the growing interface. Ion implantation of Ni was shown to lower the crystallization temperature of an a-Si thin film by approximately 200°C. Using in situ transmission electron microscopy (TEM), precipitates of NiSi2 formed within the a-Si film during annealing, were observed to migrate, leaving a trail of epitaxial c-Si. High resolution TEM revealed an epitaxial NiSi2/Si(l11) interface which was Type A. We discuss here the enhanced nucleation of c-Si and subsequent silicide-mediated SPEG of Ni-implanted a-Si.Thin films of a-Si, 950 Å thick, were deposited onto Si(100) wafers capped with 1000Å of a-SiO2. Ion implantation produced sharply peaked Ni concentrations of 4×l020 and 2×l021 ions cm−3, in the center of the films.

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