Fluoroimmunoassay of progesterone in human serum or plasma.

1981 ◽  
Vol 27 (7) ◽  
pp. 1176-1179 ◽  
Author(s):  
B L Allman ◽  
F Short ◽  
V H James
Keyword(s):  
Liquid Phase ◽  
Solid Phase ◽  
Clinical Use ◽  
Plasma Samples ◽  
Mixed Anhydride ◽  
Analytical Recovery ◽  
Covalently Linked ◽  
Solid Phase Assay ◽  
Antigen Antibody ◽  
Free Antigen

Abstract We describe a fluoroimmunoassay for progesterone in serum or plasma. The assay involves use of an antiserum to progesterone-11-hemisuccinate and a labeled antigen prepared by linking fluoresceinamine to progesterone-3-carboxymethyloxime by use of a mixed-anhydride procedure. Serum or plasma samples are extracted with hexane. After incubation of a portion of the evaporated extract with antiserum and labeled antigen, antibody-bound and free antigen are separated and the fluorescence of the bound fraction is measured. Separation is effected either by using ammonium sulfate to precipitate liquid-phase antiserum or by using a solid-phase antiserum: antiserum covalently linked to magnetizable cellulose particles, which are separated with a magnet. With either separation technique, analytical recovery, linearity, and precision were acceptable and results compared satisfactorily with those obtained with a conventional radioimmunoassay. The solid-phase assay is more precise and more technically convenient, but the performance of both fluoroimmunoassays was adequate for clinical use in the detection of ovulation.

scholarly journals A novel method for liquid-phase extraction of cell-free DNA for detection of circulating tumor DNA

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Filip Janku ◽  
Helen J. Huang ◽  
David Y. Pereira ◽  
Masae Kobayashi ◽  
Chung Hei Chiu ◽  
...  
Keyword(s):  
Liquid Phase ◽  
Solid Phase Extraction ◽  
Extraction Method ◽  
Solid Phase ◽  
Cancer Diagnostics ◽  
Phase Extraction ◽  
Plasma Samples ◽  
Cell Free Dna ◽  
Dna Yield ◽  
Free Dna

AbstractLow yields of extracted cell-free DNA (cfDNA) from plasma limit continued development of liquid biopsy in cancer, especially in early-stage cancer diagnostics and cancer screening applications. We investigate a novel liquid-phase-based DNA isolation method that utilizes aqueous two-phase systems to purify and concentrate circulating cfDNA. The PHASIFY MAX and PHASIFY ENRICH kits were compared to a commonly employed solid-phase extraction method on their ability to extract cfDNA from a set of 91 frozen plasma samples from cancer patients. Droplet digital PCR (ddPCR) was used as the downstream diagnostic to detect mutant copies. Compared to the QIAamp Circulating Nucleic Acid (QCNA) kit, the PHASIFY MAX method demonstrated 60% increase in DNA yield and 171% increase in mutant copy recovery, and the PHASIFY ENRICH kit demonstrated a 35% decrease in DNA yield with a 153% increase in mutant copy recovery. A follow-up study with PHASIFY ENRICH resulted in the positive conversion of 9 out of 47 plasma samples previously determined negative with QCNA extraction (all with known positive tissue genotyping). Our results indicate that this novel extraction technique offers higher cfDNA recovery resulting in better sensitivity for detection of cfDNA mutations compared to a commonly used solid-phase extraction method.

Interaction of mouse sperm with purified sperm receptors covalently linked to silica beads

1989 ◽  
Vol 92 (4) ◽  
pp. 713-722
Author(s):  
M.H. Vazquez ◽  
D.M. Phillips ◽  
P.M. Wassarman
Keyword(s):  
Zona Pellucida ◽  
Acrosome Reaction ◽  
Solid Phase ◽  
Galactose Oxidase ◽  
Flow Cytofluorometry ◽  
Mouse Sperm ◽  
Sperm Binding ◽  
Silica Beads ◽  
Covalently Linked ◽  
Solid Phase Assay

We describe a solid-phase assay that has permitted further analysis of zona pellucida glycoprotein, ZP3, as sperm receptor and acrosome reaction-inducer during fertilization in mice. The assay employs silica beads that contain epoxy groups to which purified, mouse oocyte ZP3 is covalently linked (ZP3-beads). ZP3-beads were characterized, e.g. by whole-mount autoradiography and flow cytofluorometry, incubated with capacitated mouse sperm under a variety of conditions, and the extent of sperm binding determined by light microscopy. Results of experiments presented suggest the following: (1) sperm bind specifically to ZP3-beads, but not to silica beads either exposed to 2-aminoethanol or derivatized with oocyte ZP2, fetuin or bovine serum albumin. (2) In nearly all cases, only one sperm binds per ZP3-bead and binding occurs via the sperm head. (3) The extent of sperm binding to ZP3-beads is dependent on ZP3 and sperm concentrations, as well as on incubation time and temperature. (4) Sperm binding to ZP3-beads is unaffected by antibodies directed against ZP3, but is inhibited in a reversible manner by treatment of ZP3-beads with galactose oxidase. (5) Only acrosome-intact sperm bind to ZP3-beads but, once bound, sperm can undergo the acrosome reaction, which results in their release from ZP3-beads. (6) Islet-activating protein and 3-quinuclidinyl benzilate, two inhibitors of the zona pellucida-induced acrosome reaction, prevent sperm bound to ZP3-beads from undergoing the acrosome reaction. These results confirm and extend previous studies of sperm-egg interaction in mice, and suggest that the solid-phase assay will be useful for both cellular and biochemical analyses of mammalian fertilization.

Rapid solid-phase immunoassay for 6-keto prostaglandin F1 alpha on microplates

1990 ◽  
Vol 36 (3) ◽  
pp. 509-514 ◽  
Author(s):  
W Schramm ◽  
R H Smith ◽  
T M Jackson ◽  
P A Craig ◽  
H E Grates ◽  
...  
Keyword(s):  
Magnetic Particles ◽  
Culture Medium ◽  
Solid Phase ◽  
Plasma Samples ◽  
Competitive Immunoassay ◽  
Immobilized Antibodies ◽  
Solid Phase Immunoassay ◽  
Hands On ◽  
Solid Phase Assay ◽  
Optimal Incubation

Abstract We describe, for the measurement of 6-keto prostaglandin F1 alpha in biological media, a solid-phase immunoassay with immobilized antibodies that requires a total processing time of less than 2 h with hands-on time less than 30 min for 40 samples. The method combines the convenience of the microplate format with the sensitivity of radiolabeled prostaglandin derivatives as tracers in a competitive immunoassay. The intra- and interassay variations at 50% displacement of the radiolabeled prostaglandin derivative as tracer were 9.0% and 11.8%, respectively. At 50% displacement of the radiolabeled tracer, the sensitivity is about 20 pg per well. Optimal incubation time is between 60 and 90 min. Nonspecific binding was less than 1% if about 8 pg of tracer (approximately 25,000 counts/min per well) was used. Inhibition curves of samples in different dilutions were parallel to standard curves. The variation of bound radiolabeled prostaglandin derivative within the wells of one microplate (n = 96) was less than 3%. Human plasma samples and medium from tissue culture assayed for 6-keto prostaglandin F1 alpha correlated well with results obtained with a solid-phase assay based on use of magnetic particles (r = 0.99, n = 24 for culture-medium samples; r = 0.99; n = 26 for plasma samples.

Living donor renal transplantation in the presence of donor-specific human leukocyte antigen antibody detected by solid-phase assay

2009 ◽  
Vol 70 (8) ◽  
pp. 584-588 ◽  
Author(s):  
Donna Phelan ◽  
T. Mohanakumar ◽  
Sabarinathan Ramachandran ◽  
Martin D. Jendrisak
Keyword(s):  
Renal Transplantation ◽  
Human Leukocyte Antigen ◽  
Living Donor ◽  
Solid Phase ◽  
Human Leukocyte ◽  
Leukocyte Antigen ◽  
Solid Phase Assay ◽  
Antigen Antibody

A monoclonal-antibody-based radioimmunoassay for measurement of protein C in plasma.

1988 ◽  
Vol 34 (2) ◽  
pp. 324-330 ◽  
Author(s):  
P R Howard ◽  
E G Bovill ◽  
K G Mann ◽  
R P Tracy
Keyword(s):  
Monoclonal Antibody ◽  
Protein C ◽  
Warfarin Therapy ◽  
Solid Phase ◽  
Functional Assay ◽  
Analytical Recovery ◽  
The Mean ◽  
Solid Phase Assay ◽  
Liquid Chromatographic

Abstract A monoclonal-antibody-based competitive radioimmunoassay for measuring human protein C is reported. With use of a purified protein C standard, the solid-phase assay was sensitive to less than 80 micrograms of protein C per liter. Intraassay CVs ranged from 5% to 8%; the inter-assay CV was 5.4%. Analytical recovery averaged 104% for purified protein C added to 10 samples of normal plasmas. The assay antibody could deplete plasma of all protein C, as determined by immunoaffinity chromatography followed by polyclonal Western blot analysis. Liquid-chromatographic gel permeation of plasma indicated a single immunoreactive species that had an appropriate molecular mass for monomeric protein C. Studies of monoclonal-antibody specificity showed no significant interferences by other vitamin K-dependent proteins. The mean protein C antigen concentration in plasma of 54 healthy subjects was 3.21 (SD 0.56) mg/L and was 1.51 (SD 0.52) mg/L for 22 patients on long-term warfarin therapy. Results of the monoclonal RIA correlated well with those by a polyclonal RIA also developed in our laboratory (r = 0.93) and an amidolytic functional assay (r = 0.88) for both normal plasma and plasma from patients on long-term warfarin therapy.

Development and Application of a Fully Automated Continuous Flow Radioimmunoassay System

1977 ◽  
Vol 14 (1-6) ◽  
pp. 1-11 ◽  
Author(s):  
Gordon C. Forrest
Keyword(s):  
Continuous Flow ◽  
Solid Phase ◽  
External Electromagnetic Field ◽  
New Approach ◽  
Flow Through ◽  
Uptake Test ◽  
Covalently Linked ◽  
Carry Over ◽  
Free Antigen ◽  
Gamma Counter

A fully automated, continuous flow radioimmunoassay system is described. The antibodies are covalently linked to a magnetic cellulose solid phase and separation of bound and free antigen is achieved by applying an external electromagnetic field. The bound fraction, after washing, is counted in a flow-through gamma counter chamber. The system, employing AutoAnalyzer modules, is highly reproducible and allows assays to be performed without equilibrium being reached, and only 10 minutes' incubation time is required. The method is universally applicable to the assay of ligands, either protein, hormone, or drug, and for obtaining information about binding proteins. A new approach to the latter has been developed and is illustrated by a fully automated thyroid hormone uptake test which gives better discrimination between normal and pathological states than existing assays. The system operates at 30 samples/h and is characterised by the high precision, speed, and minimal carry-over which can be achieved.

scholarly journals A Sensitive, Specific, Solid-Phase Enzymeimmunoassay for Plasma Progesterone

1981 ◽  
Vol 18 (1) ◽  
pp. 42-47 ◽  
Author(s):  
Brian G Joyce ◽  
Ann H Othick ◽  
Graham F Read ◽  
Diana Riad-Fahmy
Keyword(s):  
Solid Phase ◽  
World Health ◽  
Plasma Samples ◽  
Plasma Progesterone ◽  
Department Of Health ◽  
The United Kingdom ◽  
Freeze Dried ◽  
Enzyme Label ◽  
Covalently Linked ◽  
Health Organization

A homologous enzymeimmunoassay (EIA) for plasma progesterone, using a horseradish peroxidase conjugate as enzyme label and an antiserum raised against a progesterone-11α-hemisuccinyl/BSA conjugate, is described. The antiserum was covalently linked to microcrystalline cellulose to facilitate separation of bound and free steroid; this solid-phase antiserum was stable for at least nine months when stored at 4°C. The freeze-dried enzyme label is also stable, having retained both enzymic and immunological activity for about four years. The EIA developed was specific and had the sensitivity (4·8 pg/tube) required for determining progesterone concentrations in plasma samples collected at any time during the menstrual cycle. EIA of plasma samples provided results which were in good agreement with a well validated radioimmunoassay (RIA). The specificity and inter- and intra-assay coefficients of variation in the EIA were strictly comparable with those of the RIA. The method described has been in use for two years and has been assessed in external quality assurance programmes established by the World Health Organization and the United Kingdom Department of Health and Social Security.

A SOLID-PHASE RADIOIMMUNOASSAY FOR PROGESTERONE IN BOVINE PLASMA

1980 ◽  
Vol 60 (3) ◽  
pp. 783-786 ◽  
Author(s):  
G. de BOER ◽  
R. J. ETCHES ◽  
J. S. WALTON
Keyword(s):  
Aqueous Phase ◽  
Solid Phase ◽  
Plasma Samples ◽  
Bovine Plasma ◽  
Coefficients Of Variation ◽  
Solid Phase Radioimmunoassay ◽  
Highly Correlated ◽  
Solid Phase Assay ◽  
The Relationship

A solid-phase radioimmuniassay using antibody-coated tubes and 125I-labelled progesterone is described for determining concentrations of progesterone in unextracted bovine plasma. Samples were assayed for progesterone by both an aqueous-phase assay (X) using extracted plasma and the solid-phase assay (Y). The estimates of the concentration of progesterone using these two methods were highly correlated (r = 0.93) and the relationship between them was best described by the equation Y = 1.15X − 0.09. The slope and intercept of this equation were not significantly different from one and zero, respectively (P < 0.001). The technique is sufficiently sensitive to measure progesterone in 0.05 mL of plasma, is specific for progesterone, and has intra- and inter-assay coefficients of variation of 4.7 and 16.7%, respectively.

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.

SYNTHESIS OF HYDROGEN IN ACOUSTOPLASMA DISCHARGE IN A LIQUID-PHASE STREAM

2019 ◽  
pp. 42-48 ◽  
Author(s):  
N. A. Bulychev
Keyword(s):  
Liquid Phase ◽  
Organic Compounds ◽  
Electrode Materials ◽  
Solid Phase ◽  
Plasma Discharge ◽  
Raw Material ◽  
Two Phase ◽  
Reduced Pressure ◽  
External Power Source ◽  
Phase Medium

In this paper, the plasma discharge in a high-pressure fluid stream in order to produce gaseous hydrogen was studied. Methods and equipment have been developed for the excitation of a plasma discharge in a stream of liquid medium. The fluid flow under excessive pressure is directed to a hydrodynamic emitter located at the reactor inlet where a supersonic two-phase vapor-liquid flow under reduced pressure is formed in the liquid due to the pressure drop and decrease in the flow enthalpy. Electrodes are located in the reactor where an electric field is created using an external power source (the strength of the field exceeds the breakdown threshold of this two-phase medium) leading to theinitiation of a low-temperature glow quasi-stationary plasma discharge.A theoretical estimation of the parameters of this type of discharge has been carried out. It is shown that the lowtemperature plasma initiated under the flow conditions of a liquid-phase medium in the discharge gap between the electrodes can effectively decompose the hydrogen-containing molecules of organic compounds in a liquid with the formation of gaseous products where the content of hydrogen is more than 90%. In the process simulation, theoretical calculations of the voltage and discharge current were also made which are in good agreement with the experimental data. The reaction unit used in the experiments was of a volume of 50 ml and reaction capacity appeared to be about 1.5 liters of hydrogen per minute when using a mixture of oxygen-containing organic compounds as a raw material. During their decomposition in plasma, solid-phase products are also formed in insignificant amounts: carbon nanoparticles and oxide nanoparticles of discharge electrode materials.

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