scholarly journals Utilizing Time-Resolved Protein-Induced Fluorescence Enhancement to Identify Stable Local Conformations One α-Synuclein Monomer at a Time

10.3791/62655 ◽  
2021 ◽  
Author(s):  
Sofia Zaer ◽  
Eitan Lerner
Keyword(s):  
Fluorescence Enhancement ◽  
Time Resolved ◽  
Local Conformations

Two-site time-resolved immunofluorometric assay of human insulin.

1986 ◽  
Vol 32 (4) ◽  
pp. 637-640 ◽  
Author(s):  
E Toivonen ◽  
I Hemmilä ◽  
J Marniemi ◽  
P N Jørgensen ◽  
J Zeuthen ◽  
...  
Keyword(s):  
Human Insulin ◽  
Fluorescence Enhancement ◽  
Microtiter Plate ◽  
The Other ◽  
Plasma Samples ◽  
Serum Samples ◽  
Time Resolved ◽  
Standard Curve ◽  
Sandwich Type ◽  
Plate Strip

Abstract We describe a two-site "sandwich"-type time-resolved immunofluorometric assay for human insulin, based on use of two monoclonal antibodies with different specificities. The first antibody is immobilized on the surface of microtiter plate strip wells, the other is labeled with Eu3+. Serum samples can be assayed with one incubation step; two incubation steps are required when plasma samples are assayed. After the immunoreactions are complete, the bound fraction of Eu3+-label is quantified by dissociating it in a fluorescence-enhancement solution and measuring its fluorescence with a fluorometer with time-resolution. The sensitivity of the assay is 0.24 micro-int. units/mL. The standard curve is linear from 0.24 to 2400 micro-int. units/mL.

An alternative fluorescence enhancement solution for use in lanthanide-based time-resolved fluoroimmunoassays.

1987 ◽  
Vol 33 (12) ◽  
pp. 2292-2295 ◽  
Author(s):  
J A Keelan ◽  
J T France ◽  
P M Barling
Keyword(s):  
Fluorescence Enhancement ◽  
Fluorescence Emission ◽  
Lanthanide Complex ◽  
Fluorescence Yield ◽  
Assay Sensitivity ◽  
Time Resolved ◽  
Lanthanide Chelates ◽  
All Solutions ◽  
And Performance ◽  
Economical Alternative

Abstract Time-resolved fluoroimmunoassays that make use of lanthanide chelates as labels require the addition of an enhancement solution to elicit the formation of a fluorescent lanthanide complex. All solutions previously described are based on 2-naphthoyltrifluoroacetone (NTA), a beta-diketone. Currently, this compound is not commercially available. We report here the properties and performance of an enhancement solution prepared with a commercially available beta-diketone, thenoyltrifluoroacetone. Use of this solution in a commercial time-resolved fluoroimmunoassay gave results essentially identical to those obtained with the NTA-based solution, although fluorescence emission was approximately 27% lower. The lower fluorescence yield did not, however, significantly reduce assay sensitivity. We conclude that this solution represents a viable and highly economical alternative to the preparation currently in use, particularly for laboratories wishing to develop their own assays based on lanthanide fluorescence.

A new fluorescence enhancement solution for Europium-based time-resolved fluoroimmunoassays

1990 ◽  
Vol 5 (3) ◽  
pp. 207-212 ◽  
Author(s):  
Paolo Degan ◽  
Angelo Abbondandolo ◽  
Giorgio Montagnoli
Keyword(s):  
Fluorescence Enhancement ◽  
Time Resolved

Time-resolved immunofluorometric assay for the ovarian carcinoma-associated antigenic determinant CA 125 in serum.

1987 ◽  
Vol 33 (12) ◽  
pp. 2191-2194 ◽  
Author(s):  
O C Boerman ◽  
C M Thomas ◽  
M F Segers ◽  
P Kenemans ◽  
T Lövgren ◽  
...  
Keyword(s):  
Ovarian Carcinoma ◽  
Antigenic Determinant ◽  
Fluorescence Enhancement ◽  
Ca 125 ◽  
Serum Samples ◽  
Measuring Range ◽  
Time Resolved ◽  
Europium Chelate ◽  
Analytical Range ◽  
Automated Procedures

Abstract A time-resolved immunofluorometric assay (IFMA) is described for quantifying the ovarian carcinoma-associated antigenic determinant CA 125 in human serum. Monoclonal antibody to CA 125 is immobilized onto a microtiter well, and the same antibody labeled with a europium chelate is used as a tracer. After the immunoreaction the bound portion of the labeled antibody is quantified by dissociating the Eu3+ in a fluorescence-enhancement solution and measuring its fluorescence with a time-resolved fluorometer. The detection limit of the IFMA is 1.5 arb. units/mL, being about the same as that of a commercially available immunoradiometric assay (IRMA) for CA 125 (1.4 arb. units/mL). The analytical range of the IFMA extends to 2000 arb. units/mL, whereas the range of the IRMA is 500 arb. units/mL. For 29 serum samples from ovarian-cancer patients measured simultaneously in the IFMA and IRMA, orthogonal regression analysis gave the equation CA 125 (IFMA) = 0.9937 CA 125 (IRMA) - 1.211 arb. units/mL (Syx = 6.8681, r = 0.9932). Apparently, the IFMA for CA 125 is a convenient alternative to the IRMA for CA 125 because of short counting times, the use of nonradioactive, stable reagents, and the much-extended measuring range. Additionally, the microtiter format should lend itself to more fully automated procedures in laboratories doing many such analyses.

scholarly journals A dissociative fluorescence enhancement technique for one-step time-resolved immunoassays

2010 ◽  
Vol 399 (4) ◽  
pp. 1677-1682 ◽  
Author(s):  
Kaj R. Blomberg ◽  
Veli-Matti Mukkala ◽  
Harri H. O. Hakala ◽  
Pauliina H. Mäkinen ◽  
Mikko U. Suonpää ◽  
...  
Keyword(s):  
Fluorescence Enhancement ◽  
Time Resolved ◽  
Enhancement Technique ◽  
One Step

Double-label time-resolved immunofluorometry of lutropin and follitropin in serum.

1987 ◽  
Vol 33 (12) ◽  
pp. 2281-2283 ◽  
Author(s):  
I Hemmilä ◽  
S Holttinen ◽  
K Pettersson ◽  
T Lövgren
Keyword(s):  
Monoclonal Antibodies ◽  
Detection Limit ◽  
Specific Antibody ◽  
Fluorescence Enhancement ◽  
Trioctylphosphine Oxide ◽  
Time Resolved ◽  
Double Label ◽  
Capture Antibody ◽  
Triton X ◽  
Label Time

Abstract We describe a procedure for the simultaneous immunofluorometric assay of lutropin and follitropin in human serum, based on the use of monoclonal antibodies and of the fluorescent lanthanides Eu3+ and Tb3+. The alpha-chain-specific antibody was used as a common capture antibody on the surfaces of microtitration strips. The anti-beta-follitropin antibody was labeled with Tb3+, the anti-beta-lutropin antibody with Eu3+. After the immunoreactions had taken place, the bound fractions of the labels were dissociated in a fluorescence enhancement solution of pivaloyltrifluoroacetone, trioctylphosphine oxide, and Triton X-100 surfactant. In this solution both lanthanides can be measured successively with a time-resolved fluorometer. The detection limit of the assay is 0.1 int. unit/L for lutropin and 1 int. unit/L for follitropin. Results correlated well with those by commercial immunofluorometric assays and radioimmunoassays.

Simultaneous Quadruple-Label Fluorometric Immunoassay of Thyroid-Stimulating Hormone, 17 α-Hydroxyprogesterone, Immunoreactive Trypsin, and Creatine Kinase MM Isoenzyme in Dried Blood Spots

1992 ◽  
Vol 38 (10) ◽  
pp. 2038-2043 ◽  
Author(s):  
Y Y Xu ◽  
K Pettersson ◽  
K Blomberg ◽  
I Hemmilä ◽  
H Mikola ◽  
...  
Keyword(s):  
Creatine Kinase ◽  
Monoclonal Antibodies ◽  
Fluorescence Enhancement ◽  
Dried Blood Spots ◽  
Thyroid Stimulating Hormone ◽  
Lanthanide Ions ◽  
Time Resolved ◽  
Immunoreactive Trypsin ◽  
Rabbit Igg ◽  
Stimulating Hormone

Abstract We describe a quadruple-label fluorometric immunoassay for simultaneously measuring four analytes: thyroid-stimulating hormone (TSH), 17 alpha-hydroxyprogesterone (17 alpha-OHP), immunoreactive trypsin (IRT), and creatine kinase MM (CK-MM). The assay is based on immunoreagents labeled with four different lanthanide ions (Eu3+, Tb3+, Sm3+, and Dy3+), on dissociative fluorescence enhancement applying the principle of co-fluorescence, and on time-resolved fluorometry. The monoclonal anti-alpha-TSH and anti-IRT antibodies and the polyclonal anti-CK-MM antibody were labeled with Eu3+, Sm3+, and Dy3+, respectively; 17 alpha-OHP was labeled with Tb3+. The assay was performed in microtitration strip wells coated with a mixture of monoclonal antibodies against beta-TSH, IRT, and CK-MM and a polyclonal goat anti-rabbit IgG for capture of the rabbit anti-17 alpha-OHP antibodies. After completion of the immunoreactions, the bound fractions of the lanthanides were dissociated into the co-fluorescence enhancement solution, creating highly fluorescent chelates. The four lanthanide-specific signals were subsequently measured in a time-resolved fluorometer. The detection limits of the assay were 0.1 mIU/L for TSH, 2 nmol/L for 17 alpha-OHP, 2 micrograms/L for IRT, and 4 U/L for CK-MM.

Microtubule nucleation sequence studied by time-resolved x-ray scattering and electron microscopy

1983 ◽  
Vol 41 ◽  
pp. 766-767
Author(s):  
Eva-Maria Mandelkow ◽  
Eckhard Mandelkow ◽  
Joan Bordas
Keyword(s):  
Electron Microscopy ◽  
Dynamic Equilibrium ◽  
Time Resolved ◽  
X Ray ◽  
Additional Information ◽  
X Ray Scattering ◽  
Low Concentrations ◽  
Microtubule Growth ◽  
Ray Patterns ◽  
Ray Scattering

When a solution of microtubule protein is changed from non-polymerising to polymerising conditions (e.g. by temperature jump or mixing with GTP) there is a series of structural transitions preceding microtubule growth. These have been detected by time-resolved X-ray scattering using synchrotron radiation, and they may be classified into pre-nucleation and nucleation events. X-ray patterns are good indicators for the average behavior of the particles in solution, but they are difficult to interpret unless additional information on their structure is available. We therefore studied the assembly process by electron microscopy under conditions approaching those of the X-ray experiment. There are two difficulties in the EM approach: One is that the particles important for assembly are usually small and not very regular and therefore tend to be overlooked. Secondly EM specimens require low concentrations which favor disassembly of the particles one wants to observe since there is a dynamic equilibrium between polymers and subunits.

Electron Channeling Patterns

1977 ◽  
Vol 35 ◽  
pp. 12-13
Author(s):  
David C. Joy
Keyword(s):  
Electron Diffraction ◽  
Incidence Angle ◽  
Photographic Plate ◽  
Diffraction Effect ◽  
Angle Of Incidence ◽  
Bulk Single Crystal ◽  
Time Resolved ◽  
Electron Channeling ◽  
Spatially Resolved ◽  
Large Bulk

Electron channeling patterns (ECP) were first found by Coates (1967) while observing a large bulk, single crystal of silicon in a scanning electron microscope. The geometric pattern visible was shown to be produced as a result of the changes in the angle of incidence, between the beam and the specimen surface normal, which occur when the sample is examined at low magnification (Booker, Shaw, Whelan and Hirsch 1967).A conventional electron diffraction pattern consists of an angularly resolved intensity distribution in space which may be directly viewed on a fluorescent screen or recorded on a photographic plate. An ECP, on the other hand, is produced as the result of changes in the signal collected by a suitable electron detector as the incidence angle is varied. If an integrating detector is used, or if the beam traverses the surface at a fixed angle, then no channeling contrast will be observed. The ECP is thus a time resolved electron diffraction effect. It can therefore be related to spatially resolved diffraction phenomena by an application of the concepts of reciprocity (Cowley 1969).

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