scholarly journals A liquid-phase two-site immunoradiometric assay for human prolactin

1984 ◽  
Vol 217 (1) ◽  
pp. 273-279 ◽  
Author(s):  
S C Hodgkinson ◽  
J Landon ◽  
P J Lowry
Keyword(s):  
Solid Phase ◽  
High Sensitivity ◽  
Immunoradiometric Assay ◽  
Specific Antibodies ◽  
Simultaneous Addition ◽  
Operating Range ◽  
Specific Effects ◽  
Rabbit Igg ◽  
Human Prolactin ◽  
Time Required

The development of a ‘two-site’ immunoradiometric assay for human prolactin (hPrl) is described. The assay is based on the addition of radio-iodinated sheep anti-hPrl immunoglobulin G (IgG) and rabbit anti-hPrl serum to standards and unknowns followed by 3 h incubation. The use of solid phase reagents was avoided in order to minimize non-specific effects and the time required for reactants to reach equilibrium. Instead, the separation of hPrl-bound and free labelled antibody is achieved by the addition of sheep anti-(rabbit IgG) serum which precipitates bound labelled antibody by complex formation with rabbit anti-hPrl antibodies which are also hPrl-bound. Varying the order of addition of specific antibodies had a pronounced effect on the ‘operating range’ and sensitivity of resultant assays. This was attributed to competition between labelled and unlabelled antibodies for binding sites on the hPrl molecule. The immunoradiometric assay employing ‘simultaneous addition’ of specific antibodies was compared to a ‘simultaneous addition’ hPrl radioimmunoassay developed using the same sheep antiserum as that used to prepare the radioiodinated sheep anti-hPrl IgG. This immunoradiometric assay is characterized by rapid equilibration of reactants, a wide ‘operating range’ (the precision of dose estimates was less than 10% over the range 8-10000 mU/l), and high sensitivity (2.6 mU/l, 13 pg). In contrast, the hPrl radioimmunoassay required an incubation of 18 h, demonstrated a much reduced ‘operating range’ (the precision of dose estimates was less than 10% only over the range 25-1500 mU/l) and reduced sensitivity (9.8 mU/l, 49 pg).

scholarly journals Development of a non-extracted ‘two-site’ immunoradiometric assay for corticotropin utilizing extreme amino- and carboxy-terminally directed antibodies

1984 ◽  
Vol 218 (3) ◽  
pp. 703-711 ◽  
Author(s):  
S C Hodgkinson ◽  
B Allolio ◽  
J Landon ◽  
P J Lowry
Keyword(s):  
Complex Formation ◽  
Confidence Interval ◽  
Immunoglobulin G ◽  
Solid Phase ◽  
High Sensitivity ◽  
Immunoradiometric Assay ◽  
Specific Effects ◽  
Rabbit Igg ◽  
Time Required ◽  
Wide Operating

The development of a ‘two-site’ immunoradiometric assay (i.r.m.a.) for the direct estimation of human corticotropin-(1-39)-peptide in plasma is described. The assay is based on the simultaneous addition of 125I-labelled sheep anti-(N-terminal corticotropin) IgG (immunoglobulin G) antibodies and rabbit anti-(C-terminal corticotropin) antiserum to standards and unknowns (0.5 ml) followed by 18h incubation. The use of solid-phase reagents was avoided in order to minimize non-specific effects and the time required for reactants to reach equilibrium. Instead, the separation of corticotropin-bound from free labelled antibody is achieved by the addition of sheep anti-(rabbit IgG) antiserum, which precipitates bound labelled antibody by complex-formation with rabbit anti-corticotropin antibodies, which are also hormone-bound. Several 125I-labelled sheep anti-(N-terminal corticotropin) IgG preparations were assessed in the i.r.m.a. Although each was derived from antisera raised to a thyroglobulin conjugate of synthetic corticotropin-(1-24)-peptide (Synacthen), purification of immunoglobulins before iodination by selective immunoadsorption resulted in preparations with distinct specificities which demonstrated marked differences in binding to intact human corticotropin-(1-39)-peptide. These preparations are compared in combination with two rabbit anti-(C-terminal corticotropin) antisera. A ‘two-site’ assay based on the use of 125I-labelled sheep anti-[corticotropin-(2-16)-peptide] IgG and rabbit anti-[corticotropin-(34-39)-peptide] antiserum was optimized, since steric inhibition of antibody binding was avoided with this combination and because the measurement of only intact human corticotropin-(1-39)-peptide and not fragments was assured by the use of terminal antibodies. This i.r.m.a. is characterized by rapid equilibration of reactants, a wide ‘operating range’ (the precision of dose estimates was less than 4% over the range 30-2200 pg/ml) and high sensitivity [8 pg of corticotropin/ml (95% confidence interval 3.7-12.0) (4 pg minimal detectable mass) can be detected directly in plasma].

The Relation Between VIIIR:AG and VIII:C Studied with Two Antisera

1979 ◽  
Author(s):  
H. P. Muller ◽  
N. H. van Tilburg ◽  
R. M. Bertina ◽  
J. J. Veltkamp
Keyword(s):  
Molecular Weight ◽  
High Salt ◽  
Low Molecular Weight ◽  
Gel Chromatography ◽  
Specific Antibodies ◽  
Specific Effects ◽  
Normal Plasma ◽  
Coagulant Activity ◽  
Sterical Hindrance ◽  
Inhibitory Antibodies

FVIII was separated into low molecular weight FVIII (LMW FVIII) and high molecular weight FVIII (HMW FVIII) by gel chromatography in the presence of high salt concentration or by high salt elution of LMW FVIII from FVIII bound to anti HMW FVII-Sepharose. Specific antibodies were raised in rabbits against HMW FVIII and LMW FVIII. After removal of the contaminating anti HMW activities the rabbit anti LMW FVIII was still able to neutralize the FVIII coagulant activity of normal plasma and of IMW FVIII with canparable efficiency and it had no effect on the VIIIR:WF of FVIII in normal plasma or in HMW FVIII. Anti LMW FVIII does not bind to HMW FVIII and does not precipitate FVIII as tested by counter immunoelectrophoresis. Rabbit anti HMW FVIII precipitates FVIII in normal plasma, inhibits VIIIR:WF activity, while it has no effect on the FVIII coagulant activity of LMW FVIII. The coagulant activity of FVIII in normal plasma is slightly inhibited by anti HMW FVIII presumably by non-specific effects (sterical hindrance). It is concluded that inhibitory antibodies against VIII:C raised in rabbits recognize antigenic structures only present on LMW FVIII. Antibodies against HMW FVIII raised in rabbits appears to recognize structures only present on HMW FVIII.

scholarly journals A Single LC-MS/MS Analysis to Quantify CoA Biosynthetic Intermediates and Short-Chain Acyl CoAs

Metabolites ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 468
Author(s):  
Anthony E. Jones ◽  
Nataly J. Arias ◽  
Aracely Acevedo ◽  
Srinivasa T. Reddy ◽  
Ajit S. Divakaruni ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
Biosynthetic Pathway ◽  
Solid Phase ◽  
High Sensitivity ◽  
Short Chain ◽  
Extraction Column ◽  
Chain Acyl ◽  
Precision And Accuracy ◽  
Mass Spectrometry Mass Spectrometry

Coenzyme A (CoA) is an essential cofactor for dozens of reactions in intermediary metabolism. Dysregulation of CoA synthesis or acyl CoA metabolism can result in metabolic or neurodegenerative disease. Although several methods use liquid chromatography coupled with mass spectrometry/mass spectrometry (LC-MS/MS) to quantify acyl CoA levels in biological samples, few allow for simultaneous measurement of intermediates in the CoA biosynthetic pathway. Here we describe a simple sample preparation and LC-MS/MS method that can measure both short-chain acyl CoAs and biosynthetic precursors of CoA. The method does not require use of a solid phase extraction column during sample preparation and exhibits high sensitivity, precision, and accuracy. It reproduces expected changes from known effectors of cellular CoA homeostasis and helps clarify the mechanism by which excess concentrations of etomoxir reduce intracellular CoA levels.

A new photometric method for oxygen consumption measurements in cell suspensions

1986 ◽  
Vol 61 (2) ◽  
pp. 449-455 ◽  
Author(s):  
W. Mueller-Klieser ◽  
R. Zander ◽  
P. Vaupel
Keyword(s):  
Volume Fraction ◽  
High Sensitivity ◽  
Cell Suspensions ◽  
Measuring System ◽  
Photometric Method ◽  
O2 Consumption ◽  
Physical Density ◽  
O2 Concentration ◽  
Consumption Rates ◽  
Time Required

A new technique is described for measuring O2 consumption rates and O2 concentrations in suspensions of respiring cells. Aliquots of a cell suspension kept in a special thermostated precision syringe are injected into the measuring system in defined time intervals. The O2 content of these samples is determined photometrically, as reported previously. The O2 consumption per cellular wet weight and/or per single cell can be calculated from the cell volume fraction, the physical density, the cell concentration in the suspension, and the time-dependent decline of the O2 concentration in the precision syringe. The minimum detectable amount of O2 is 0.1 microliter O2, which corresponds to 0.001 (vol/vol) of O2 if a 100-microliters sample of suspended cells is analyzed. Reproducibility of the O2 consumption measurement is 9% of the measured value. The advantages offered by this method are the straightforward calibration in absolute terms, the short time required for one analysis (2–6 min), a high sensitivity, the simultaneous determination of overall O2 concentration and O2 consumption rates in cell suspensions, and the great variability in the application.

Combustion Synthesis of La0.65Sr0.35MnO3 and its Sensing Properties for NO2

2016 ◽  
Vol 680 ◽  
pp. 208-211
Author(s):  
Lian Lian Wu ◽  
Qiang Li ◽  
Dan Yu Jiang ◽  
Jin Feng Xia
Keyword(s):  
Solid Phase ◽  
High Sensitivity ◽  
Combustion Method ◽  
Fast Response ◽  
Size Analysis ◽  
Phase Method ◽  
X Ray Diffraction ◽  
Ultrafine Structure ◽  
Nox Sensor ◽  
Solid Phase Method

In this paper, La0.65Sr0.35MnO3 (LSM) oxide powder with ultrafine structure has been synthesized by self-propagating combustion method. The powders were characterized by X-ray diffraction, scanning electron microscopy and laser size analysis. Compared to the powders prepared by traditional solid-phase method, the grain size of powders prepared by self-propagating combustion method is relatively small and uniform. Starting from ultrafine LSM powders, sensing electrode (SE) for NO2 mixed-potential sensors based on yttria-stablized zirconia (YSZ) was fabricated. As-obtained NO2 sensor displays fast response and high sensitivity (25.4mV/decade). The response values of the sensor have good linear relationship with the logarithm of NO2 concentration varying from 30ppm to 500ppm.Keywords:Self-propagating combustion method; La0.65Sr0.35MnO3; NOx sensor; YSZ

High-sensitivity troponin assays: analytical and clinical aspects

2016 ◽  
Vol 51 (4) ◽  
pp. 315-320
Author(s):  
Magdalena Krintus
Keyword(s):  
Myocardial Infarction ◽  
Clinical Symptoms ◽  
High Sensitivity ◽  
Reference Population ◽  
Clinical Aspects ◽  
Analytical Sensitivity ◽  
Reference Limit ◽  
Highly Sensitive ◽  
Time Required ◽  
Highly Sensitive Troponin

Cardiac troponins are considered the most sensitive and specific biomarkers for the diagnosis of acute coronary syndromes (ACS). According to the Third Universal Definition of Acute Myocardial Infarction, the diagnosis requires a rise / or fall of troponin concentration with at least one value exceeding the 99th percentile upper reference limit (URL) in a reference population with the coexistence of clinical symptoms of ischemia. The introduction of highly sensitive assays has resulted in lower detection limits for the concentration of troponin, allowing for early diagnosis of, as well as the detection of quantifiable concentrations of this biomarker in healthy subjects. According to current guidelines, the use of high-sensitivity tests can shorten the time required to make clinical decisions from the current 3-6 hours to 1-2 hours. The use of highly sensitive troponin assays also carries other potential benefits associated with their predictive value, as well as challenges that include reduced specificity for myocardial infarction, lack of standardization or the presence of biological variability. Given the increasing availability of new, highly sensitive troponin assays we should be aware that their increased analytical sensitivity and precision is accompanied by accurate clinical assessment of the patient, and takes into account other non-cardiac causes of their increased concentrations.

scholarly journals IMMUNOREGULATION OF BLOOD SERUM ESTRADIOL AND PROGESTERONE LEVELS IN POSTMENOPAUSAL WOMEN

2019 ◽  
Vol 21 (5) ◽  
pp. 869-876
Author(s):  
A. N. Glushkov ◽  
E. G. Polenok ◽  
S. A. Mun ◽  
L. A. Gordeeva ◽  
V. A. Lutsenko ◽  
...  
Keyword(s):  
Blood Serum ◽  
Human Blood ◽  
Solid Phase ◽  
Biological Effects ◽  
Serum Levels ◽  
Human Blood Serum ◽  
Breast Cancer Patients ◽  
Specific Antibodies ◽  
Healthy Women ◽  
Antibody Levels

Specific antibodies against estradiol (Es) and progesterone (Pg) are known to modulate blood serum concentrations of these hormones and their biological effects after immunization of animals. It was suggested that specific IgA-Es and IgA-Pg could influence on Es and Pg levels in human blood serum. The purpose of this study was to identify the suggested correlations between serum Es and Pg and specific IgA-Es and IgA-Pg in postmenopausal healthy women (HW) and breast cancer patients (BCP). The serum levels of Es, Pg, IgA-Es and IgA-Pg were studied in 226 HW and 633 BCP by means of solid-phase immunoassay. The following results were obtained. The levels of Es in BCP (0.25 nmol/l) were higher than in HW (0.16; р < 0.0001). The levels of Pg were lower (0.79 vs 0.87; р < 0.0001), and individual Pg/Es ratios were lower (3.19 vs 6.64; р < 0.0001). Individual IgA-Pg/IgA-Es ratios correlated with decrease of Es (rs = -0.15; p = 0.029), with increase in Pg (rs = 0.38; р < 0.0001), and with increased Pg/Es ratio (rs = 0.29; р < 0.0001) in healthy women. Similar correlations were determined in BCP (correspondingly: rs = -0.14, р < 0.001; rs = 0.1, р = 0.009; rs = 0.15, р < 0.0001). The decrease of Es and increase of Pg and Pg/Es in BCP were less significant than in HW: the a quotients in regression у = ах+b (y = hormones levels and x = antibodies levels) in BCP were 3 to 4-fold lower than in HW. These peciliarities of interrelations between hormones and specific antibody levels were revealed only in ER+/PR+ BCP but not in ER+/PR- and ER-/PR- BCP. In conclusion, we have confirmed a suggestion about participation of specific antibodies in regulation of steroids levels in human blood serum. The immune regulation of hormonal status was weakened in BCP.

scholarly journals Greener Monolithic Solid Phase Extraction Biosorbent Based on Calcium Cross-Linked Starch Cryogel Composite Graphene Oxide Nanoparticles for Benzo(a)pyrene Analysis

Molecules ◽  
2021 ◽  
Vol 26 (20) ◽  
pp. 6163
Author(s):  
Aree Choodum ◽  
Nareumon Lamthornkit ◽  
Chanita Boonkanon ◽  
Tarawee Taweekarn ◽  
Kharittha Phatthanawiwat ◽  
...  
Keyword(s):  
Graphene Oxide ◽  
Solid Phase Extraction ◽  
Limit Of Detection ◽  
Solid Phase ◽  
High Sensitivity ◽  
Rice Flour ◽  
Oxide Nanoparticles ◽  
Phase Extraction ◽  
Limit Of Quantification ◽  
Significant Difference

Benzo(a)pyrene (BaP) has been recognized as a marker for the detection of carcinogenic polycyclic aromatic hydrocarbons. In this work, a novel monolithic solid-phase extraction (SPE) sorbent based on graphene oxide nanoparticles (GO) in starch-based cryogel composite (GO-Cry) was successfully prepared for BaP analysis. Rice flour and tapioca starch (gel precursors) were gelatinized in limewater (cross-linker) under alkaline conditions before addition of GO (filler) that can increase the ability to extract BaP up to 2.6-fold. BaP analysis had a linear range of 10 to 1000 µgL−1 with good linearity (R2 = 0.9971) and high sensitivity (4.1 ± 0.1 a.u./(µgL−1)). The limit of detection and limit of quantification were 4.21 ± 0.06 and 14.04 ± 0.19 µgL−1, respectively, with excellent precision (0.17 to 2.45%RSD). The accuracy in terms of recovery from spiked samples was in the range of 84 to 110% with no significant difference to a C18 cartridge. GO-Cry can be reproducibly prepared with 2.8%RSD from 4 lots and can be reused at least 10 times, which not only helps reduce the analysis costs (~0.41USD per analysis), but also reduces the resultant waste to the environment.

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