scholarly journals Introduction of a PEGylated EPO‐conjugate as internal standard for EPO analysis in doping controls

2021 ◽  
Author(s):  
P. Reihlen ◽  
M. Blobel ◽  
P. Weiß ◽  
J. Harth ◽  
J. Wittmann ◽  
...  
Keyword(s):  
2018 ◽  
Vol 56 (11) ◽  
pp. 1905-1912 ◽  
Author(s):  
Kees J. Bronsema ◽  
Frank Klont ◽  
Frank B. Schalk ◽  
Rainer Bischoff ◽  
Ido P. Kema ◽  
...  

Abstract Background: Insulin-like growth factor 1 (IGF1) is a biomarker with various applications in medicine and also in doping control. Methods: A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed that employs 15N-IGF1 as an internal standard. The method features urea-based IGF1/IGFBP-complex dissociation which is directly followed by tryptic digestion. Following solid-phase extraction (SPE) sample clean-up of the digest, IGF1 is detected by means of two signature peptides that enable quantification of total IGF1 as well as discrimination between IGF1 proteoforms with ‘native’ and modified or extended N-terminal sequences. Results: Our method is capable of measuring plasma IGF1 concentrations over the clinically relevant range of 10–1000 ng/mL and was validated according to regulatory guidelines. Comparison with the IDS-iSYS IGF1 immunoassay revealed good correlation (R2>0.97) and no proportional bias between both assays was observed after normalizing the results against the WHO reference standard for IGF1 (02/254). Evaluation of several commercially available IGF1 preparations showed varying responses which were due to inconsistencies in purity and absolute amount of IGF1 present in these products. Conclusions: Our LC-MS/MS method introduces urea-based dissociation of IGF1/IGFBP-complexes to enable reliable quantification of IGF1 in plasma. Furthermore, the method is able to detect clinically relevant IGF1 levels without an enrichment procedure at the protein-level and thereby minimizes the risk of losing IGF1 proteoforms during sample preparation.


Steroids ◽  
2005 ◽  
Vol 70 (2) ◽  
pp. 103-110 ◽  
Author(s):  
Wolfgang Felzmann ◽  
Günter Gmeiner ◽  
Peter Gärtner

Steroids ◽  
2007 ◽  
Vol 72 (5) ◽  
pp. 429-436 ◽  
Author(s):  
Peter Gaertner ◽  
Katharina Bica ◽  
Wolfgang Felzmann ◽  
Guro Forsdahl ◽  
Günter Gmeiner

Author(s):  
Songquan Sun ◽  
Richard D. Leapman

Analyses of ultrathin cryosections are generally performed after freeze-drying because the presence of water renders the specimens highly susceptible to radiation damage. The water content of a subcellular compartment is an important quantity that must be known, for example, to convert the dry weight concentrations of ions to the physiologically more relevant molar concentrations. Water content can be determined indirectly from dark-field mass measurements provided that there is no differential shrinkage between compartments and that there exists a suitable internal standard. The potential advantage of a more direct method for measuring water has led us to explore the use of electron energy loss spectroscopy (EELS) for characterizing biological specimens in their frozen hydrated state.We have obtained preliminary EELS measurements from pure amorphous ice and from cryosectioned frozen protein solutions. The specimens were cryotransfered into a VG-HB501 field-emission STEM equipped with a 666 Gatan parallel-detection spectrometer and analyzed at approximately −160 C.


Author(s):  
R.D. Leapman ◽  
S.Q. Sun ◽  
S-L. Shi ◽  
R.A. Buchanan ◽  
S.B. Andrews

Recent advances in rapid-freezing and cryosectioning techniques coupled with use of the quantitative signals available in the scanning transmission electron microscope (STEM) can provide us with new methods for determining the water distributions of subcellular compartments. The water content is an important physiological quantity that reflects how fluid and electrolytes are regulated in the cell; it is also required to convert dry weight concentrations of ions obtained from x-ray microanalysis into the more relevant molar ionic concentrations. Here we compare the information about water concentrations from both elastic (annular dark-field) and inelastic (electron energy loss) scattering measurements.In order to utilize the elastic signal it is first necessary to increase contrast by removing the water from the cryosection. After dehydration the tissue can be digitally imaged under low-dose conditions, in the same way that STEM mass mapping of macromolecules is performed. The resulting pixel intensities are then converted into dry mass fractions by using an internal standard, e.g., the mean intensity of the whole image may be taken as representative of the bulk water content of the tissue.


2018 ◽  
Vol 84 (12) ◽  
pp. 5-19
Author(s):  
D. N. Bock ◽  
V. A. Labusov

A review of publications regarding detection of non-metallic inclusions in metal alloys using optical emission spectrometry with single-spark spectrum registration is presented. The main advantage of the method - an extremely short time of measurement (~1 min) – makes it useful for the purposes of direct production control. A spark-induced impact on a non-metallic inclusion results in a sharp increase (flashes) in the intensities of spectral lines of the elements that comprise the inclusion because their content in the metal matrix is usually rather small. The intensity distribution of the spectral line of the element obtained from several thousand of single-spark spectra consists of two parts: i) the Gaussian function corresponding to the content of the element in a dissolved form, and ii) an asymmetric additive in the region of high intensity values ??attributed to inclusions. Their quantitative determination is based on the assumption that the intensity of the spectral line in the single-spark spectrum is proportional to the content of the element in the matter ablated by the spark. Thus, according to the calibration dependence constructed using samples with a certified total element content, it is possible not only to determine the proportions of the dissolved and undissolved element, but also the dimensions of the individual inclusions. However, determination of the sizes is limited to a range of 1 – 20 µm. Moreover, only Al-containing inclusions can be determined quantitatively nowadays. Difficulties occur both with elements hardly dissolved in steels (O, Ca, Mg, S), and with the elements which exhibit rather high content in the dissolved form (Si, Mn). It is also still impossible to determine carbides and nitrides in steels using C and N lines. The use of time-resolved spectrometry can reduce the detection limits for inclusions containing Si and, possibly, Mn. The use of the internal standard in determination of the inclusions can also lower the detection limits, but may distort the results. Substitution of photomultipliers by solid-state linear radiation detectors provided development of more reliable internal standard, based on the background value in the vicinity of the spectral line. Verification of the results is difficult in the lack of standard samples of composition of the inclusions. Future studies can expand the range of inclusions to be determined by this method.


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