scholarly journals Optimization of RNA storage in a biobank, as well as methods for manual and automated isolation of RNA from whole blood and leukocyte fraction

2022 ◽  
Vol 20 (8) ◽  
pp. 3105
Author(s):  
S. A. Romanyuk ◽  
O. S. Popov ◽  
N. N. Sushentseva ◽  
S. V. Apalko ◽  
I. A. Polkovnikova ◽  
...  

Aim. To optimize the technique for the isolation and storage of ribonucleic acid (RNA) from whole blood and leukocyte fraction.Materials and methods. Comparison of isolation quality was carried out for RNA samples obtained from 228 leukocyte samples and 198 whole blood samples. Isolation was performed from fresh and frozen samples using ExtractRNA™ reagent and a MagNA Pure Compact automated system. Various methods of removing erythrocytes (centrifugation and treatment with hemolytic agents from two manufacturers) were tested, as well as freezing with and without preservatives for subsequent RNA isolation.Results. Twenty-one combinations of conditions were tested. The highest quality RNA was isolated by manual extraction using the ExtractRNA™ reagent from a fresh leukocyte fraction, purified by the Amplisens hemolytic agent (successful extraction — 94%, median RIN=8,4); frozen in IntactRNA™, purified by leukocyte fraction centrifugation (successful extraction — 100%, median RIN=8); frozen in ExtractRNA™, purified by leukocyte fraction centrifugation (successful extraction — 100%, median RIN=9,3).Conclusion. RNA can be isolated from frozen blood fractions, which is not inferior in quality to that isolated from fresh samples. Thus, it is not necessary to isolate RNA immediately after the receipt of biological material.

2018 ◽  
Vol 1 (1) ◽  
Author(s):  
Michael A. Wiley, MS ◽  
Nathan J. Alves, PhD

Background and Hypothesis:  Thromboelastography is torsional method of assessing coagulation efficiency (e.g. rate of clot formation and maximum clot strength) by measuring the viscosity changes of blood under low-shear stress. Limited research has been performed to date on how centrifugation and storage of whole blood affects the coagulability when recombined. We hypothesized that centrifuged blood that was stored at its component-specific ideal temperatures would produce a clot with the same clotting characteristics as whole blood.  Experimental Design or Project Methods:  Whole blood samples were collected from healthy volunteers into citrated tubes and were assigned to the following conditions: whole at 25[Symbol]C, whole at 4[Symbol]C, and centrifuged and stored as RBCs, platelets, and plasma (at 4[Symbol]C, 25[Symbol]C, and -20[Symbol]C, respectively) and then recombined. TEG tracings for each condition were obtained over the course of three weeks. We also explored how freezing and thawing whole blood samples and its components affected clotting.  Results:  When compared to room temperature and refrigerated whole blood, separated blood maintains its normal physiologic clotting dynamics and kinetics for a longer period of time. Whole blood at 25[Symbol]C maintains maximum clot strength for approximately one week post draw, whereas freezing whole blood drastically reduces coagulability. Additionally, we discovered that there is no practical difference in the clotting parameters between the first blood draw and subsequent draws.  Conclusion and Potential Impact:  Our study demonstrates the viability of storing blood in its components and recombining them at a later date while minimally affecting its clotting ability, thereby potentially reducing wasted blood products and the need for multiple blood draws for coagulation studies. 


2010 ◽  
Vol 41 (02) ◽  
Author(s):  
N Shazi ◽  
A Böss ◽  
HJ Merkel ◽  
F Scharbert ◽  
D Hannak ◽  
...  

Separations ◽  
2021 ◽  
Vol 8 (6) ◽  
pp. 78
Author(s):  
Sevasti Karampela ◽  
Jessica Smith ◽  
Irene Panderi

An ever-increasing need exists within the forensic laboratories to develop analytical processes for the qualitative and quantitative determination of a broad spectrum of new psychoactive substances. Phenylethylamine derivatives are among the major classes of psychoactive substances available on the global market and include both amphetamine analogues and synthetic cathinones. In this work, an ultra-high-performance liquid chromatography-positive ion electrospray ionization tandem mass spectrometric method (UHPLC-ESI-MS/MS) has been developed and fully validated for the determination of 19 psychoactive substances, including nine amphetamine-type stimulants and 10 synthetic cathinone derivatives, in premortem and postmortem whole blood. The assay was based on the use of 1 mL premortem or postmortem whole blood, following solid phase extraction prior to the analysis. The separation was achieved on a Poroshell 120 EC-C18 analytical column with a gradient mobile phase of 0.1% formic acid in acetonitrile and 0.1% formic acid in water in 9 min. The dynamic multiple reaction monitoring used in this work allowed for limit of detection (LOD) and lower limit of quantitation (LOQ) values of 0.5 and 2 ng mL−1, respectively, for all analytes both in premortem and postmortem whole blood samples. A quadratic calibration model was used for the 12 quantitative analytes over the concentration range of 20–2000 ng mL−1, and the method was shown to be precise and accurate both in premortem and postmortem whole blood. The method was applied to the analysis of real cases and proved to be a valuable tool in forensic and clinical toxicology.


2021 ◽  
Vol 2 (1) ◽  
pp. 100311
Author(s):  
Daniella C. Terenzi ◽  
Ehab Bakbak ◽  
Justin Z. Trac ◽  
Mohammad Al-Omran ◽  
Adrian Quan ◽  
...  

Author(s):  
Polina A. Dyachenko Timoshina ◽  
Leonid E. Dolotov ◽  
Ekaterina N. Lazareva ◽  
Anastasiia A. Kozlova ◽  
Olga A. Inozemtseva ◽  
...  

1994 ◽  
Vol 42 (3) ◽  
pp. 231-241 ◽  
Author(s):  
C. Shenberg ◽  
S. Spiegel ◽  
S. Chaitchik ◽  
P. Jordan ◽  
M. Kitzis ◽  
...  

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