cardiovascularКардиоваскулярная терапия и профилактикаCardiovascular Therapy and Prevention1728-88002619-0125«SILICEA-POLIGRAF» LLC10.15829/1728-8800-2021-3105cardiovascular-3105Research ArticleБИОБАНКИРОВАНИЕBIOBANKINGОптимизация условий хранения РНК в биобанке, а также методов ручного и автоматизированного выделения РНК из цельной крови и лейкоцитарной фракцииOptimization of RNA storage in a biobank, as well as methods for manual and automated isolation of RNA from whole blood and leukocyte fractionhttps://orcid.org/0000-0002-4267-5941РоманюкС. А.RomanyukS. A.

Софья Андреевна Романюк — биолог.

Сестрорецк, SPIN: 6028-9050

Sestroretsk

sofya.romanyuk@gmail.comhttps://orcid.org/0000-0003-1778-0165ПоповО. С.PopovO. S.

Олег Сергеевич Попов — специалист.

Сестрорецк, SPIN: 5220-9174

Sestroretsk

ospopov@outlook.comhttps://orcid.org/0000-0002-5100-5229СушенцеваН. Н.SushentsevaN. N.

Наталья Николаевна Сушенцева — биолог.

Сестрорецк. Тел.: +7 (911) 159-83-03, SPIN: 5187-2286

Sestroretsk

navicula@yandex.ruhttps://orcid.org/0000-0002-3853-4185АпалькоС. В.ApalkoS. V.

Светлана Вячеславовна Апалько — кандидат биологических наук, биолог, заведующий лабораторией.

Сестрорецк,SPIN: 7053-2507

Sestroretsk

svetlana.apalko@gmail.comhttps://orcid.org/0000-0002-9178-8512ПолковниковаИ. А.PolkovnikovaI. A.

Ирина Андреевна Полковникова — биолог.

Сестрорецк, SPIN: 1081-6432

Sestroretsk

polkovnikovaia@gb40.ruhttps://orcid.org/0000-0001-5047-2792ЩербакС. Г.ShcherbakS. G.

Сергей Григорьевич Щербак — доктор медицинских наук, главный врач, профессор, заведующий кафедрой последипломного медицинского образования.

Сестрорецк; Санкт-Петербург, SPIN: 1537-9822

Sestroretsk; St. Petersburg

sgsherbak@mail.ruСПб ГБУЗ Городская больница № 40РоссияCity Hospital № 40Russian FederationСПб ГБУЗ Городская больница № 40; Санкт-Петербургский государственный университетРоссияCity Hospital № 40; Saint Petersburg State UniversityRussian Federation20210901202220831053105Copyright © Романюк С.А., Попов О.С., Сушенцева Н.Н., Апалько С.В., Полковникова И.А., Щербак С.Г., 20222022Романюк С.А., Попов О.С., Сушенцева Н.Н., Апалько С.В., Полковникова И.А., Щербак С.Г.Romanyuk S.A., Popov O.S., Sushentseva N.N., Apalko S.V., Polkovnikova I.A., Shcherbak S.G.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://cardiovascular.elpub.ru/jour/article/view/3105Цель

Цель. Оптимизация методики выделения и хранения рибонуклеиновой кислоты (РНК) из цельной крови и лейкоцитарной фракции.

Материалы и методы

Материалы и методы. Сравнение качества выделения проводилось для образцов РНК, полученных из 228 образцов лейкоцитарной фракции и 198 образцов цельной крови. Выделение было проведено из свежих и замороженных образцов с помощью реагента ExtractRNA™ и автоматизированной станции MagNA Pure Compact. Были опробованы различные способы удаления эритроцитов — с помощью центрифугирования и обработки гемолитиками двух производителей, а также способы заморозки материала для последующего выделения РНК — с консервантами и без них.

Результаты

Результаты. Была проверена 21 комбинация условий. Наиболее качественная РНК выделялась при ручной экстракции с использованием реагента ExtractRNA™ из свежей, очищенной гемолитиком “Амплисенс” лейкоцитарной фракции (успешность выделения — 94%, медиана RIN=8,4); замороженной в IntactRNA™, очищенной центрифугированием лейкоцитарной фракции (успешность выделения — 100%, медиана RIN =8); замороженной в ExtractRNA™, очищенной центрифугированием лейкоцитарной фракции (успешность выделения — 100%, медиана RIN =9,3).

Заключение

Заключение. Из замороженных фракций крови возможно выделение РНК, не уступающей по качеству выделенной из свежих образцов. Таким образом, проводить выделение РНК сразу после поступления биологического материала не обязательно.

Aim

Aim. To optimize the technique for the isolation and storage of ribonucleic acid (RNA) from whole blood and leukocyte fraction.

Materials and methods

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

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

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.

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The authors declare that there are no conflicts of interest present.