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Serbian Journal of Experimental and Clinical Research
2020, vol. 21, br. 4, str. 307-315
Fiksacija humanih zdravih autopsijskih tkiva formalinom - uticaj vrste tkiva, temperature i vremena inkubacije na kvalitet izolovane DNK
Todorović Danijelaa ORCID, Vitošević Katarinab ORCID, Todorović Milošb ORCID, Slović Živanab ORCID
aUniverzitet u Kragujevcu, Fakultet medicinskih nauka, Srbija
bUniverzitet u Kragujevcu, Fakultet medicinskih nauka, Katedra za anatomiju i sudsku medicinu, Srbija

e-adresa: dtodorovic@medf.kg.ac.rs
Projekat:
This study was funded by Faculty of Medical Sciences, University of Kragujevac, Serbia (grant number JP: 05/13)

Ključne reči: sudsko-medicinska obdukcija; formalin; izolovanje DNK; PCR; spektrofotometrija
Sažetak
Formalinska fiksacija je široko korišćena metoda u histopatologiji, koja ima i određena ograničenja. Formalin često dovodi do degradacije molekula DNK u tkivu tumora, što čini fiksirano tkivo neupotrebljivim za molekularnu analizu. I drugi faktori mogu uticati na kvalitet molekula DNK izolovanih iz fiksiranih tkiva. Cilj ovog rada je utvrditi uticaj dužine inkubacije tkiva u formalinu i uticaj temperature na kvalitet molekula DNK izolovanih iz različitih zdravih ljudskih tkiva. Tkiva mozga, pluća i bubrega, koja su izuzeta tokom sudsko-medicinskih obdukcija ljudi koji su umrli nasilnom smrću, fiksirana su u puferizovanom formalinu od 24h do dva meseca. Nakon isteka određenog inkubacionog perioda, iz tkiva je izolovana DNK ekstrakcijom pomoću fenol-hloroform-izoamil alkohola i spektrofotometrijski je određena koncentracija i čistoća uzoraka. Stepen degradacije molekula DNK procenjen je PCR reakcijom, amplifikacijom fragmenata gena dužine 150bp (GPD1) i 262bp (b-actin). Najveća koncentracija, čistoća i očuvan integritet DNK dobijeni su iz tkiva mozga. Sa produženim vremenom inkubacije tkiva u formalinu koncentracija i integritet DNK opadaju u svim ispitivanim tkivima, posebno u mozgu, dok čistoća uzoraka ostaje konstantna. Takođe, fiksacija tkiva na +4°S doprinosi boljem kvalitetu izolovane DNK u odnosu na DNK izolovane iz tkiva fiksiranih na sobnoj temperaturi. Može se zaključiti da vrsta zdravih humanih tkiva, temperatura i vreme inkubacije tkiva u formalinu imaju značajan uticaj na koncentraciju, čistoću i integritet DNK tokom procesa fiksacije tkiva izuzetih tokom sudsko-medicinske obdukcije.
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O članku

jezik rada: engleski
vrsta rada: izvorni naučni članak
DOI: 10.2478/sjecr-2019-0020
primljen: 16.12.2018.
prihvaćen: 08.05.2019.
objavljen u SCIndeksu: 26.11.2021.
Creative Commons License 4.0

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