PURPOSE: To assess the parameters for postmortem retinal tissue recovery and processing that affect the quality of RNA extracted from the retina/retinal pigment epithelium (RPE) complex. METHODS: RNA was extracted from retina/RPE samples. The RNA quality was determined based on qualitative/quantitative measurements made with a Bioanalyzer (Agilent) and on the expression of a long retinal gene (RPE65). After a pilot analysis on rats, ocular RNA was extracted from human donor eyeballs (group A) explanted according to conventional procedures for cornea transplantation. In a second experiment, another group of human donor eyeballs (group B) were processed in a much shorter time. The postmortem interval (T) comprised two periods: T1, the time between a donor's death and enucleation, and T2, the time between eyeball explantation and immersion of the excised retina/RPE sample in preservative solution (T = T1 + T2). RESULTS: A short T2 was correlated with good quality of RNA extracted from the retina/RPE complex (p = 0.043) and successful expression of a tissue-specific gene (p = 0.007). No other parameter appeared to influence RNA quality. CONCLUSIONS: The time between eyeball explantation and immersion of the retina/RPE sample in preservative solution was the chief parameter affecting the quality of RNA extracted from the retina/RPE complex.
PURPOSE: To assess the parameters for postmortem retinal tissue recovery and processing that affect the quality of RNA extracted from the retina/retinal pigment epithelium (RPE) complex. METHODS: RNA was extracted from retina/RPE samples. The RNA quality was determined based on qualitative/quantitative measurements made with a Bioanalyzer (Agilent) and on the expression of a long retinal gene (RPE65). After a pilot analysis on rats, ocular RNA was extracted from humandonor eyeballs (group A) explanted according to conventional procedures for cornea transplantation. In a second experiment, another group of humandonor eyeballs (group B) were processed in a much shorter time. The postmortem interval (T) comprised two periods: T1, the time between a donor's death and enucleation, and T2, the time between eyeball explantation and immersion of the excised retina/RPE sample in preservative solution (T = T1 + T2). RESULTS: A short T2 was correlated with good quality of RNA extracted from the retina/RPE complex (p = 0.043) and successful expression of a tissue-specific gene (p = 0.007). No other parameter appeared to influence RNA quality. CONCLUSIONS: The time between eyeball explantation and immersion of the retina/RPE sample in preservative solution was the chief parameter affecting the quality of RNA extracted from the retina/RPE complex.
Authors: Davina J Hensman Moss; Michael D Flower; Kitty K Lo; James R C Miller; Gert-Jan B van Ommen; Peter A C 't Hoen; Timothy C Stone; Amelia Guinee; Douglas R Langbehn; Lesley Jones; Vincent Plagnol; Willeke M C van Roon-Mom; Peter Holmans; Sarah J Tabrizi Journal: Sci Rep Date: 2017-03-21 Impact factor: 4.379
Authors: Patrice E Fort; Thekkelnaycke M Rajendiran; Tanu Soni; Jaeman Byun; Yang Shan; Helen C Looker; Robert G Nelson; Matthias Kretzler; George Michailidis; Jerome E Roger; Thomas W Gardner; Steven F Abcouwer; Subramaniam Pennathur; Farsad Afshinnia Journal: JCI Insight Date: 2021-10-08
Authors: Elke K Markert; Holger Klein; Coralie Viollet; Werner Rust; Benjamin Strobel; Stefan G Kauschke; Bar Makovoz; Heike Neubauer; Remko A Bakker; Timothy A Blenkinsop Journal: Front Cell Dev Biol Date: 2022-08-26