Evaluating RNA Preparation Options for Archived Myocardial Biopsies

Ip, W.; Huggins, Catherine E.; Pepe, Salvatore; Delbridge, L.

doi:10.1016/j.hlc.2011.01.023

Cited by 3 publications

(2 citation statements)

References 11 publications

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“…To the best of our knowledge, few similar systematic analyses have been performed on other cardiovascular clinical tissue sample types. Ip et al compared an organic extraction method with a solid-phase extraction method on archived right atrial appendage samples,[50] however these samples are different in size and composition from the more clinically-relevant left ventricular biopsies obtained in the ARCADIA study.…”

Section: Discussionmentioning

confidence: 99%

Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

et al. 2019

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This study aimed to optimise techniques for whole transcriptome and small RNA analyses on clinical tissue samples from patients with cardiovascular disease. Clinical samples often represent a particular challenge to extracting RNA of sufficient quality for robust RNA sequencing analysis, and due to availability, it is rarely possible to optimise techniques on the samples themselves. Therefore, we have used equivalent samples from pigs undergoing cardiopulmonary bypass surgery to test different protocols for optimal RNA extraction, and then validated the protocols in human samples. Here we present an assessment of the quality and quantity of RNA obtained using a variety of commercially-available RNA extraction kits on both left ventricular biopsies and blood plasma. RNA extraction from these samples presents different difficulties; left ventricular biopsies are small and fibrous, while blood plasma has a low RNA content. We have validated our optimised extraction techniques on human clinical samples collected as part of the ARCADIA (Association of non-coding RNAs with Coronary Artery Disease and type 2 Diabetes) cohort study, resulting in successful whole transcriptome and small RNA sequencing of human left ventricular tissue.

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Section: Discussionmentioning

confidence: 99%

Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

et al. 2019

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“…Dividing the muscle after dissection to source different applications is the best option in many cases. However, the muscle pieces are often too small to use pulverization under liquid nitrogen or mechanical grinding techniques for homogenization [2][3][4][5] . As muscle is a highly structural tissue packed with extracellular matrix and contractile proteins, inadequate mechanical homogenization leads to a low yield of subsequent DNA, RNA or protein.…”

Section: Introductionmentioning

confidence: 99%

Cryosectioning of Contiguous Regions of a Single Mouse Skeletal Muscle for Gene Expression and Histological Analyses

Beedle

2016

JoVE

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With this method, consecutive cryosections are collected to enable both microscopy applications for tissue histology and enrichment of RNA for gene expression using adjacent regions from a single mouse skeletal muscle. Typically, it is challenging to achieve adequate homogenization of small skeletal muscle samples because buffer volumes may be too low for efficient grinding applications, yet without sufficient mechanical disruption, the dense tissue architecture of muscle limits penetration of buffer reagents, ultimately causing low RNA yield. By following the protocol reported here, 30 μm sections are collected and pooled allowing cryosectioning and subsequent needle homogenization to mechanically disrupt the muscle, increasing the surface area exposed for buffer penetration. The primary limitations of the technique are that it requires a cryostat, and it is relatively low throughput. However, high-quality RNA can be obtained from small samples of pooled muscle cryosections, making this method accessible for many different skeletal muscles and other tissues. Furthermore, this technique enables matched analyses (e.g., tissue histopathology and gene expression) from adjacent regions of a single skeletal muscle so that measurements can be directly compared across applications to reduce experimental uncertainty and to reduce replicative animal experiments necessary to source a small tissue for multiple applications.

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A Small Cohort Omega-3 PUFA Supplement Study: Implications of Stratifying According to Lipid Membrane Incorporation in Cardiac Surgical Patients

Chandramouli

Smith

et al. 2017

Heart, Lung and Circulation

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Evaluating RNA Preparation Options for Archived Myocardial Biopsies

Cited by 3 publications

References 11 publications

Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

Cryosectioning of Contiguous Regions of a Single Mouse Skeletal Muscle for Gene Expression and Histological Analyses

A Small Cohort Omega-3 PUFA Supplement Study: Implications of Stratifying According to Lipid Membrane Incorporation in Cardiac Surgical Patients

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