siRNA / miRNA gene silencing Human Primary Endometrial Stromal Cells hsa-miR-542-3p

As autophagy is a multi-step process which includes not just the formation of autophagosomes, but most importantly, flux through the entire system, including the degradation upon fusion with lysosomes, which makes it quite challenging for detection. There are several methods for detection in mammalian cells, including immunoblotting analysis of LC3 and p62 and detection of autophagosome formation/maturation by fluorescence microscopy, Currently, there is no single “gold standard” for determining the autophagic activity that is applicable in every experimental context, hence it is recommended to go for the combined use of multiple methods to accurately assess the autophagic activity in any given biological setting.

Get tips on using Lipofectamine® 2000 Transfection Reagent to perform siRNA / RNAi /miRNA transfection Mouse - HeLa cells Lipofectamine

RNA-Seq is a method to sequence RNA by applying Next Generation Sequencing (NGS). The quality of RNA is critical for the success of RNA-Seq. The integrity of RNA is measured by the RNA integrity number (RIN). RIN is computed from RNA electrophoresis and electropherogram profiles (the peak area of the 28S rRNA should be approximately twice the peak area of the 18S rRNA). If you get the RIN value lower than 7, the possibility of getting the low quality of RNA-seq data is high. To get a high quality RNA, it is better to work with fresh samples or snap-freeze the tissues in liquid nitrogen as quickly as possible and store them at -80°C until further use. Make sure designated areas and all your filter tips, microfuge tubes, plastic, and glassware are RNase-free.

An alternative to culture-based cell death detection is an assessment of other cell viability indicators using fluorescent dyes, including membrane potential and membrane integrity. Live/Dead assays differentiates live and dead cells using membrane integrity as a proxy for cell viability and are based on a fluorescent staining procedure followed by detection using flow cytometry. However, samples preparation for such flow cytometry-based techniques could be challenging. Cell harvesting by trypsinization, mechanical or enzymatic cell disaggregation from tissues, extensive centrifugation steps, may all lead to preferential loss of apoptotic cells. To overcome this strictly follow manufacturers instruction of the detection kit.

Get tips on using mirVana™ miRNA Isolation Kit, with phenol to perform RNA isolation / purification Tissue - Human Bone marrow

Get tips on using RNAqueous™ Total RNA Isolation Kit to perform RNA isolation / purification Cells - primary human lung fibroblasts

Get tips on using ARCTURUS® PicoPure® DNA Extraction Kit to perform RNA isolation / purification Cells - primary human melanocytes

Get tips on using miRNeasy Micro Kit to perform RNA isolation / purification Cells - primary rat cortical neurons

Get tips on using AmpFLSTR™ Identifiler™ PCR Amplification Kit to perform Cell line authentication Endometrial Adenocarcinoma cell line HEC-1B

Get tips on using AmpFLSTR™ Identifiler™ Plus PCR Amplification Kit to perform Cell line authentication Endometrial Cancer cell line KLE