Immunohistochemistry Rat

Isolating RNA from tissues and paraffin-embedded tissue samples can be challenging due to cross-linking of biomolecules and fragmented nucleic acids. The best solution is to slice the tissues into smaller pieces and make a homogenate solution (using tissue homogenizer or grinding liquid nitrogen frozen samples) in presence of RNAse inhibitors. The homogenization process should be carried out on dry ice to maintain the integrity of RNA.

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 TRIzol Reagent to perform RNA isolation / purification Tissue - Mouse Retina

Get tips on using TRIzol Reagent to perform RNA isolation / purification Tissue - Human Retina

Get tips on using RNeasy Mini Kit to perform RNA isolation / purification Tissue - Mouse Retina

Get tips on using NucleoSpin® RNA Plus to perform RNA isolation / purification Tissue - Mouse Retina

Get tips on using LAMP-1 Antibody (H4A3) to perform Autophagy assay cell type - Proximal tubular cells (rPT)

Get tips on using AllPrep DNA/RNA Mini Kit to perform RNA isolation / purification Tissue - Human Retina

Get tips on using Gibco DMEM/F-12, HEPES to perform 3D Cell Culture Media hiPSC-derived retinal organoids

Get tips on using Anti-Beclin 1 antibody (ab62557) to perform Autophagy assay cell type - Proximal tubular cells (rPT)