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Past Issue in 2012
Volume: 2, Issue: 12
Biochemistry
An Improved Method for PAGE-based Detection of Phosphorylated Protein in Yeast
One dimensional polyacrylamide gel electrophoresis has been successfully used to detect protein phosphorylation. This method is very simple and highly reproducible. Hyperhosphorylated proteins usually migrate slowlier than dephosphorylated proteins. However, not all phosphorylated proteins can be readily detected, due to sub-optimal sample preparation and electrophoresis conditions. Here, an improved method is described that can detect phosphorylation of yeast proteins ranging from 15 kD to 200 kD. The improvement in gel electrophoresis should also be applicable to mammalian culture cells.
Cancer Biology
Cell Survival Rate Assay
This protocol utilizes PicoGreen 96-well plate technology. This method is applied to estimate the sensitivity of different tumor cell lines to chemodrugs.
Primary Tumor Preparation
This protocol has been developed for culturing primary glioblastoma cells. We have most experience in using it on rodent preparations, but it can also be used in culturing cells from other species.
Immunology
Generation of Mouse Bone Marrow-Derived Dendritic Cells (BM-DCs)
Generating mouse dendritic cells from bone-marrow progenitor cells is a useful tool to study biological functions of mouse dendritic cells. Dendritic cells are one of the major populations of phagocytes able to activate both innate and adaptive immune cells.
Generation of Mouse Bone Marrow-Derived Macrophages (BM-MFs)
Generating mouse macrophages from bone-marrow progenitor cells is a useful tool to study biological functions of mouse macrophages. Macrophages are one of the major populations of phagocytes and play many different roles during inflammatory process initiation and termination.
Quantitative Enzyme-linked Immunosorbent Assay (ELISA) to Measure Serum Levels of Murine Anti-cardiolipin Antibodies
The circulating anticardiolipin antibody is a hallmark of antiphospholipid syndrome. It also appears in a number of autoimmune mouse models and is indicative of the break of tolerance against self antigens. This protocol describes a reliable method to determine the relative serum titer of anticardiolipin in autoimmune mouse models.
Molecular Biology
A Simple RNA Preparation Procedure from Yeast for Northern Blot Using Hot Phenol
Compared to several expensive RNA extraction kits, the following protocol provides an economic and simple method for researchers to extract yeast RNA. This method can achieve RNA quality that is sufficient for most northern blot studies in yeast.