Protein lipid overlayLipid–protein interactionsPLOSchematic overview of the protein lipid overlay (PLO) assay. Schematic overview of the classical protein lipid overlay (CPLO) assay. Different types or concentrations of lipids are spotted onto a PVDF membrane, blocked with PBST blocking buffer ...
The quantification of the protein–lipid overlay assay revealed a strong and significant binding of GFP-mGBP2 to cardiolipin (CL), among other lipids that will be analyzed elsewhere. Cardiolipin represents a mitochondria-specific phospholipid, where it is mainly located in the inner membrane (up to...
The lipid overlay assay has been commonly used due to its convenience but it suffers from many drawbacks, including low sensitivity, poor reliability, and an inability to yield quantitative information (Dowler, Kular, & Alessi, 2002). Also, lipids are presented in a poorly defined, non...
(n = 3, biologically independent).kLBP lipid overlay assay using purified LBP with non-treatment or 1 h 1 mM H2O2treatment lipid-bound membranes.lDose-response curves for determination of Kd for the binding of LBP to tridocosahexaenoin using a labeled MST assay. Shown are means...
8A) and was subsequently used for protein lipid overlay assay. Two membranes pre-spotted with a variety of lipid species were probed with the apoL1.FLAG fusion protein. As expected, Fig. 8B showed that the apoL1.FLAG, but not 3× FLAG itself (negative control), bound to a variety of ...
Methods for transfection include calcium phosphate (Chen and Okayama, 1987; Wigler et al., 1977), lipid-mediated (Felgner et al., 1989; Felgner and Ringold, 1989) and electroporation (Chu et al., 1987; Shigekawa and Dower, 198...
Samples were prepared as described using the lipid-DNA-method [9, 10]. Briefly, DNA mixtures containing reporter and prey (for PR slides) or reporter, prey and bait constructs (for PRB slides) in EC buffer supplemented with 0.2 M sucrose were treated for 5 minutes at RT with 1.5 μl En...
While DNA and RNA methods are routine to disrupt the expression of specific genes, complete understanding of developmental processes requires also protein methods, because: oocytes and early embryos accumulate proteins and these are not directly affected
Methods for transfection include calcium phosphate (Chen and Okayama, 1987; Wigler et al., 1977), lipid-mediated (Felgner et al., 1989; Felgner and Ringold, 1989) and electroporation (Chu et al., 1987; Shigekawa and Dower, 198...
The brain slices were lysed, and total RNA was extracted using the RNeasy Lipid Tissue kit (Qiagen, France) according to the manufacturer’s instructions. The RNA (1 μg) was denatured for 10 min at 68°C, and cDNA was generated using reverse transcription with 200 nmol/L of dNTPs, 1...