How can I increase the transduction efficiency with the BacMam 2.0 reagents such as the the CellLight and Premo products?
Try varying particle-to-cell ratio (PPC), incubation volume, temperature and, cell density (if adherent cells are transduced). For adherent cells, we recommend a confluence of about 70%. Following the PPC, adjusting the volume is the next best parameter to change to optimize protein expression. If that doesn't work, you can also use the BacMam Enhancer Kit (Cat. No. B10107).
Dynamic colocalization microscopy to characterize intracellular trafficking of nanomedicines.
Authors:Vercauteren D, Deschout H, Remaut K, Engbersen JF, Jones AT, Demeester J, De Smedt SC, Braeckmans K,
Journal:ACS Nano
PubMed ID:21923168
'To gain a better understanding of intracellular processing of nanomedicines, we employed quantitative live-cell fluorescence colocalization microscopy to study endosomal trafficking of polyplexes in retinal pigment epithelium cells. A new, dynamic colocalization algorithm was developed, based on particle tracking and trajectory correlation, allowing for spatiotemporal characterization of internalized polyplexes in ... More
Polycations such as polyethylenimine (PEI) are used in many novel nonviral vector designs and there are continuous efforts to increase our mechanistic understanding of their interactions with cells. Even so, the mechanism of polyplex escape from the endosomal/lysosomal pathway after internalization is still elusive. The ... More
Inhibitors of intravesicular acidification protect against Shiga toxin in a pH-independent manner.
Authors:Dyve Lingelem AB, Bergan J, Sandvig K,
Journal:Traffic
PubMed ID:22132807
Shiga toxin inhibits protein synthesis after being transported from the cell surface to endosomes and retrogradely through the Golgi apparatus to the endoplasmic reticulum (ER) and into the cytosol. In this study, we have abolished proton gradients across internal membranes in different ways and investigated the effect on the various ... More
Effective endogenous gene silencing mediated by pH responsive peptides proceeds via multiple pathways.
Authors:Lam JK, Liang W, Lan Y, Chaudhuri P, Chow MY, Witt K, Kudsiova L, Mason AJ,
Journal:J Control Release
PubMed ID:22138072
Cationic amphipathic histidine rich peptides possess high plasmid DNA and siRNA delivery capabilities. To further understand the pH responsive siRNA delivery process and evaluate the capabilities of such peptides we have investigated their ability to mediate specific silencing of endogenous GAPDH gene activity in MCF-7 and A549 cells and compared ... More
Engineering of blended nanoparticle platform for delivery of mitochondria-acting therapeutics.
Authors:Marrache S, Dhar S,
Journal:Proc Natl Acad Sci U S A
PubMed ID:22991470
Mitochondrial dysfunctions cause numerous human disorders. A platform technology based on biodegradable polymers for carrying bioactive molecules to the mitochondrial matrix could be of enormous potential benefit in treating mitochondrial diseases. Here we report a rationally designed mitochondria-targeted polymeric nanoparticle (NP) system and its optimization for efficient delivery of various ... More