We have developed a straightforward method for producing a stable, aqueous suspension of hydrophobic, fluorescent pi-conjugated polymer nanoparticles consisting primarily of individual conjugated polymer molecules. Features of the method are the facile preparation, purity, unique optical properties, and small size (approximately 5-10 nm) of the resulting nanoparticles. The results of TEM, scanning force microscopy, and near-field scanning optical microscopy of particles cast from the suspension indicate that the particles are single conjugated polymer molecules. The NSOM results yield estimates of the optical cross-sections of individual conjugated polymer molecules. The UV-vis absorption spectra of the nanoparticle suspensions indicate a reduction in conjugation length attributed to deformations of the polymer backbone. Fluorescence spectra of the aqueous nanoparticle suspensions indicate interactions between segments of the polymer chain and intramolecular energy transfer.
T cells recognize and kill a myriad of pathogen-infected or cancer cells using a diverse set of T cell receptors (TCR). The affinity of TCR to cognate antigen is of high interest in adoptive T cell transfer immunotherapy and antigen-specific T cell repertoire immune profiling because it is widely known to correlate with downstream T cell responses. Here, we introduce the in situ TCR affinity and sequence test (iTAST) for simultaneous measurement of TCR affinity and sequence from single primary CD8+ T cells in human blood. We demonstrate that the repertoire of primary antigen-specific T cells from pathogen inexperienced individuals has a surprisingly broad affinity range of 1000-fold composed of diverse TCR sequences. Within this range, samples from older individuals contained a reduced frequency of high affinity T cells compared to young individuals, demonstrating an age-related effect of T cell attrition that could cause holes in the repertoire. iTAST should enable the rapid selection of high affinity TCRs ex vivo for adoptive immunotherapy and measurement of T cell response for immune monitoring applications.
Escherichia coli is used intensively for recombinant
protein production, but one key challenge with recombinant E.
coli is the tendency of recombinant proteins to misfold and
aggregate into insoluble inclusion bodies (IBs). IBs contain high concentrations
of inactive recombinant protein that require recovery steps to salvage a
functional recombinant protein. Currently, no universally effective method
exists to prevent IB formation in recombinant E. coli. In this
study, DNA microarrays were used to compare the E. coli gene
expression response dynamics to soluble and insoluble recombinant protein
production. As expected and previously reported, the classical heat-shock genes
had increased expression due to IB formation, including protein folding
chaperones and proteases. Gene expression levels for protein synthesis-related
and energy-synthesis pathways were also increased. Many transmembrane
transporter and corresponding catabolic pathways genes had decreased expression
for substrates not present in the culture medium. Additionally, putative genes
represented over one-third of the genes identified to have significant
expression changes due to IB formation, indicating many important cellular
responses to IB formation still need to be characterized. Interestingly, cells
grown in 3% ethanol had significantly reduced gene expression responses
due to IB formation. Taken together, these results indicate that IB formation is
complex, stimulates the heat-shock response, increases protein and energy
synthesis needs, and streamlines transport and catabolic processes, while
ethanol diminished all of these responses.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.