Tgen was the third best performing stock this
past week--in the list of regional stocks in
the Seattle Times. At just under a buck a share,
the volume does not suggest that any real
money is changing hands, but it is nice to see a
ray of hope in a crummy market.
And here are a couple of 2002 abstracts from Pubmed,
the second one has an author who went to tgen, fwiw.
Biotechnol Appl Biochem 2002 Aug 1;36(1):13-20
Quantitative physical characterization of lipid-polycation-DNA lipopolyplexes.
Tsai JT, Furstoss KJ, Michnick T, Sloane DL, Paul RW.
Targeted Genetics Corporation, 1100 Olive Way, Suite 100, Seattle, WA 98101, U.S.A.
Quantitative assays for the characterization of multi-component lipopolyplexes and their individual constituents are crucial for determining the consistency of formulation protocols which are ultimately reflected in biological activity. Lipid-polycation-DNA formulations consisting of lipids, polycations and DNA are of interest because they have been demonstrated to be efficient gene-delivery vehicles when administered systemically. We have developed a panel of analytical techniques to characterize these lipopolyplexes. Complexes were measured for size by dynamic light scattering and surface-charge characteristics by zeta potential. Interaction between DNA and the polycation, protamine sulphate, was determined using a PicoGreen dye-exclusion technique. Total DNA in the lipopolyplex was assayed through decomplexation of the formulation by addition of heparin sulphate and subsequent DNA quantification by PicoGreen reagent. Protamine sulphate in the lipopolyplex was determined using a novel Amido Black-staining protocol which is linearly sensitive in a range of 0.25-3 &mgr;g of protein. Lipids were quantified by HPLC after extraction in chloroform/methanol (2:1). In this method elution is conducted over 40 min, with 1,2-dioleoyl-3-trimethylammonium-propane and cholesterol being resolved by greater than 10 min. Such assays are essential for product characterization and release tests, as well as development of a better understanding of the correlation between physical structure and biological function.
PMID: 12149118
Int Immunol 2002 Apr;14(4):389-400
Expression and characterization of recombinant soluble human CD3 molecules: presentation of antigenic epitopes defined on the native TCR-CD3 complex.
Law CL, Hayden-Ledbetter M, Buckwalter S, McNeill L, Nguyen H, Habecker P, Thorne BA, Dua R, Ledbetter JA.
Xcyte Therapies, Inc., Seattle, WA 98104, USA. Pacific Northwest Research Institute, 720 Broadway, Seattle, WA 98122, USA. Present address: Seattle Genetics, Inc., Bothell, WA 98021, USA. Present address: Targeted Genetics Corp., Seattle, WA 98101, USA.
The TCR-CD3 complex consists of the clonotypic disulfide-linked TCRalphabeta or TCRdeltagamma heterodimers, and the invariant CD3delta, epsilon, gamma and zeta chains. We generated plasmid constructs expressing the extracellular domains of the CD3delta, epsilon or gamma subunits fused to human IgG1 Fc. Recombinant fusion proteins consisting of individual CD3delta, epsilon or gamma subunits reacted poorly with anti-CD3 mAb including G19-4, BC3, OKT3 and 64.1. Co-expression of the CD3epsilon-Ig with either the CD3delta-Ig (CD3epsilondelta-Ig) or the CD3gamma-Ig (CD3epsilongamma-Ig) resulted in fusion proteins with much increased binding to G19-4. A brief acid treatment of the purified CD3epsilondelta-Ig fusion protein substantially improved its binding to BC3, OKT3 and 64.1. Surface plasmon resonance analysis revealed that the dissociation constants for CD3epsilondelta-Ig and anti-CD3 mAb ranged from 10(-8) to 10(-9) M. Based on these results, a single-chain (sc) construct encoding the CD3delta chain linked to the CD3epsilon chain with a flexible linker followed by human IgG1 Fc was expressed. The sc CD3deltaepsilon-scIg reacted with anti-CD3 mAb without requiring acid treatment. Moreover, anti-CD3 mAb bound CD3epsilondelta-Ig at a higher affinity than CD3epsilongamma-Ig, suggesting potential structural differences between the CD3epsilondelta and CD3epsilongamma subunits. In summary, we report the expression of soluble recombinant CD3 proteins that demonstrate structural characteristics of the native CD3 complex expressed on the T cell surface. These CD3 fusion proteins can be used to further analyze the structure of the TCR-CD3 complex, and to identify molecules that can interfere with TCR-CD3-mediated signal transduction by disrupting the interaction between CD3 and TCR subunits.
PMID: 11934875 [PubMed - in process] |
