Chromatographic methods for large-scale preparation of immunoglobulin G2a monoclonal antibodies Lym-1 and TNT-1 F(ab')2 fragments

Lym-1 and TNT-1 are two murine immunoglobulin G_2a monoclonal antibodies (MAbs) which have been used for clinical trials in cancer patients. This paper describes methods for large-scale preparation of F(ab')₂ fragments from 50 mg to 4 g of MAbs Lym-1 and TNT-1. Digestion of MAbs with pepsin was optimized and performed at pH 3.8, a pepsin/antibody ratio of 1:250, and 3–4 h of incubation at 37°C. The F(ab')₂ fragments were purified by tandem column procedures using fast protein liquid chromatography. Quality control analyses of the products included protein purity, isoelectric point, immunoreactivity, and endotoxin level. The results revealed that the chromatographic procedures are practical, simple, and effective, and can be used to produce gram quantities of clinical-grade F(ab')₂ fragments for the diagnosis of cancer in patients.     

Modest Interference with Actin Dynamics in Primary T Cell Activation by Antigen Presenting Cells Preferentially Affects Lamellal Signaling

Dynamic subcellular distributions of signaling system components are critical regulators of cellular signal transduction through their control of molecular interactions. Understanding how signaling activity depends on such distributions and the cellular structures driving them is required for comprehensive insight into signal transduction. In the activation of primary murine T cells by antigen presenting cells (APC) signaling intermediates associate with various subcellular structures, prominently a transient, wide, and actin-associated lamellum extending from an interdigitated T cell:APC interface several micrometers into the T cell. While actin dynamics are well established as general regulators of cellular organization, their role in controlling signaling organization in primary T cell:APC couples and the specific cellular structures driving it is unresolved. Using modest interference with actin dynamics with a low concentration of Jasplakinolide as corroborated by costimulation blockade we show that T cell actin preferentially controls lamellal signaling localization and activity leading downstream to calcium signaling. Lamellal localization repeatedly related to efficient T cell function. This suggests that the transient lamellal actin matrix regulates T cell signaling associations that facilitate T cell activation.     

BOOK REVIEWS

Sukopp, H. & Hejny, S. (eds.) & Kowarik, I. (co-ed). 1990. Urban Ecology. Plants and Plant Communities in Urban Environments. Short reviews Holten, J. I. (ed.) 1990. Effects of climate change on terrestrial ecosystems. Höner, D. 1991. Mehrjáhrige Beobachtungen kleiner Vegetationsfláchen im Raume von Karpathos (Nomos Dhodhekanisou, Griechenland).     

Comparative bacterial clearances of muscle and skin/subcutaneous tissues with and without dead bone: a laboratory study

Seventeen New Zealand White rabbits underwent implantation of three different concentrations of bacteria and a sterile saline control solution with and without dead autologous bone in eight separate muscular and eight separate subcutaneous sites. Following a period of 1 week, each site was surgically explored and samples of tissue were taken for histology and quantitative culture. Results reveal that final bacterial concentrations in the subcutaneous sites were significantly lower than in the muscle sites (p less than or equal to 0.0001) for each concentration of bacteria, with and without dead bone. Dead bone resulted in very significantly greater bacterial concentrations in both subcutaneous and muscle sites. Clinically, these results indicate that a thorough bony wound debridement is more important than the type of tissue used to close the wound. Flap tissue should be selected with regard to the perfusion, contour, and appearance of the recipient site.