Specificity grafting of human antibody frameworks selected from a phage display library: generation of a highly stable humanized anti-CD22 single-chain Fv fragment

A prerequisite for the enrichment of antibodies screened from phage display libraries is their stable expression on a phage during multiple selection rounds. Thus, if stringent panning procedures are employed, selection is simultaneously driven by antigen affinity, stability and solubility. To take advantage of robust pre-selected scaffolds of such molecules, we grafted single-chain Fv (scFv) antibodies, previously isolated from a human phage display library after multiple rounds of in vitro panning on tumor cells, with the specificity of the clinically established murine monoclonal anti-CD22 antibody RFB4. We show that a panel of grafted scFvs retained the specificity of the murine monoclonal antibody, bound to the target antigen with high affinity (6.4-9.6 nM), and exhibited exceptional biophysical stability with retention of 89-93% of the initial binding activity after 6 days of incubation in human serum at 37degreesC. Selection of stable human scaffolds with high sequence identity to both the human germline and the rodent frameworks required only a small number of murine residues to be retained within the human frameworks in order to maintain the structural integrity of the antigen binding site. We expect this approach may be applicable for the rapid generation of highly stable humanized antibodies with low immunogenic potential.

SHIMMER (1.0): a novel mathematical model for microbial and biogeochemical dynamics in glacier forefield ecosystems

SHIMMER (Soil biogeocHemIcal Model for Microbial Ecosystem Response) is a new numerical modelling framework designed to simulate microbial dynamics and biogeochemical cycling during initial ecosystem development in glacier forefield soils. However, it is also transferable to other extreme ecosystem types (such as desert soils or the surface of glaciers). The rationale for model development arises from decades of empirical observations in glacier forefields, and enables a quantitative and process focussed approach. Here, we provide a detailed description of SHIMMER, test its performance in two case study forefields: the Damma Glacier (Switzerland) and the Athabasca Glacier (Canada) and analyse sensitivity to identify the most sensitive and unconstrained model parameters. Results show that the accumulation of microbial biomass is highly dependent on variation in microbial growth and death rate constants, Q10 values, the active fraction of microbial biomass and the reactivity of organic matter. The model correctly predicts the rapid accumulation of microbial biomass observed during the initial stages of succession in the forefields of both the case study systems. Primary production is responsible for the initial build-up of labile substrate that subsequently supports heterotrophic growth. However, allochthonous contributions of organic matter, and nitrogen fixation, are important in sustaining this productivity. The development and application of SHIMMER also highlights aspects of these systems that require further empirical research: quantifying nutrient budgets and biogeochemical rates, exploring seasonality and microbial growth and cell death. This will lead to increased understanding of how glacier forefields contribute to global biogeochemical cycling and climate under future ice retreat.