Automated tension infiltrometer

An automated tension infiltrometer including a soil contacting base to which is mounted a porous plate for interfacing the infiltrometer with the soil to be analyzed. A Marriotte column is positioned in the base so that its open bottom end abuts the porous plate. A bubble tower is also positioned in the base and has a bubbling tube operatively connected between its interior and interior of the Marriotte column. The bubble tower is adjustable to provide variable tension to the Marriotte column. First and second transducers are positioned at the upper and lower parts of the Marriotte column and continuously measure pressure changes at those positions while water from the column infiltrates into the soil. By correlating these measurements, improved precision in measuring water level is achieved, which in turn allows improved results regarding deriving soil characteristic information

A Strategy and Case Study Example for Designing and Implementing Environmental Long-Term Monitoring at Legacy Management Sites

Environmental monitoring objectives of site owners, regulators, consultants, and scientists typically share the common elements of (1) cost management, (2) risk management, and (3) information management (Figure 1). Many site owners focus on minimizing monitoring costs while regulators typically focus on risk and regulatory compliance. Scientists and consultants typically provide information management in the form of spreadsheets with extracted information provided in reports to other users. This common piecemeal approach upon individual focus on elements of the monitoring objectives, rather than the common objective of minimizing cost and risk using site information, results in missed opportunities for cost savings, environmental protection, and improved understanding of site performance

Overland Flow Implications on Infiltration Along a Hill Slope

Water flow and solute transport on a hill slope is a complex nonlinear problem. Rainwater initially infiltrates at a rate equal to the rainfall rate. Once the soil infiltration capacity is reached, surface runoff is generated redistributing water along sloped surfaces. More water usually infiltrates at the lower parts of a hill slope, even for homogeneous soil profiles, because of generally longer surface ponding times and vegetation density. The variable infiltration along a hill slope has significant consequences for plant growth and the overall water balance of evapotranspiration covers. To describe these complex interactions we have coupled the HYDRUS-2D software package, simulating water flow and solute transport in variably saturated porous media, with a newly developed overland flow routine. The overland flow solver uses fully implicit four-point finite difference method to numerically solve the one-dimensional kinematic wave equation, with overland fluxes evaluated using Manning’s hydraulic resistance law. A Picard iterative solution scheme, similar to one used for solution of the Richards equation, is invoked to solve the resulting system of nonlinear equations. The subsurface flow module determines the main time step for the coupled system. If required for numerical stability, the overland flow module can use multiple smaller time steps during the main time step. This type of time management considers the fact that overland flow and variably-saturated subsurface flow often run at quite different time scales. We will present several relatively small-scale examples of the updated HYDRUS-2D program showing the development of overland flow as a function of storm intensity and slope angle. Simple examples will verify the correctness of the numerical implementation against an analytical solution. More complex examples will examine infiltration with and without the overland flow modifications along a hill slope. The interaction of runon, vegetative growth, and permeability changes will be examined through these simulations

Metabolic exploitation of the sialic acid biosynthetic pathway to generate site-specifically labeled antibodies.

Lack of a universal site-specific conjugation methodology for antibodies limits their potential to be developed as tumor-specific imaging agents or targeted therapeutics. A potential mechanism for site-specific conjugation involves utilization of the conserved N-glycosylation site in the CH2 domain. We sought to develop an antibody with an altered azido-sugar at this site whereby site-specific label could be added. The HB8059 hybridoma was cultured with peracetylated N-azidoacetlymannosamine (Ac4ManNAz). The resulting azido-sugar antibody was conjugated to phosphine-polyethylene glycol (PEG3)-biotin via a modified Staudinger reaction. Biochemical and functional characterization of the biotinylated antibody was performed. The azido-sugar antibody was also labeled with DyLight-650-Phosphine and injected into mice harboring pancreatic cancer xenografts. The tumors were dissected and imaged utilizing an IVIS fluorescent camera. The antibody was successfully produced in 100 μM Ac4ManNAz. The biotinylated antibody demonstrated a 50 kDa heavy and 25 kDa light chain on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but demonstrated a single band at 50 kDa on western blot. Treatment with a N-linked glycosidase extinguished the band. Flow cytometry demonstrated antigen-specific binding of CA19-9-positive cells and the antibody localized to the antigen-positive tumor in vivo. We successfully produced an antibody with an azido-sugar at the conserved CH2 glycosylation site. We were able to utilize this azide to label the antibody with biotin or fluorescent label and demonstrate that the label is added in a site-specific manner to the heavy chain, N-linked glycosylation site. Finally, we demonstrated functionality of our antibody for in vitro and in vivo targeting of pancreatic cancer cells

Metabolic exploitation of the sialic acid biosynthetic pathway to generate site-specifically labeled antibodies.

Lack of a universal site-specific conjugation methodology for antibodies limits their potential to be developed as tumor-specific imaging agents or targeted therapeutics. A potential mechanism for site-specific conjugation involves utilization of the conserved N-glycosylation site in the CH2 domain. We sought to develop an antibody with an altered azido-sugar at this site whereby site-specific label could be added. The HB8059 hybridoma was cultured with peracetylated N-azidoacetlymannosamine (Ac4ManNAz). The resulting azido-sugar antibody was conjugated to phosphine-polyethylene glycol (PEG3)-biotin via a modified Staudinger reaction. Biochemical and functional characterization of the biotinylated antibody was performed. The azido-sugar antibody was also labeled with DyLight-650-Phosphine and injected into mice harboring pancreatic cancer xenografts. The tumors were dissected and imaged utilizing an IVIS fluorescent camera. The antibody was successfully produced in 100 μM Ac4ManNAz. The biotinylated antibody demonstrated a 50 kDa heavy and 25 kDa light chain on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but demonstrated a single band at 50 kDa on western blot. Treatment with a N-linked glycosidase extinguished the band. Flow cytometry demonstrated antigen-specific binding of CA19-9-positive cells and the antibody localized to the antigen-positive tumor in vivo. We successfully produced an antibody with an azido-sugar at the conserved CH2 glycosylation site. We were able to utilize this azide to label the antibody with biotin or fluorescent label and demonstrate that the label is added in a site-specific manner to the heavy chain, N-linked glycosylation site. Finally, we demonstrated functionality of our antibody for in vitro and in vivo targeting of pancreatic cancer cells

A mutated anti-CA19-9 scFv-Fc for positron emission tomography of human pancreatic cancer xenografts.

PurposeIntact antibodies have a long serum persistence resulting in high background signal that inhibits their direct translation as imaging agents. Engineering of antibody fragments through the introduction of mutations in the fragment crystallizable (Fc) region can dramatically reduce serum persistence. We sought to develop a Fc-mutated, anti-CA19-9 antibody fragment (anti-CA 19-9 scFv-Fc H310A) to provide micro-positron emission tomography (microPET) imaging of pancreatic cancer xenografts.ProceduresThe anti-CA19-9 scFv-Fc H310A was successfully expressed and purified. Biochemical characterization included size exclusion chromatography, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), Western blot, and flow cytometry. The antibody fragment was labeled with iodine-124 ((124)I) and injected into mice containing human pancreatic cancer xenografts. MicroPET/CT images were then obtained. Blood, organ, and tumor radioactivity was measured and expressed as the percent of injected dose per gram of tissue (%ID/g).ResultsBiochemical characterization was consistent with the creation of a 105 kD dimer containing a human Fc region. Flow cytometry demonstrated antigen-specific binding, and cell-based ELISA further established a dissociation constant (K D) of 10.7 nM. (124)I-labeled scFv-Fc H310A localized to the antigen-positive tumor xenografts as detected by microPET. Objective confirmation of targeting was demonstrated by higher %ID/g in the antigen-positive tumor compared to the blood, antigen-negative tumor, and liver.ConclusionsWe successfully engineered and produced an anti-CA19-9 scFv-Fc H310A antibody fragment that retains similar affinity when compared to the parental intact murine antibody. Additionally, our engineered and mutated fragment exhibited antigen-specific microPET imaging of both subcutaneous and orthotopic pancreatic cancer xenografts at early time points secondary to decreased serum half-life

Research Plan: Foam Delivery of Remedial Amendments to Deep Vadose Zone for Metals and Radionuclides Remediation

Research proposals were submitted to the Scientific and Technical Basis for In Situ Treatment of Metals and Radionuclides Technical Working Group under the US Department of Energy (DOE) Environmental Management Office (specifically, EM-22). After a peer review and selection process, the proposal, “Foam Delivery of Remedial Amendments to Deep Vadose Zone for Metals and Radionuclides Remediation,” submitted by Pacific Northwest National Laboratory (PNNL) was selected for support by the program. A research plan was requested for this EM funded project. The overall objective of this project is to develop foam delivery technology for the distribution of remedial amendments to deep vadose zone sediments for in situ immobilization of metal and radionuclide contaminants. The focus of this research in FY 2009 is on the physical aspects of the foam delivery approach. Specific objectives are to 1) study the foam quality (i.e. the gas volume fraction in foam) influence on injection pressure, 2) study the sediment air permeability influence on injection pressure, 3) investigate liquid uptake in sediment and determine whether a water front will be formed during foam delivery, 4) test amendment distance (and mass) delivery by foam from the injection point, 5) study the enhanced sweeping over heterogeneous systems (i.e., low K zones) by foam delivery relative to water-based delivery under vadose zone conditions, and 6) numerically simulate foam delivery processes in the vadose zone. Laboratory scale experiments will be conducted at PNNL to study a range of basic physical aspects of the foam propagation in sediments, including foam quality and sediment permeability influence on injection pressure, liquid uptake, and foam sweeping across heterogeneous systems. This study will be augmented with separate studies to be conducted at MSE Technology Applications, Inc. (MSE) to evaluate foam transport and amendment delivery at the intermediate-scale. The results of intermediate-scale tests will be used to bridge the gap between the small-scale foam transport studies and the field-scale demonstration. Numerical simulation studies on foam delivery under vadose conditions will be performed to simulate observed foam transport behavior under vadose zone conditions and predict the foam delivery performance at field-scale