Biomedical subjects

Carl K Fredrickson

Publications and source records attributed to Carl K Fredrickson.

4 recordsLinked to original sources

PubMed2006

Ricin detection by biological signal amplification in a well-in-a-well device.

This paper presents a ricin detection method based on ricin's inhibitory effects on protein synthesis. Biological synthesis (expression) of a protein includes the steps of gene transcription (DNA --> RNA) and protein translation (RNA --> proteins); these reactions can be coupled into a one-step operation and carried out in a cell-free medium. Ricin is known to inhibit protein synthesis by interacting with 28S ribosome RNA; the inhibitory effect is exploited as the sensing mechanism in this work. For each copy of DNA, thousands of copies of proteins can be produced. As a result, the inhibitory effects of ricin are amplified, leading to a significantly enhanced detection signal (the difference between the positive control and samples). An array of protein expression units is developed to accommodate positive/negative controls and multiple samples. The array device contains a solution without any reagent captured on a solid surface, offering flexibility without comprising the activities of biomolecules. The miniaturized well-in-a-well design possesses a mechanism to supply nutrients continuously and remove byproducts, leading to higher protein expression yields and thus larger detection signals (lower detection limit) when ricin is present. We demonstrate the production of green fluorescent protein and luciferase in the device. A calibration curve has been obtained between the luciferase expression yield and the ricin concentration, showing a detection limit of 0.01 nM (0.3 ng/mL) ricin. The nested-well device is also used for measuring the toxicity level of ricin after physical or chemical treatment.

Calibration↗

PubMed2005

Toxin detection by a miniaturized in vitro protein expression array.

A novel concept for toxin detection is presented that is based on the inhibitory effects of certain toxins on protein synthesis. Biological synthesis of proteins is called protein expression, which consists of the steps of DNA transcription and protein translation. In the toxin detection concept, a group of proteins are expressed simultaneously in a device consisting of an array of miniaturized wells. The expression yields of these proteins are inhibited by presence of a toxin. Differential inhibitory effects of each toxin on the expression of proteins in wells result in a response pattern (or signature) of the array device, which can be used as a tool for detection and identification of known and unknown agents. The concept has been demonstrated by in vitro protein expression of three proteins, including green fluorescent protein (GFP), chloramphenicol acetyltransferase (CAT), and luciferase. The production yields of these proteins in a cell-free medium were inhibited differentially by two toxin simulants, tetracycline (TC) and cycloheximide (CH). We confirmed that TC has an inhibitory effect on the production of GFP and CAT in the E. coli expression system whereas CH has a negligible effect. Moreover, the degree of TC's inhibitory effect on GFP is different from that on CAT. We also observed an opposite disparity; TC has a negligible inhibitory effect on the luciferase production in the rabbit reticulocyte expression system, whereas CH has a significant inhibitory effect. In addition, we obtained a response pattern in a 3 x 4 array device; the results suggest the feasibility of toxin detection based on the mechanisms of toxin actions. The device and method are also likely applicable to high-throughput drug screening, evident from the fact that TC is an antibiotic medicine.

Animals↗

PubMed2005

Conductivity properties of carrier ampholyte pH gradients in isoelectric focusing.

The conductivity properties of natural pH gradient created by carrier ampholytes were studied during the process of isoelectric focusing (IEF). IEF was performed in capillaries (10-30 mm long) or in microchips with the same channel length. A 10-30x reduction of the conductivity of the separation medium was observed during the establishment of pH gradient. Results obtained using different IEF voltages indicate that there is a nonlinear relationship between the conductivity of an established pH gradient and the applied electric field. Our theoretical analysis using a simplified model generated values that reasonably agree with the experimental data. In addition, we found that above a certain electric field ( approximately 300 V/cm), resolution does not increase with the applied voltage as predicated; we observed band-broadening and gel breakdown. The approach presented in this work can be used for optimization of the IEF separation and judicious selection of IEF conditions.

Ampholyte Mixtures↗

PubMed2004

Macro-to-micro interfaces for microfluidic devices.

Since the concept of miniaturized total analysis systems (microTAS) was invented, a great number of microfluidic devices have been demonstrated for a variety of applications. However, an important hurdle that still needs to be cleared is the connection of a microfluidic device with the rest of the world, which is often referred to as the macro-to-micro interface, interconnect, or world-to-chip interface. In this review, we will examine the methods used by pioneers in the field and other investigators, review the approaches for capillary electrophoresis-based devices and those using pneumatic pumping, and present additional discussion on interface standardization and choosing and designing interconnects for your applications.

Electrophoresis, Capillary↗