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Biomedical subjects

K A Kang

Publications and source records attributed to K A Kang.

At least 19 recordsLinked to original sources
PubMed1999

Preliminary study of real-time fiber optic based protein C biosensor.

Deficiency of protein C (PC), one of the human body's key anticoagulants, can lead to massive thrombotic complications. There is a diagnostic need to perform real-time assays, in order to quickly identify and treat this disease. An immuno-optical biosensor for the diagnosing of PC deficiencies and monitoring of PC concentrations is being developed for this purpose. Monoclonal antibody against PC (anti-PC) is immobilized on the surface of a tapered quartz fiber that is enclosed in a glass tube (capacity approximately 200 microL). Following sample injection, PC within a sample binds to the anti-PC in a highly specific reaction. The system is then probed with a fluorophore-tagged secondary antibody against PC. Excitation light is applied through the fiber, and the fluorescence intensity is correlated with the PC concentration in the sample. This study presents (1) a feasibility, direct binding assay, (2) a comparison of methods to immobilize anti-PC upon the fiber (direct immobilization vs an avidin-biotin bridge), and (3) effectiveness of an elution step to regenerate the fiber. PC-deficient patients typically have a concentration range less than 2.5 microg/mL. It was found that the sensor could detect PC levels down to 0.1 microg/mL in pure buffer with minimal optimization. Avidin-biotin immobilization of the primary antibody produced enhanced signals, up to 470% of the original intensities. Preliminary fiber regeneration tests achieved nearly a 50% increase in fiber lifetime with the use of a CaCl(2) elution step. Ultimately, further development may lead to automation and the use of the system as a multi-blood factor analyzer.

Animals↗
PubMed1998

Preliminary study of biosensor optimization for the detection of protein C.

Methods to develop an immuno-optical biosensor for the detection and monitoring of Protein C (PC) concentrations are described. A tapered quartz fiber is enclosed in a glass tube (capacity approximately 300 microliters) and monoclonal antibody against PC (anti-PC) is immobilized on the surface of this fiber. PC within a sample, when injected into the chamber, will bind to the anti-PC in a specific reaction. The system is then probed with a fluorophore tagged secondary antibody against PC, also binding to PC in a specific reaction. Excitation light is applied through the fiber, and the amount of fluorescence is correlated with the PC concentration in the sample. This study offers encouraging results for the detection of PC deficiency in real-time.

Antibodies, Monoclonal↗
PubMed1997

Chromatographic process identification for protein C purification using frequency response analysis.

Frequency response analysis is applied for the analysis of liquid chromatography output of protein separation. Reduced data from simple chromatograms suggest that various Bode plot parameters, magnitude ratios, phase shift, the steady state gain, break frequency, and system order in the frequency domain, can be used to gain phenomenological insights on the system. Such an approach is advantageous because the validity of the model can be checked for two plots, the magnitude ratio vs. frequency and the phase shift vs. frequency, as compared to a single plot in the time domain. This approach also provides a useful empirical-tool which can be quantifiably used for process validation and scale-up, especially for immunoaffinity and immobilized metal affinity chromatographic systems used for protein C purification.

Animals↗
PubMed1997

Separation of recombinant human protein C from transgenic animal milk using immobilized metal affinity chromatography.

Protein C is an important serine protease due to its ability to proteolytically cleave activated Factors V and VIII. Excess coagulation and blood agglutination can lead to plugged capillaries, thereby reducing oxygen transport to interstitial tissues. To treat patients with hereditary and acquired protein C deficiency would require a greater amount of Protein C than that available from human plasma. However, the potential demand for this protein could be met by the production of human protein C from transgenic animal mammary glands. Thus, research into inexpensive, efficient methods to purify proteins from transgenic animal milk will be a critical area of study for the large scale production of protein C. Immobilized metal affinity chromatography (IMAC) is a novel method for the purification of protein C. A proposed method of purification is to take advantage of protein C's strong metal ion binding characteristics with IMAC to assist in the separation from transgenic animal milk. The separation procedure is benchmarked against current systems in use by the American Red Cross for purification of Protein C from transgenic porcine milk. Common problems in developing separation schemes for new therapeutics are the initial availability of the product (protein), and time-to-market concerns. Extensive experimental tests for scaleable purification schemes are often cost and time prohibitive. In order to optimize an IMAC protocol with minimal waste of time and resources, total quality management tools have been adopted. Initial experiments were designed to choose buffer conditions, eluents, immobilized valence metals, and flow rates using Taguchi experimental design, which is a total quality management (TQM) tool. One of the values of Taguchi methods lies in the use of Latin orthogonal sets. Through the use of the orthogonal sets, the total number of experiments may be reduced, shortening the focus time on optimal conditions.

Animals↗
PubMed1997

Reusable, real-time, immuno-optical protein C biosensor.

A Protein C (PC) biosensor can be used to diagnose PC deficiency, to monitor the PC level in the blood of PC deficient patients, and to measure the PC concentration in other PC-containing samples, such as PC producing animal cell culture broth or transgenic animal milk. A fully functional biosensor requires extremely high sensitivity and specificity, and real-time measurement. To satisfy these requirements, it is proposed to develop an immuno-optical fiber biosensor that utilizes PC-specific biomolecules (PC probes) tagged with fluorophores. The method involves immobilizing monoclonal antibody against PC (anti-PC) on the surface of an optical fiber. When PC in a sample is adsorbed to the anti-PC on the fiber, it can be reached with the fluorophore tagged PC-probe. The intensity of light transported through the optical fiber, therefore, can be correlated with the concentration of PC in the sample. The sensor will be designed so it can be reused, following a simple elution step, thus reducing diagnostic expense. The preliminary study shows encouraging future for the real-time optical PC biosensor.

Animals↗