The Characterization of Human Epidermal Stem Cells
The role of Notch signaling in the regulation of growth and differentiation of epithelial stem cells is poorly understood. While specific markers for epidermal stem cells have not yet been identified, members the Notch signaling pathway have been reported to be differentially expressed in the human epidermis. This study sought to demonstrate the presence and distribution of Notch and its ligands, Delta and Jagged, in human keratinocytes, and thereby characterize this subpopulation. Human neonatal foreskin samples were used to obtain isolated epidermal cells. Cells that were shown to be negative for connexin43, a gap junction protein, and positive for keratin14, a basal marker, were classified as presumptive stem cells (PSC). This sorted subpopulation was shown to be small and agranular by flow cytometry analysis. After two weeks in cell culture, PSC revealed a proliferative potential three times greater than non-sorted cells. The PSC exhibited increased expression of Delta and Jagged ligands than the general population. Additionally, RT-PCR confirmed the presence of Jagged and Delta in keratinocytes; however, only Jagged was detected in immunohistochemistry tests. Members of the Notch family were identified by immunohistochemistry in the epithelium and also at the protein- and mRNA-level. The data suggests that variations in the expression of members of the Notch signaling pathway could potentially be used as markers for stem cells of the epithelium; however, further research is necessary to make definitive conclusions, which would provide better insight into Notch regulatory pathways. This understanding could one day allow for the eventual treatment of epithelial damage caused by various skin diseases, injuries, or burns.
Novel Biotechnological Approach for Recognition and Purification of Antibody: Lectin Affinity Membranes
Immunoglobulin G is a glycoprotein structured molecule that is produced by the immune system and protects organism from harmful effects of antigens. Ig G amount in the blood plasma is an appropriate indicator of; infection, cancer, diabetes, cardiovascular diseases, Alzheimer and other autoimmune diseases. Besides, purification of Ig G used in the treatment of these diseases from naturel sources is carried out at high costs on the World market. It is hard to obtain Ig G in high amounts and without any decomposes, that’s why it is important to develop new systems that will help to recognize and purify Ig G antibody. In this project, my purpose was; recognizing Ig G antibody with efficient, high amounted, fast, easily, with less toxicity, economically and purifying Ig G in high ratios from its natural sources. For this purpose p(HEMA-EDMA) membranes are synthesized with free radical photo polymerization method and characterized according to SEM images, swelling behaviors FTIR analysis and elemental analysis. In order to adsorb Ig G to polymeric membranes; polymeric membranes are activated with silanization agent (IMEO) and derivatized with Con A which is a lectin affinity ligand. In the SEM results it is examined that membranes are in spherical structures. Highest swelling value is determined as 224.8%.Binding of IMEO was demonstrated with FTIR and Elemental Analysis. Optimum conditions for Ig G adsorption to membranes are; 1.5 mg/ml initial Ig G concentration, 30 minutes of adsorption time, pH 4 citrate buffer 37 0C and without any different ion strength. Optimum adsorption capacity is determined as 253.8 mg/cm2 and it is also determined that this value is 7 times higher than nonspecific Ig G adsorption to p(HEMA-EDMA) membranes. Ig G adsorption-desorption cycles (5 times) proved that product is reusable without losing its adsorption capacity. According to the electrophoresis, Ig G could be desorbed in pure form without any denaturation to its structure.
A New Generation Colorimetric Method for Lead Analysis: APTAMER MODIFIED GOLD NANOPARTICLES
Lead is a toxic element which is used in the production of chemicals, dyes, accumulators and various industrial areas. It may cause complications even extended to death when it is taken consistently in high amounts. Lead poisoning is in the first place among the occupational diseases. It is gaining importance to develop new and sensitive methods for lead analysis. Because lead poisoning can progress without any symptoms and poisoning level (10µg/dL) is low. Disadvantages of the systems used for lead analysis are such as longer detection time, being expensive and difficult to implement. The aim of this project is to develop a new generation method in order to detect lead in blood, based on aptamer modified gold nanoparticles. We detected the lead in terms of color change obtained in gold nanoparticle solutions, with composite biochemosensor that is prepared with 20 & 80 nm sized gold nanoparticles and TBA(Thrombin Binding Aptamer). While immobilizing TBA to the gold nanoparticles, we benefitted from the magnificent surface affinity of the –SH (Thiol) groups that modified to the TBA. Gold nanoparticles that are used in development of our biotechnological method do not stimulate the immune system. The preparation of aptamers in completely sterile medium provides us to use our system in the lead detection of blood. Our method can also be used in the lead detection of mediums such as waste water, food and soil. We have developed a biochemosensor that can be used to detect the presence and absence of Pb2+ by taking into consideration the toxic effect in the human body. Also we detected the presence of lead colorimetrically, in low concentration levels and wide interval values of 4.4 – 11 µg/dL. The developed system is first that; it provides TBA to be used with its complimentary sequence detects the presence of lead colorimetrically and can be used in physiological media such as blood. Also our system can detect lead in amounts that are lower than the poisoning threshold.