Jean Piaget: The Four Stages Of Cognitive Theory

Jean Piaget: The Four Stages Of Cognitive Theory Cognitive theory of Jean Piaget includes four stages of development that children move through during which the explanatory behaviors of infants transform into the abstract, logical intelligence of adulthood. There are three important specific characteristics of Piagets theory of which the first one is being a general theory, that is, cognitions all aspects undergo a similar course of change. Another characteristic is that children move through the stages in an invariant sequence. Piaget believed that there is a same order that children follow. Third, the stages are universal. Stages in cognitive theory assume the theory to include all children everywhere (Berk, 2003). Biological concepts are used in a limited way in Piagets theory. However, he stated the importance of genetic and environmental factors on the way that children move through the stages (Crain, 2005). He emphasized that the speed of children while passing those stages is affected by differences in genetic and environmen tal factors. Jean Piaget used the term scheme while explaining human beings organized way of making sense of experience (Mark, 1969). Traill (2008) explains that the term scheme used by Piaget is different from peoples everyday usage of scheme. The term can be any pattern for exploring and learning from the environment and it has three different intellectual structures. Piaget calls first intellectual structures to emerge as behavioral schemes, ones that appear after 2 years as symbolic schemes, and structures that appear after 7 years as operational schemes (Piaget, 1972, as cited in Traill 2008). For instance, dropping scheme of an 8 month old baby and a 25 months of will not be the same, as sooner it will become more deliberate and creative. Toddlers, different from infants, begin to think before acting and Piaget identifies that transition from sensorimotor to cognitive approach to the world which depends on mental representations. (Piaget, 1926, as cited in Berk 2003) Images and concepts are the two powerful mental representations. Especially, the shift from sensorimotor to cognitive approach is accounted for two processes; adaptation, consisting assimilation and accommodation, and organization. Interpretation of new structures into already existing schemes is called as assimilation and modification of existing schemes into adaptation of new experiences is called as accommodation. Cognitive adaptation aims to adjust to the environment and is a result of the equilibrium between assimilation and accommodation (Block, 1982). While trying to grasp an object, a baby is experiencing the assimilation process, while removing an obstacle and grasping an object, a baby now accommodates the scheme (Crain, 2005). During the organization process more complex intellectual structures are combined with existing schemes by children. For instance, after the baby experienced and covered dropping movement, then he/she will relate it with throwing movement as well as understanding the concepts of near and far (Berk, 2003). The Sensorimotor Stage (Birth to 2 Years) Jean Piaget observed his children during their developmental period and constructs the stages based on his observations. His books mostly involve many examples from his dialogues and interactions with his children. The sensorimotor stage consists of six substages. (Santrock, 2004) That stage starts with the use of reflexes from birth to 1 month. Newborn reflexes take important place in sensorimotor stage. According to Piaget inborn reflexes are consisted from first schemes. He states that as children use inborn reflexes and experience assimilation, they desire to put them to active use (Crain, 2005). After one month, children begin to repeat their chance behaviors and primary circular reactions period (one to four months) starts. A baby experiences the thumb sucking by bringing her hand to her mouth by a chance, when the hand falls she wants to get it back and experiences many failures until she gets it back (Crain, 2005). At that example the child organizes the hand movement and sucking which is a kind of circular reaction. Piaget also states that children at that period indicate the first efforts at imitation (Berk, 2003). The next substage is secondary circular reactions and is observed between fourth and eighth months. Infants start to experience motor achievements that encourage them to play attention to their environment. Infants begin to get enjoyment from the response of the environment to their attempts and they repeat their movements that get reaction from their surrounding (Santrock, 2004). Coordination of secondary schemes substage takes place during eight to twelve months. At this stage infants begin to coordinate tow or more actions to achieve simple objectives. In addition with an intentional purpose, babies try to imitate behaviors after watching a person. One may be able to observe a baby at this stage trying to stir with a spoon. In addition, a baby may begin to cry when she sees her mother wearing her coat in order to stop her mother leaving (Berk, 2003). In substage 5, tertiary circular reactions (twelve to eighteen months), children are interested with different outcomes. Piaget had observed one of his children hitting on a table at different rates in order to listen different sounds that he creates (Crain, 2005). It should be noted that all experiences are results of childrens intrinsic curiosity about the environment around them that Piaget emphasizes within his cognitive development theory. The last substage of the sensorimotor period is named as beginnings of thought or internalization of schemes lasting from eighteen to twenty months. During that substage children have the capacity to remember the behaviors that are not present (deferred imitation). Their efforts on imitation also indicate progress and they experiment with actions inside their heads. Besides, children can be observed to engage in make-believe play during that period (Santrock, 2004). Object Permanence: Piaget and many researchers concluded that infants appreciate concepts of permanence objects. Up to four months, children do not make any attempt to an object leaving in front of their eyes. During secondary circular reactions stage children are more able to explore their surrounding and they have a better sense of permanence of objects. At stage four children have the ability to find the hidden objects. If an adult takes a toy behind a box, the baby will look at the behind of the box and find the toy. During the stages five and six children are able to follow displacements and follow invisible shifts (Crain, 2005). Beginnings of Categorization: Before the capability of mental representation children are not able to categorize objects. During the first year of their life, children experience perceptual categorization. For example they can categorize the legs of an animal. Conceptual categorization begins with the end of first year; they are now able to categorize similar characteristics and behaviors. Active categorization period starts with the beginning of the second year. It is stated that sorting objects into two classes can be observed in eighteen months babies. In the second year babies can group two different kinds of objects without grasping them (Berk, 2003). When the observed milestones of research and the description of substages of Piaget are compared from birth to two years, both similarities and differences are seen. There are points that seem to occur earlier than Piaget accepted such as categorization, deferred imitation, and analogical problem solving. Those differences are explained differently from many researchers. Some of the surveys indicate that some children born with different intellectual capacities and some of them with a set off limits which causes those differences. The latter argue the theory of Piaget in terms of biological considerations. The Preoperational Stage (2 to 7 Years) Preoperational stage is lasting from two to seven ages in which the child is more capable while dealing with the environment. Although the reasoning of child is still unsystematic and illogical, that is the period that children begin to use symbols and rapidly develop representation. One of the important symbols that indicate increase during that period is language (Santrock, 2004). Piaget believed that experience of internal images occurs before labeling words and he did not take language as an important tool in cognitive development of children. Berk (2003) argues that Piaget had misadjusted the role of language in early intellectual development. She proposes that conceptual abilities of children are highly affected from the dialogues of children with adults. Moreover, there are many psychologists that believe as children develop their language ability, they begin to think more logically. Children experience transductive reasoning during that stage which means shifting from one particular to another. Children place two unrelated situations into the same case as if they have a relationship. One of Piagets children had concluded that she hadnt had her nap yet so it wasnt afternoon (Piaget, 1924). Piaget (1924) explains that statement as an example of transductive reasoning, because the child did not catch the understanding that afternoons include many different events and having nap is only one of them. An important milestone of the increase in mental representation is make-believe play during preoperational stage. The differences in make-believe play between sensorimotor and preoperational stage can be clearly observed. By the middle of preoperational stage make-believe play of children indicate real life conditions. In addition, by preoperational stage children begin to engage in sociodramatic play, they coordinate variety of roles and story lines during their play. One of the criticized points of cognitive theory of Piaget is based on the belief of Piaget that play reflects childrens cognitive and social skills, however there are many recent studies indicating the contribution of play on those skills. Especially during sociodramatic play, children interact with their peers longer and they are more cooperative. Many psychologists believed the role of strengthening of make believe play on a wide range of mental abilities and logical reasoning (Berk, 2003). Egocentrism: Piaget stated that children look at their surrounding from their own viewpoint and they ignore perspectives of others. Three-mountains study is one of the famous observations of Piaget explaining egocentric behavior of children at preoperational stage. He had used a model of three mountains and taken a child for a walk around the model in order to give opportunity for the child to look at the model from different view. Piaget had placed the child from one point of the model and placed a toy to another place. The child had been asked what he/she saw while looking at the model and what the toy would be seen while looking at it. All the children could correctly explain what they were seeing, however children at preoperational stage gave the same answer with their own view (Crain, 2005). Studies emphasize on the relation between egocentrism and social communication. Children at preoperational stage, according to Piaget, fail to recognize the needs of their peers during verbal interaction (Rubin, 1973). As they look only from their own view, they are able to understand view of the person interacting with them. They think that they can be seen from everywhere, everybody see and hear them. An adult may observe a child at this period telling that nobody could see him/her while closing his/her eyes with hands. Animism: Piaget (1951) proposes that the child recognizes no limits between himself and the external world and it is expected that the child would see many nonliving and non acting things as living and conscious and he explains this phenomenon as animism. In his book The Childs Conception of World, 1951, he identifies the reason for him to use the term animisim. He accepts that animism was term used for primitive human beings and responds the criticisms by telling that he had used that term as a generic term and emphasizing on the different types of animism in psychological origins (Piaget, 1951). Children at preoperational stage have a belief that objects are alive because they move and grow. For example, a child may tell that there are not any cars on the road, because they are sleeping. Piaget described animism inside four stages. Initially children accepted useful things as living. At this first stage broken or damaged objects were not alive for them. At the second stage, moving objects, whether are moved by an external factor or by themselves, were considered as alive. In stage three, to be categorized as living, things should move by themselves. Lastly, at the fourth stage, adults know that plants and animals are living things only (Moriarty, 2005). Irreversibility: Going through a series of steps and after changing direction is difficult for children at preoperational stage. Another well known experiment of Piaget indicates that problem in a way that there are children shown 16 boxes, 6 of which are yellow and 10 of which are red. When children are asked whether red boxes are more or boxes, children at this stage responds as red boxes and fails to be aware of that both yellow and red boxes are boxes. In his book The Childs Conception of World, 1951, Piaget gives examples about irreversibility. There are dialogues indicating their inability such as, asking a child about her sister, the child responds that she has a sister named A, then Piaget asks the child whether A has a sister or not, the child responds that A has not a sister. (Piaget, 1951) Inability to Conserve: Piaget propounds preoperational childs lack of conservation by applying experiments of liquids and number. He shows two same size glasses to the children and fulls the glasses with water. He asks children which of the water was more. All the children respond that they were equal in amount. Then he puts the water in one of the glasses into a different size glass (wider or taller) and repeats his question. Children at preoperational stage tell that they are now different. They have not the capability to perceive that certain physical features of objects remain same, even their physical appearance changes (Santrock, 2004) Based on experiments of Piaget, at the beginning of seven children begin to give the correct answer to the conservation tests. Before that age children indicates at conservation but not totally achieve it. They give answers like one is more because it is taller and then change their answers the other one is more because it is wider. Besides, irreversibility of the child can be concluded based on the conservation of liquid experiment. The child cannot understand the end result as a reverse of the original one. Jean Piaget also had thought about the failures of children from the linguistic point. Terms such as taller, more, wider takes time to be understood. He suggests ways to overcome that problem and tells adults to apply experiments by using different sentences and establishing questions by using different words within a particular case (Berk, 2003). Piaget experimented conservation of children also with using number. A row of egg cups and a bunch of eggs are given to children and they are asked to take enough eggs to fill the cups. At the first half of preoperational stage children ignored the number of eggs in the cups and they made an equal length with the rows. Children at the second half of preoperational stage made one-to-one correspondence. However, those children failed in the second question. Piaget then brought together the eggs and asked which of them was more and they could not respond that they are still same. Most of them thought that the longer was more in number. Piaget explains that phenomenon as been influenced by their sudden perceptions than by logic (Crain, 2005). Categorization: In coordinating and extending knowledge in cognitive development theory of Piaget, categorization takes an important place. Researchers state that children begin to make similar categorizations and form some categories with the age of two such as animals, plants, and vehicles (Scholnick, Nelson, Gelman, Miller, 2008). Things or objects in all of those categories have differences in perceptual characteristics, so a conflict emerges with the main idea of Piaget that childrens reasoning is governed by the way they see and objects appear. By the age of three children become able to make distinctions between basic and general categories such as furniture versus tables. However, children at preoperational stage are not able to organize objects into classes and subclasses based on their basic similarities and differences (Meadows, 1986). It is argued that Piaget was partly right and partly wrong with his conclusions of preoperational stage. Researchers give simplified tasks for children and preschoolers indicate the beginnings of logical operations, however there are important differences between their reasoning and children at school age. Difficulties were observed in conservation tests, three mountains tests and appearance-reality tests. As opposed to perceptual approaches to solve problems, children rely on increasing effective mental. In order to give an example, research indicate that children who have not the capability to use counting while comparing two groups of objects, do not also conserve number. They begin to find effective solution ways for proms with more objects when they become capable of counting. Piaget states that, as many other psychologists accept, children move through variety of stages of understating, although they do not totally handle conservation up to school years. The Concrete Operational Stage (7 to 11 years) The beginning of concrete operational stage is furnished by logical reasoning. That period is accepted as a major turning point in cognitive development. The time that a child begins to mentally resemble an adult starts with attaining concrete operational stage. A school-aged childs cognitive performance is more evident in terms of concrete operations. For example an eight year of childs response to conservation of liquid test is that the amount has not changed and they are still same. This explanation of the child also indicates the reversibility of the reasoning that the child has now the capability to understand that the reverse is the same with the original. Their ability to achieve conservation tasks indicate their logical thinking (Richardson, 2003). Between ages seven and ten, children take attention on relations between a general and two specific categories within the same case. Their awareness of classification is getting developed. In middle childhood period many children are interested in collections such as coins, rocks, and stamps which show their level of classification. Children in the concrete operational stage are fairly good at the use of inductive reasoning which involves going from a specific experience to a general principle, whereas children at this age have difficulty with using deductive reasoning. Deductive reasoning involves using a general principle to determine the outcome of a specific event. Seriation: Seriation refers to the capability to arrange objects along a quantitative dimension. Ordering sticks from different lengths is one of Piagets tests. Children at five and six years could create series but they made many errors. However, children at concrete operation stage, by starting the shortest and putting the next, indicate a perfect arrangement with an orderly plan. Children between ages seven to eleven have an ability called transitive inference which refers to mentally arrangement of items. Piaget had showed children three stick with different colors. Children had observed that first stick is longer than second stick and second stick is longer than third stick and they had made the inference that first stick is longer than third stick. Children at concrete operation stage mentally integrate three relations at once. On the other hand, preoperational stage children can achieve analogical reasoning by encouragement of an adult (Meadows, 1989). Spatial Reasoning: The ability of understanding space is more developed in concrete operational children. School age children begin to give directions as they gain more advanced space understanding. Children at five or six years indicate difference while representing an object on a persons, in front of them, left or right. Starting from seven age children achieve mental rotations, that is, they can recognize their own frame and of a person in a different direction (Santrock, 2004). While drawing large scale areas their performance indicate important changes representing their development of cognitive skills. In the middle period of concrete operational stage children achieve to place notes showing the location of objects in their classroom. Their capability to use a rotated map is improving and also their map drawing become more accurate. Research indicate that concrete operational thinking has a limitation that while children are reasoning in a logical manner, they are always dealing with concrete knowledge. With abstract information, they cannot catch the achievement (Crain, 2005). When they are given ideas that do not appear in the real world they are not able to indicate the same responds with concrete ones. That can be easily observed from transitional inference that when children are shown the sticks they give the correct responds whether which one is smaller or taller than the others. However, when the questions turned to a more concrete version such as As hair is longer than B, B2 hair is longer than C. Who has the longest hair?, up to eleven years children cannot give the right answer (Richardson, 2003). Jean Piaget believed the importance of rich and appropriate environment for the speed of concrete operational cognitive skills of children. Many research done in small towns or villages represent the lack of rich environment and its consequences on children. Even the easiest conservation tasks are not easily achieved until the age of eleven. On the contrary, many children living in big cities, especially Western nations, have many opportunities in terms of environment and from very young periods of childhood they meet with rich and varied materials. Hence, they easily understand the conservation tasks. The same comparison had done between children receiving a professional early childhood education and the ones that spend their early period at home or street. The conclusions indicate better results of the children continuing a preschool program (Berk, 2003). A survey had been made between Brazilian street vendors and Brazilian economically advantaged children on informal version and Piagetian version of class inclusion test of Piaget. In the informal one, the researcher asks children for the price of two chewing gums which are different. In the Piagetian one, four units of one type (mint) chewing gum and two units of the other (strawberry) one is set aside and the researcher asks In which one do you get more money, whether you sell me the chewing gum with mint or all the chewing gum? The success of two groups children indicates difference that on the informal version of the conservation task street vendors performed better and on the other version economically advantaged children performed well (Ceci Roazzi 1994, as cited in Berk 2003). The Formal Operational Stage (11 Years and Older) The child in sensorimotor period can do things and play with objects, the child in sensorimotor period and concrete operational period can think and make reasoning about those objects. Thinking about propositions and relations different from objects and events is another period. Now human beings are bale to make coordinations of coordinations, in other words, they can combine two different groupings of concrete operations within reversibility by reciprocity (Richardson, 2003). In formal operational stage adolescents do not require concrete things as objects of thought. Adolescents at this stage are capable of deductive reasoning. In order to overcome a problem, they start more generally and think evaluate all possible factors and then proceed to more specific predictions. Piaget used pendulum problem in that stage in which he had showed many strings with different lengths, objects differing in weight, and in order to hang the strings a bar to children. He asked the factors that will affect the speed with which a pendulum swings. Children in concrete operational level made an unsystematic experimentation. The influences of each variable could not be distinguished by them. Adolescents of formal operational stage separate the factors that have the possibility to affect the speed into four groups; strings length, objects weight, the level of force when pushing the object, and how high the object is raised before it is released. At the end they come up to a right conclusion that the length of the string affects the speed (Berk, 2003). They work systematically in terms of all possibilities as a scientist. Cognitive development theory of Piaget is mostly depending on mathematical and scientific reasoning; however he has some speculations on social life of formal operational stage adolescents. Piaget thinks that children between seven and ten live in here and now, whereas at the formal operational stage they start to think about long-term periods. They handle abstract ideas such as justice and love and they start to construct theories for better living world (Inhelder Piaget, 1955, as cited in Crain 2005). Piaget identifies egocentrism again and believes that a new kind of egocentrism is carried by utopian during that period. Piaget takes into account and repeat the forms of egocentrism from beginning to later periods: At the beginning, sensorimotor period, infants have no idea bout the environment around them and they are egocentric, still the objects that they cannot see have no existence on their own. At the next level, preoperational thought, children enter a representative world in terms of language and symbols. They indicate difficulty more than their own view. After a time they become aware of other perspectives and they start to think about concrete objects. Finally, adolescents, formal operational period, meet with a world full of problems and possibilities. Adolescents feel unlimited power in themselves for their thoughts and egocentrism reappears and they are now not able to distinguish self and others abstract views.(Crain, 2005). Teenagers feel that they are at the centre of their environment and they assume that everyone is looking at them. They spend many hours in front of a mirror feeling anxiety for the peoples criticism about them. This is called as imaginary audience. Then with having a thought that everyone is observing them, they start to feel themselves special and unique and this is named as personal fable. Propositional Thought: Adolescents do not need to refer real-life conditions and they are capable of evaluating the logic of propositions. Previously, they were evaluating statements when they were concrete in the real world. Piaget did not give an important role for language in childhood period, but he thought that it was more significant during adolescence. So, verbal reasoning about abstract situations and events is one of the important concepts in formal operational thought. Adolescents thoughts and opinions about abstract conditions and their capacity while expressing those views are demonstration of their propositional thought. Theory and research of Piaget have greatly changed the view at infant action and thought and provided a new way of looking at children (Fischer Heneke, 1996). Evaluation of Piagets Theory Jean Piaget has contributed to the field of child development than many other theorists, as many things he expressed were the first for his time. He was the first to tell that children were different from adults and they are curious and active learners (Berk, 2003). Piaget is the founder of discipline of cognitive development. He was interested in reasoning of children and that ensured the idea of that field would be developmental and should be different from the study of adult thinking. Piagets considerations through the idea about active roles of children on their development can be seen as obvious recently, but it was innovative for his time (Shaffer Kipp, 2001). Piaget tried to explain the process development and from that part he was one of the firsts who not only described the theory. Piagets explanations had a major impact about social and emotional developmental theories. By asking many questions about intellectual development of children and creating thousands of resource for that field, he had important contributions for the educators and researchers (Shaffer Kipp, 2001). Many studies criticize Piaget for underestimating mental capabilities of children due to his concern with defining the underlying cognitive structure which considered performance of children on a conservation task. Piaget is criticized as assuming that child making mistake in one problem will lack the underlying concepts he was testing. Validity of this assumption is refuted by stating that performance of a child may be influenced by many other factors instead of lack in cognitive skills (Scholnick et all, 2008). Researchers have challenged stage theory of Piaget by asking if intellectual development is at all stagelike. It is proposed that transitions in mental gradually occur and the consistency of a performance of a child in a test would be very little. For instance, problems of concrete operations and problems of formal operations may be achieved in different orders by many children with different abilities. Many theorists believe that cognitive functions are so complex that would be evaluated within a stage manner. For instance, a nine years old child may do well on verbal reasoning tests if he likes to deal with word puzzles and play verbal games, however he may not do well in mathematical reasoning (Shaffer Kipp, 2001). Besides, Piaget ignores cultural differences while claiming that his stages move on the same sequence in all cultures. Children from different cultures are educated differently and a s environment has a direct interaction with cognitive development, cultural differences should be taken into account to get a valid conclusion. Implications for Education Piaget did influence much from Montessori and Rousseau and emphasized on the importance of active learning of children. He believed that learning is a process of active discovery and should be related with the level of the child. The role of educator at giving the child appropriate and rich environment based on childs interests and modes of learning takes significant place in terms of encouraging innate curiosity of children. An environment with rich, variety, and interesting materials will encourage children to discover and become active learners (Crain, 2005). According to Piaget educators should not teach children in a direct way, children should be allowed to construct their own knowledge through experience. Children should be given the opportunities to make mistakes and learn through those mistakes, and look for solution ways. Piaget helps educators to be aware of that, meaningful interactions of children with environment and real activities will enable children to learn. All of those factors are the roots of constructivism which is a Apoptosis: Techniques for Measuring and Observing Apoptosis: Techniques for Measuring and Observing INTRODUCTION   The number of cells within an organism is tightly regulated- not simply by controlling the rate of cell division, but also by controlling the rate of cell death. If cells are no longer needed, they commit suicide by activating an intracellular death program. (Liao, 2008) Apoptosis, is defined by distinct morphological and biochemical changes mediated by a family of cysteine aspartic acid-specific proteases (caspases), which are expressed as inactive precursors or zymogens (pro-caspases) and are proteolytically processed to an active state following an apoptotic stimulus. (Liao, 2008) The aim of this paper is to discuss the techniques for measuring and observing apoptosis, whilst commenting on any limitations reported to date. DISCUSSION The intracellular machinery responsible for apoptosis Kerr, Wylie and Currie (1972) observed controlled cell death that was distinct from uncontrolled necrotic death. They noticed a characteristic, identical sequence of events in many different types of cells and published their observations in 1972 and used the term apoptosis. (Alberts, 2007; Cancer, 1972) In apoptosis, cell shrinkage and membrane ruffling (blebbing) occur, and the cell disintegrates into small membrane-bound apoptopic bodies. Inside the cell chromatin condensation and nuclear fragmentation occur, which are accompanied by breakdown of the DNA into regular size fragments. On the surface of the cell lipids are rearranged in the bilayer of the plasma membrane with the lipid phosphatidylserine becoming exposed to the outside. (Hancock, 2005) Measuring and observing apoptosis Apoptosis is essentially a morphological state arrived at by a whole variety of different biochemical pathways. Some routes may result in the expression or loss of an antigen but there is no assurance that the same biochemical alterations occur in every cell. The identification of protein markers permits ready detection by conventional immunohistochemistry, which facilitates rapid and confident assessment of apoptosis. (Harrison, 1996) The in situ end labelling and in situ nick translation techniques rely on the presence of DNA strand breaks characteristic of the nuclear implosion and fragmentation seen in apoptosis. Thus, labelling with biotinylated nucleotides and subsequent immunodetection can be used to identity sensitively cells with strand breakage. (Harrison, 1996) Activation of the caspase-3 pathway is a hallmark of apoptosis and can be used in cellular assays to quantify activators and inhibitors of the death cascade by molecular identification. The response is both time and concentration dependent suggesting that multiple pathways play a role in triggering the caspase-3 activation. (Bio Medicine, 2008) TUNEL assay is a technique used for observing activation of the caspase-3 pathway by biochemical verification. The enzyme TdT is able to add nucleotides to the ends of DNA fragments; most commonly, biotin-labelled nucleotides (usually dUTP) are added. The biotinylated DNA can be detected by using streptavidin, which binds to biotin, coupled to enzymes that convert a colourless substrate into a coloured insoluble product. Cells stained in this way can be detected by light microscopy. (Janeway, 2001) The cells with unsuccessful DNA repair may undergo apoptosis; in some cases, false positive labelling may result. TUNEL staining has been reported to give false positive staining in the kidneys of nude and BALB/c mice. In situ localization of activated or cleaved caspase-3 is starting to replace TUNEL as the molecular verification of the apoptotic process. (Hughes, 2007) Extrinsic and Intrinsic signal activation The extrinsic pathway is initiated by ligation of transmembrane death receptors (DR) with their respective to activate membrane-proximal caspases, which in turn cleave and activate effector caspases. A killer lymphocyte carrying the Fas ligand binds and activates Fas proteins on the surface of the target cell. Adaptor proteins bind to the intracellular region of aggregated Fas proteins, causing the aggregation of procaspase-8 molecules. These then cleave one another to initiate the caspase cascade. This pathway can be regulated by c-FLIP, which inhibits upstream initiator caspases, and inhibitor of apoptosis proteins (IAPs), which affect both initiator and effector caspases. (Alberts, 2007; Liao, 2008) The intrinsic pathway requires disruption of the mitochondrial membrane and the release of mitochondrial proteins, such as cytochrome c. Cytochrome c, which binds to and causes the aggregation of the adaptor protein Apaf-1. Apaf-1 binds and aggregates procaspase-9 molecules, which leads to the cleavage of these molecules and the triggering of a caspase cascade. (Alberts, 2007, Liao, 2008) The primary regulatory step for mitochondrial-mediated caspase activation might be at the level of cytochrome c release. The known regulators of cytochrome c release are Bcl-2 family proteins. Members of this family are divided into two main groups, the anti-apoptotic proteins with Bcl-2 and Bcl-Xl as archetypes and the pro-apoptotic proteins such as Bax. The ratio between these different proteins determines the sensitivity of the cell to apoptosis; antiapoptotic proteins inhibit apoptosis by counteracting Bax and Bak, and BH3-only proteins either trigger apoptosis through direct interaction with Bax or sensitize cells to death by inactivating Bcl-2 or Bcl-Xl. (Cartron, 2003; Liao, 2008) RT-PCR primer sets that are specific for genes involved in inducing and regulating the apoptotic response. These primers are specific for genes encoding proteins from the Fas and Fas ligand, Bcl-2 and ICE protein families. The RT-PCR primer sets for studying the apoptotic response are designed to meet several criteria. The primers sets, based on known genomic sequences, amplify a region that spans at least one intron. To distinguish the amplification products from genomic sequences, which are longer than the cDNA products, the primer sets amplify PCR products that are 400 bp to 650 bp in length. The primers are synthesized as 18- to 27-mer oligonucleotides. Each set of primers amplifies only a specific target. (Biomedicine, 2008) Direct measurement using fluorescence resonance energy transfer has shown the interaction between Bax and Bcl-2. (Cartron, 2003) Light microscopy and electron microscopy, including staining, are frequently used to observe the morphological changes of the cells undergoing apoptosis. There are many staining protocols used for identification of apoptotic cells, and the choice varies subject to the laboratory and the tissue being studied. (Wang, 2008) The advantages of Near-Field Scanning Optical Microscope (NSOM) are observing in normal environment, observing in nanometre scale resolution, and observing in non-contact mode. (Wang, 2008) CONCLUSION Using morphology, biochemical or molecular methods to identify, localize and quantify apoptosis gives strength to many research studies. The measurement of the level of apoptosis within tissue sections represents only a à ¢Ã¢â€š ¬Ã‹Å"snapshotà ¢Ã¢â€š ¬Ã¢â€ž ¢ of one time point during a developmental, physiological or pathological process. Given the rapid nature of apoptosis and its cryptic nature in tissue sections, these measurements may often be underestimates of the actual extent of apoptosis. The ability to measure the levels of apoptosis within living organisms, including humans, non-invasively at repeated intervals over time would facilitate the analysis of apoptosis in many organs as well as tumours. (Hughes, 2007) These investigative techniques have broadened the understanding of disease development and will undoubtedly present real opportunities for novel therapeutic intervention. For example, the gene-driven nature of apoptosis and its modulation by various controlling molecules have provided a basis to develop therapies for selectively protecting or deleting cell populations. (Hughes, 2007)